Viruses, Serums, Toxins, and Analogous Products; Organisms and Vectors (9 CFR 101.1-124.43)

666 SUBCHAPTER E—VIRUSES, SERUMS, TOXINS, AND ANALOGOUS PRODUCTS; ORGANISMS AND VECTORS PART 101—DEFINITIONS Sec. 101.1 Applicability. 101.2 Administrative terminology. 101.3 Biological products and related terms. 101.4 Labeling terminology. 101.5 Testing terminology. 101.6 Cell cultures. 101.7 Seed organisms. AUTHORITY: 21 U.S.C. 151–159; 7 CFR 2.22, 2.80, and 371.4. SOURCE: 38 FR 8426, Apr. 2, 1973, unless oth erwise noted. § 101.1 Applicability. When used in parts 101 through 117 of this subchapter, the meaning of the words and phrases listed shall be as de fined in this part. § 101.2 Administrative terminology. The following administrative words and phrases shall mean: Adjacent herd. Adjacent herds are herds physically contiguous to the herd of origin; there are no herds between an adjacent herd and the herd of origin. Administrator. The Administrator, Animal and Plant Health Inspection Service, or any person authorized to act for the Administrator. Animal and Plant Health Inspection Service. The agency in the Department of Agriculture responsible for admin istering the Virus-Serum-Toxin Act. Biological products. The term biologi cal products, also referred to in this subchapter as biologics, biologicals, or products, shall mean all viruses, se rums, toxins (excluding substances that are selectively toxic to microorga nisms, e.g., antibiotics), or analogous products at any stage of production, shipment, distribution, or sale, which are intended for use in the treatment of animals and which act primarily through the direct stimulation, sup plementation, enhancement, or modu lation of the immune system or im mune response. The term ‘‘biological products’’ includes but is not limited to vaccines, bacterins, allergens, anti bodies, antitoxins, toxoids, immunostimulants, certain cytokines, antigenic or immunizing components of live organisms, and diagnostic com ponents, that are of natural or syn thetic origin, or that are derived from synthesizing or altering various sub stances or components of substances such as microorganisms, genes or ge netic sequences, carbohydrates, pro teins, antigens, allergens, or anti bodies. (1) A product’s intended use shall be determined through an objective stand ard and not a subjective one, and would be dependent on factors such as rep resentations, claims (either oral or written), packaging, labeling, or ap pearance. (2) The term analogous products shall include: (i) Substances, at any stage of pro duction, shipment, distribution, or sale, which are intended for use in the treatment of animals and which are similar in function to biological prod ucts in that they act, or are intended to act, through the stimulation, sup plementation, enhancement, or modu lation of the immune system or im mune response; or (ii) Substances, at any stage of pro duction, shipment, distribution, or sale, which are intended for use in the treatment of animals through the de tection or measurement of antigens, antibodies, nucleic acids, or immunity; or (iii) Substances, at any stage of pro duction, shipment, distribution, or sale, which resemble or are represented as biological products intended for use in the treatment of animals through appearance, packaging, labeling, claims (either oral or written), rep resentations, or through any other means. (3) The term treatment shall mean the prevention, diagnosis, management, or cure of diseases of animals. Department. The U.S. Department of Agriculture. Distributor. A person who sells, dis tributes, or otherwise places in chan nels of trade, one or more biological VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00676 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 667 Animal and Plant Health Inspection Service, USDA § 101.2 products he does not produce or im port. Division. A marketing unit estab lished by the licensee which may be named on labels, advertisements and promotional material in addition to the name and address of the producer. Domestic animals. All animals, other than man, including poultry. Establishment. One or more premises designated on the establishment li cense. Guidelines. Guidelines establish prin ciples or practices related to test pro cedures, manufacturing practices, product standards, scientific protocols, labeling, and other technical or policy considerations. Guidelines contain pro cedures or standards of general applica bility that are usually not regulatory in nature, but that are related to mat ters that fall under the Virus-Serum Toxin Act. Guidelines issued by the agency include Veterinary Biologics Licensing Considerations, Memoranda, Notices, and Supplemental Assay Methods. Herd. Any group of animals, includ ing birds, fish, and reptiles, maintained at a common location (e.g. lot, farm or ranch) for any purpose. The herd (or flock) includes all animals subse quently housed at the common loca tion. If the principal animals of a group are moved to a different location, the group is still considered the same herd. Herd of origin. The herd from which the microorganism used as seed for production of an autogenous biologic is isolated. Offspring and excess breeding stock (not the principal animals) moved or sold from one group of ani mals to another have changed herds and are no longer considered part of the herd they originated from. Groups of animals under the same ownership but at different locations are separate herds. Inspection. An examination made by an inspector to determine the fitness of animals, establishments, facilities, and procedures used in connection with the preparation, testing, and distribution of biological products and the examina tion or testing of biological products. Inspector. Any officer or employee of Animal and Plant Health Inspection Service who is authorized by the Ad ministrator to do inspection work. Licensed establishment. An establish ment operated by a person holding an unexpired, unsuspended, and unrevoked U.S. Veterinary Biologics Establish ment License. Licensee. A person to whom an estab lishment license and at least one prod uct license has been issued. Microorganisms. Microscopic or sub microscopic organisms, which are sometimes referred to as organisms, which may introduce or disseminate disease of animals. Nonadjacent herd. Nonadjacent herds are all herds other than the herd of ori gin and other than herds adjacent to the herd of origin. Herds adjacent to the herd of origin but in a different State from the herd of origin are also considered nonadjacent herds. Permittee. A person who resides in the United States or operates a business establishment within the United States, to whom a permit to import bi ological products has been issued. Person. Any individual, firm, partner ship, corporation, company, associa tion, educational institution, State or local governmental agency, or other organized group of any of the fore going, or any agent, officer, or em ployee of any thereof. Premises. All buildings, appur tenances, and equipment used to produce and store biological products located within a particular land area shown on building plans or drawings furnished by the applicant or the li censee and designated by an address adequate for identification. Prepare or preparation. Sometimes re ferred to as manufacture or produce, means the steps and procedures used in the processing, testing, packaging, la beling, and storing of a biological prod uct. Regulations. The provisions in parts 101 through 118 of this subchapter. Research investigator or research spon sor. A person who has requested au thorization to ship an experimental bi ological product for the purpose of evaluating such product, or has been granted such authorization. Secretary. The Secretary of Agri culture of the United States or any of ficer or employee of the Department to whom authority has heretofore been delegated, or to whom authority may VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00677 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 668 9 CFR Ch. I (1–1–18 Edition) § 101.3 hereafter be delegated, to act in his stead. Subsidiary. A corporation in which a corporate licensee owns in excess of 50 percent of the voting stock. Veterinary Services. Veterinary Serv ices unit of Animal and Plant Health Inspection Service of the Department. Virus-Serum-Toxin Act. The Act of March 4, 1913, 37 Stat. 832–833; as amended December 23, 1985, Public Law 99–198, 99 Stat. 1654–1655; and as further amended September 28, 1988, Public Law 100–449, 102 Stat. 1868; 21 U.S.C. 151–159. U.S. Veterinary Biological Product Li cense. A document, sometimes referred to as a product license, which is issued pursuant to part 102 of this subchapter to the holder of an establishment li cense, as a part of and ancillary to the establishment license, and which au thorizes production of a specified bio logical product in the designated li censed establishment. U.S. Veterinary Biological Product Per mit. A document, sometimes referred to as a permit, issued to a person author izing the importation of specified bio logical products subject to restrictions and controls as provided in the regula tions. U.S. Veterinary Biologics Establishment License. A document referred to as an establishment license, which is issued pursuant to part 102 of this subchapter, authorizing the use of designated prem ises for production of biological prod ucts specified in one or more unex pired, unsuspended, and unrevoked product license(s). [38 FR 8426, Apr. 2, 1973; 38 FR 9221, Apr. 12, 1973, as amended at 40 FR 46093, Oct. 6, 1975; 41 FR 44358, Oct. 8, 1976; 49 FR 22624, May 31, 1984; 52 FR 30131, Aug. 13, 1987; 56 FR 66782, 66783, Dec. 26, 1991; 57 FR 38756, Aug. 27, 1992; 62 FR 31328, June 9, 1997; 64 FR 43044, Aug. 9, 1999] § 101.3 Biological products and related terms. When used in conjunction with or in reference to a biological product, the following terms shall mean: (a) Licensed biological product. A bio logical product prepared within a li censed establishment by a person hold ing an unexpired, unsuspended, and unrevoked product license for such product. (b) Experimental biological product. A biological product which is being eval uated to substantiate an application for a product license or permit. (c) Completed product. A biological product in bulk or final container pro duced in compliance with the regula tions to final form and composition. (d) Finished product. A completed product which has been bottled, sealed, packaged, and labeled as required by the regulations. (e) Released product. A finished prod uct released for marketing after all re quirements have been satisfactorily complied with. (f) Fraction. A specific antigen, its antibodies, or its antitoxin which con stitutes a component of a biological product. (g) Diluent. A liquid used to re hydrate a desiccated product or a liq uid used to dilute another substance. (h) Serial. The total quantity of com pleted product which has been thor oughly mixed in a single container and identified by a serial number: Provided, That, when all or part of a serial of liq uid biological product is packaged as diluent for all or part of a serial of des iccated product, the resulting combina tion packages shall be considered a se rial of the multiple fraction product. (i) Subserial. Each of two or more properly identified portions of a serial which are further processed at different times or under different conditions such as, but not limited to, being des iccated in different size final con tainers and/or at different times. (j) Outline of production. A detailed protocol of methods of manufacture to be followed in the preparation of a bio logical product and which may some times be referred to as an outline. (k) Product Code Number. A number assigned by Animal and Plant Health Inspection Service to each type of li censed biological product. (l) Harvest date. Unless otherwise specified in a filed Outline of Produc tion, the harvest date shall be the date blood or tissues are collected for pro duction or the date cultures of living microorganisms are removed from pro duction incubators. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00678 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 669 Animal and Plant Health Inspection Service, USDA § 101.4 (m) Bacterin. An inactivated bacterial product consisting of an antigenic sus pension of organisms or particulate parts of organisms, representing a whole culture or a concentrate thereof, with or without the unevaluated growth products, which has been inac tivated as demonstrated by acceptable tests written into the filed Outline of Production for the product. (n) Toxoid. An inactivated bacterial product which consists of a sterile, an tigenic toxin or toxic growth product, which has resulted from the growth of bacterial organisms in a culture me dium from which the bacterial cells have been removed, which has been in activated without appreciable loss of antigenicity as measured by suitable tests, and which is nontoxic as dem onstrated by acceptable tests written into the filed Outline of Production. (o) Bacterin-toxoid. An inactivated bacterial product which is either: (1) A suspension of organisms, rep resenting a whole culture or a con centrate thereof, with the toxic growth products from the culture which has been inactivated without appreciable loss of antigenicity as measured by suitable tests, the inactivation of orga nisms and toxins being demonstrated by acceptable tests written into the filed Outline of Production: Provided, That it shall contain cellular antigens and shall stimulate the development of antitoxin; or (2) A combination product in which one or more toxoids or bacterin-toxoids is combined with one or more bacterins or one or more bacterin-toxoids. (p) Bacterial extract. An inactivated bacterial product which consists of the sterile, nontoxic, antigenic derivatives extracted from bacterial organisms or from culture medium in which bac terial organisms have grown. (q) Combination package. Biological product consisting of two or more li censed biological products. Each com pleted product in final container is packaged together and mixed prior to administration. A combination pack age is issued a separate U.S. Veteri nary Biological Product License and assigned a product code number to dis tinguish it from its component prod ucts, which also may be marketed indi vidually unless otherwise restricted. [38 FR 8426, Apr. 2, 1973, as amended at 42 FR 63770, Dec. 20, 1977; 50 FR 24903, June 14, 1985; 56 FR 66782, Dec. 26, 1991; 60 FR 14354, Mar. 17, 1995; 81 FR 59433, Aug. 30, 2016] § 101.4 Labeling terminology. Terms pertaining to identification and packaging of biological products shall mean: (a) Label. All written, graphic, or printed matter: (1) Upon or attached to a final con tainer of a biological product; (2) Appearing upon any immediate carton or box used to package such final container; and (3) Appearing on any accompanying enclosures (leaflets, inserts, or circu lars) on which required information or directions as to the use of the biologi cal product shall be found. (b) Labeling. All labels and other written, printed, or graphic matter ac companying the final container. (c) Final container. The unit, bottle, vial, ampule, tube, or other receptacle into which any biological product is filled for distribution and sale. (d) True name. The name entered on the product license or permit at the time of issuance to differentiate the bi ological product from others: Provided, That, the principal part of such name shall be emphasized on such license or permit by being more prominently let tered than descriptive terms which may be necessary to complete the dif ferentiation. (e) Serial number. Numbers or num bers and letters used to identify and distinguish one serial from others. (f) Expiration date. A date designating the end of the period during which a bi ological product, when properly stored and handled, can be expected with rea sonable certainty, to be efficacious. (g) Label number. A number assigned by Animal and Plant Health Inspection Service to each label or sketch sub mitted for review. (h) Master label. The finished carton, container, or enclosure label for the smallest size final container that is au thorized for a biological product, that serves as the Master template label ap plicable to all other size containers or cartons of the same product that is VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00679 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 670 9 CFR Ch. I (1–1–18 Edition) § 101.5 marketed by a licensee, subsidiary, di vision, or distributor. [38 FR 8426, Apr. 2, 1973, as amended at 42 FR 63770, Dec. 20, 1977; 56 FR 66782, Dec. 26, 1991; 61 FR 29464, June 11, 1996] § 101.5 Testing terminology. Terms used when evaluating biologi cal products shall mean: (a) Standard Requirement. Test meth ods, procedures, and criteria estab lished by Animal and Plant Health In spection Service for evaluating biologi cal products to be pure, safe, potent, and efficacious, and not to be worth less, contaminated, dangerous, or harmful under the Act. (b) Log. Logarithm computed to the base 10. (c) Pure or purity. Quality of a bio logical product prepared to a final form relatively free of extraneous micro-or ganisms and extraneous material (or ganic or inorganic) as determined by test methods or procedures established by Animal and Plant Health Inspection Service in Standard Requirements or in the approved Outline of Production for such product, but free of extraneous microorganisms or material which in the opinion of the Administrator ad versely affects the safety, potency, or efficacy of such product. (d) Safe or safety. Freedom from prop erties causing undue local or systemic reactions when used as recommended or suggested by the manufacturer. (e) Sterile or sterility. Freedom from viable contaminating microorganisms as demonstrated by procedures pre scribed in part 113 of this subchapter, Standard Requirements, and approved Outlines of Production. (f) Potent or potency. Relative strength of a biological product as de termined by test methods or proce dures as established by Animal and Plant Health Inspection Service in Standard Requirements or in the ap proved Outline of Production for such product. (g) Efficacious or efficacy. Specific ability or capacity of the biological product to effect the result for which it is offered when used under the condi tions recommended by the manufac turer. (h) Dose. The amount of a biological product recommended on the label to be given to one animal at one time. (i) Vaccinate. An animal which has been inoculated, injected, or otherwise administered a biological product being evaluated. (j) Control animal. An animal, which may be referred to as a control, used in a test procedure for purposes of com parison or to add validity to the re sults. (k) Day. Time elapsing between any regular working hour of one day and any regular working hour of the fol lowing day. (l) Test results. Terms used to des ignate testing results are as follows: (1) No Test. Designation used when a deficiency in the test system has ren dered a test unsuitable for drawing a valid conclusion. (2) Satisfactory. Designation is a final conclusion given to a valid test with results that meet the release criteria stated in the filed Outline of Produc tion or Standard Requirement. (3) Unsatisfactory. Designation is a final conclusion given to a valid test with results that do not meet the re lease criteria stated in the filed Out line of Production or Standard Re quirement. (4) Inconclusive. Designation used for an initial test when a sequential test design established in the filed Outline of Production or Standard Require ment allows further testing if a valid initial test is not satisfactory. (m) Healthy. Apparently normal in all vital functions and free of signs of dis ease. (n) Unfavorable reactions. Overt ad verse changes which occur in healthy test animals subsequent to initiation of a test and manifested during the ob servation period prescribed in the test protocol which are attributable either to the biological product being tested or to factors unrelated to such product as determined by the responsible indi vidual conducting the test. (o) Master reference. A Master Ref erence is a reference whose potency is correlated, directly or indirectly, to host animal immunogenicity. The Mas ter Reference may be used as the work ing reference in in vitro tests for rel ative potency. The Master Reference VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00680 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 671 Animal and Plant Health Inspection Service, USDA § 101.7 may also be used to establish the rel ative potency of a serial of product used in requalification studies and to establish the relative potency of work ing references. The preparation of a Master Reference as described in a filed Outline of Production may be: (1) A completed serial of vaccine or bacterin prepared in accordance with a filed Outline of Production; (2) A purified preparation of a protec tive immunogen or antigen; or (3) A nonadjuvanted harvested cul ture of microorganisms. (p) Working reference. A Working Ref erence is the reference preparation that is used in the in vitro test for the release of serials of product. Working References may be: (1) Master References; or (2) Serials of product that have been prepared and qualified, in a manner ac ceptable to Animal and Plant Health Inspection Service for use as reference preparations. (q) Qualifying serial. (1) A serial of bi ological product used to test for immunogenicity when the Master or Working Reference is a purified anti gen or nonadjuvanted harvest material. Qualifying serials shall be produced in accordance with the filed Outline of Production, tested for immunogenicity in accordance with methods deemed appropriate by the Animal and Plant Health Inspection Service, and have a geometric mean relative potency, when compared to the Master Reference, of not greater than 1.0 as established by: independent parallel line assays with five or more replicates; or other valid assay methods for determining relative antigen content which demonstrate linearity, specificity, and reproduc ibility at least equivalent to the par allel line assay and are acceptable to the Animal and Plant Health Inspec tion Service. (2) Qualifying serials used to re qualify or extend the dating period of a Master Reference shall be determined to be immunogenic in accordance with methods deemed appropriate by the Animal and Plant Health Inspection Service as provided in paragraph (a)(1) of this section, and, in addition, shall be within their permitted dating period and have been prepared in accordance with the production method described in the currently filed Outline of Pro duction. (r) Immunogenicity. The ability of a biological product to elicit an immune response in animals as determined by test methods or procedures acceptable to the Animal and Plant Health Inspec tion Service. [38 FR 8426, Apr. 2, 1973, as amended at 40 FR 45419, Oct. 2, 1975; 41 FR 6751, Feb. 13, 1976; 43 FR 3701, Jan. 27, 1978; 56 FR 66782, 66783 Dec. 26, 1991; 62 FR 19037, Apr. 18, 1997; 79 FR 55969, Sept. 18, 2014] § 101.6 Cell cultures. When used in conjunction with or in reference to cell cultures, which may be referred to as tissue cultures, the following terms shall mean: (a) Batches of primary cells. A pool of original cells derived from normal tis sue up to and including the 10th sub culture. (b) Cell line. A pool of cells which are 11 or more subcultures from the tissue of origin. (c) Subculture. Each flask to flask transfer or passage regardless of the number of cell replications. (d) Master Cell Stock (MCS). The sup ply of cells of a specific passage level from which cells for production of bio logics originate. [38 FR 8426, Apr. 2, 1973, as amended at 40 FR 45419, Oct. 2, 1975; 49 FR 22624, May 31, 1984] § 101.7 Seed organisms. When used in conjunction with or in reference to seed organisms, the fol lowing shall mean: (a) Master Seed. An organism at a spe cific passage level which has been se lected and permanently stored by the producer from which all other seed pas sages are derived within permitted lev els. (b) Working Seed. An organism at a passage level between Master Seed and Production Seed. (c) Production Seed. An organism at a specified passage level which is used without further propagation for initi ating preparation of a fraction. [49 FR 22625, May 31, 1984] VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00681 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 672 9 CFR Ch. I (1–1–18 Edition) Pt. 102 PART 102—LICENSES FOR BIOLOGICAL PRODUCTS Sec. 102.1 Licenses issued by the Administrator. 102.2 Licenses required. 102.3 License applications. 102.4 U.S. Veterinary Biologics Establish ment License. 102.5 U.S. Veterinary Biological Product Li cense. 102.6 Conditional licenses. AUTHORITY: 21 U.S.C. 151–159; 7 CFR 2.22, 2.80, and 371.4. § 102.1 Licenses issued by the Adminis trator. Each establishment qualified to pre pare biological products under the Virus-Serum-Toxin Act shall hold an unexpired and unrevoked U.S. Veteri nary Biologics Establishment License issued by the Administrator and a U.S. Veterinary Biological Product License for each product prepared in such es tablishment unless the product is sub ject to the provisions of 9 CFR parts 103 or 106 of this subchapter. [60 FR 48021, Sept. 18, 1995] § 102.2 Licenses required. (a) Every person who prepares bio logical products subject to the Virus Serum-Toxin Act shall hold an unex pired, unsuspended, and unrevoked U.S. Veterinary Biologics Establishment Li cense and at least one unexpired, un suspended, and unrevoked U.S. Veteri nary Biological Product License issued by the Administrator to prepare a bio logical product. (b) An applicant who applies for an establishment license must also apply for at least one product license. An es tablishment license will not be issued without a license authorizing the pro duction of a biological product in the establishment. [52 FR 11026, Apr. 7, 1987, as amended at 56 FR 66783, Dec. 26, 1991; 61 FR 52873, Oct. 9, 1996] § 102.3 License applications. (a) U.S. Veterinary Biologics Establish ment License. (1) The operator of each establishment of the kind specified in § 102.2 shall make written application to the Administrator for a license. Blank forms of application will be fur nished upon request to Animal and Plant Health Inspection Service. (2) When a person conducts more than one establishment, a separate ap plication shall be made for each estab lishment. (3) Whenever subsidiaries are to oper ate in an establishment for which li cense application is made, the appli cant shall apply for permission for such subsidiaries to operate in the establish ment and furnish therewith a complete statement regarding the relationship between the applicant and the subsidi aries. (4) Facilities documents, prepared as prescribed in part 108 of this sub chapter, shall accompany the applica tion for license unless previously filed with Animal and Plant Health Inspec tion Service. (5) Each application for a U.S. Veteri nary Biologics Establishment License shall be accompanied by an application for one or more U.S. Veterinary Bio logical Product Licenses and the sup porting documents required by para graph (b)(2) of this section. (6) A new application shall be made when a change of ownership, operation, or location of an establishment occurs; or prior to the expiration of a U.S. Vet erinary Biologics Establishment Li cense issued for an interim period of time. (b) U.S. Veterinary Biological Product License. (1) The licensee of each estab lishment or applicant for an establish ment license shall make written appli cation to the Administrator for a U.S. Veterinary Biological Product License for each biological product to be pre pared in the licensed establishment. (2) Each application for a U.S. Veteri nary Biological Product License shall be supported by: (i) At least two copies of an Outline of Production prepared in accordance with §§ 114.8 and 114.9 of this sub chapter; and (ii) At least three copies of test re ports and research data sufficient to establish purity, safety, potency, and efficacy of the product; and (iii) Legends prepared as prescribed in § 108.5 of this subchapter designating which facilities are to be used in the preparation of each fraction; and VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00682 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 673 Animal and Plant Health Inspection Service, USDA § 102.4 (iv) Labels in finished form or sketches prepared as prescribed in § 112.5 of this subchapter, together with information regarding all claims to be made on labels and in advertising mat ter to be used in connection with or re lated to the biological product. (Approved by the Office of Management and Budget under control number 0579–0013) [39 FR 37763, Oct. 24, 1974, as amended at 48 FR 57472, Dec. 30, 1983; 49 FR 21043, May 18, 1984; 50 FR 50763, Dec. 12, 1985; 56 FR 66783, Dec. 26, 1991; 75 FR 20772, Apr. 21, 2010] § 102.4 U.S. Veterinary Biologics Estab lishment License. (a) Before a U.S. Veterinary Biologics Establishment License will be issued by the Administrator for any establish ment, an inspection shall be made to determine whether the condition, equipment, facilities, and the like, of the establishment, and the methods used to prepare biological products are in conformity with the requirements in the regulations. (b) A license shall not be issued un less: (1) In the opinion of the Adminis trator, the condition of the establish ment, including its facilities, and the methods of preparation of biological products are such as reasonably to as sure that the products shall accomplish the purpose for which they are in tended; and (2) The Administrator is satisfied on the basis of information before him that: (i) The establishment shall be oper ated in compliance with the Act and applicable regulations and be under the supervision of person(s) competent in the preparation of biological products; and (ii) The applicant, or the person hav ing the responsibility for producing bi ological products in the establishment, or both, is qualified by education and experience, and has demonstrated fit ness to produce such products in com pliance with the Act and regulations issued pursuant thereto; Provided, That, previous violations of the Act, or such regulations or both shall be rel evant to the Administrator’s deter mination of fitness. (3) Written assurance is filed with Animal and Plant Health Inspection Service that the biological products which are licensed to be prepared therein shall not be so advertised as to mislead or deceive the purchasers and that the packages or containers in which the same are to be marketed shall not bear any statement, design, or device which is false or misleading in any particular. (c) U.S. Veterinary Biologics Estab lishment Licenses shall be numbered. (d) Two or more licenses may bear the same number when they are issued for establishments under the same ownership or control, provided a serial letter is added to one or more to iden tify each license and the product pro duced thereunder. (e) When a U.S. Veterinary Biologics Establishment License is issued for an establishment, it shall not apply to more than one person at the same loca tion, except that subsidiaries of the li censee, when named in the license, may operate thereunder at the establish ment named. The licensee with its sub sidiaries will be held responsible for all operations conducted in the licensed establishment. (f) When a licensee no longer holds at least one unexpired, unsuspended, or unrevoked product license authorizing the preparation of a biological product, or is in the process of obtaining a prod uct license, the establishment license shall no longer be valid and shall be re turned to the Administrator. In the case where an establishment license expires or is suspended or revoked, any product license authorizing prepara tion of a product at such establishment shall be invalid indefinitely or for as long as the suspension is in effect. (g) Any license issued under this part to establishments in which biological products are prepared shall be issued on condition that the licensee permit the inspection of such establishments, products, product preparation, and all relevant records as provided in part 115 of this subchapter. Failure to permit inspection may result in the license being suspended or revoked. (h) The provisions of paragraph (b) of this section shall also be applicable to, VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00683 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 674 9 CFR Ch. I (1–1–18 Edition) § 102.5 and be considered by, the Adminis trator in connection with each applica tion for an additional product license. (Approved by the Office of Management and Budget under control number 0579–0013) [39 FR 37762, Oct. 24, 1974; 39 FR 38364, Nov. 1, 1974, as amended at 41 FR 44359, Oct. 8, 1976; 48 FR 57472, Dec. 30, 1983; 52 FR 11026, Apr. 7, 1987; 52 FR 30131, Aug. 13, 1987; 56 FR 66783, Dec. 26, 1991; 60 FR 48021, Sept. 18, 1995; 61 FR 52873, Oct. 9, 1996; 62 FR 13294, Mar. 20, 1997] § 102.5 U.S. Veterinary Biological Product License. (a) Authorization to produce each bi ological product shall be specified on a U.S. Veterinary Biological Product Li cense, issued by the Administrator, and supplementary to the U.S. Veterinary Biologics Establishment License named therein. (b) The following shall appear on the U.S. Veterinary Biological Product Li cense: (1) The U.S. Veterinary Biologics Es tablishment License Number for the es tablishment from which the product is released for marketing. (2) The true name of the product. (3) The product code number for the product. (4) The date of issuance. (5) Any restrictions designated by the Administrator under paragraph (e) of this section. (6) When necessary to comply with § 102.6 of this part, a termination date and a brief description of requirements to be met for reissuance. (c) The following provisions shall apply to all licensed biological prod ucts: (1) Licensed biological products shall be prepared as required by the regula tions and in accordance with a filed Outline of Production as prescribed in §§ 114.8 and 114.9 of this subchapter. No change shall be made in the prepara tion of a biological product without prior approval of the Administrator. (2) In addition to restrictions im posed by the Administrator pursuant to paragraph (e) of this section, bio logical products may be subject to re strictions which are imposed by any State or other jurisdiction pertaining to the distribution and use of such products, based on local disease condi tions. (3) When requested by the Adminis trator, a licensee shall submit a list of licensed biological products prepared in the licensed establishment. (d) Where the Administrator deter mines that the protection of domestic animals or the public health, interest, or safety, or both, necessitates restric tions on the use of a product, the prod uct shall be subject to such additional restrictions as are prescribed on the li cense. Such restrictions may include, but are not limited to, limits on dis tribution of the product or provisions that the biological product is re stricted to use by veterinarians, or under the supervision of veterinarians, or both. (e) Any person may request that the distribution and use of a veterinary bi ological product be restricted if the re striction pertains to the protection of domestic animals or the public health, interest, or safety, or both. All re quests must be sent, in writing, to the Director, Center for Veterinary Bio logics, Policy, Evaluation, and Licens ing, 1920 Dayton Avenue, P.O. Box 844, Ames, IA 50010. Requests must specify the restriction(s) being requested and must explain why the restrictions are needed. Copies of any supporting docu ments, such as scientific literature, published or unpublished articles, or data from tests, should be attached to the request. When a decision is reached regarding the request, the person sub mitting the request will be sent writ ten notification of such decision. (Approved by the Office of Management and Budget under control number 0579–0013) [39 FR 37763, Oct. 24, 1974, as amended at 48 FR 57472, Dec. 30, 1983; 50 FR 50764, Dec. 12, 1985; 52 FR 11026, Apr. 7, 1987; 56 FR 66783, Dec. 26, 1991; 57 FR 38760, Aug. 27, 1992; 59 FR 67616, Dec. 30, 1994; 62 FR 13294, Mar. 20, 1997; 64 FR 43044, Aug. 9, 1999; 75 FR 20772, Apr. 21, 2010] § 102.6 Conditional licenses. In order to meet an emergency condi tion, limited market, local situation, or other special circumstance, includ ing production solely for intrastate use under a State-operated program, the Administrator may, in response to an application submitted as specified in VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00684 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 675 Animal and Plant Health Inspection Service, USDA § 103.2 § 102.3(b) of this part, issue a condi tional U.S. Veterinary Biological Prod uct License to an establishment under an expedited procedure which assures purity and safety, and a reasonable ex pectation of efficacy. Preparation of products under a conditional license shall be in compliance with all applica ble regulations and standards and may be restricted as follows: (a) The preparation may be limited to a predetermined time period which shall be established at the time of issuance and specified on the license. Prior to termination of the license, the licensee may request reissuance. Such requests shall be substantiated with data and information obtained since the license was issued. After consid ering all data and information avail able, the Administrator shall either re issue the U.S. Veterinary Biological Product License or allow it to termi nate. (b) Distribution may be limited to the extent necessary to assure that the product will meet the basic criteria for issuance of the conditional license. (c) Labeling for the product may be required to contain information on the conditional status of the license. [52 FR 11026, Apr. 7, 1987, as amended at 60 FR 48021; Sept. 18, 1995] PART 103—EXPERIMENTAL PRO DUCTION, DISTRIBUTION, AND EVALUATION OF BIOLOGICAL PRODUCTS PRIOR TO LICENSING Sec. 103.1 Preparation of experimental biological products. 103.2 Disposition of animals administered experimental biological products or live organisms. 103.3 Shipment of experimental biological products. AUTHORITY: 21 U.S.C. 151–159; 7 CFR 2.22, 2.80, and 371.4. § 103.1 Preparation of experimental bi ological products. Except as otherwise provided in this section, experimental biological prod ucts which are neither composed of nor prepared with organisms or antigens used in biologicals already licensed, shall not be prepared in the production facilities of a licensed establishment. Upon application therefor, the Admin istrator may authorize the preparation of experimental products on the prem ises of a licensed establishment if he determines that such preparation will not result in contamination of the li censed products. Each request for per mission to prepare an experimental bi ological product on licensed premises shall indicate the nature of the unli censed product, designate facilities to be used, and specify precautions which will be taken to prevent contamination of licensed products. Such requests shall be submitted to the Adminis trator. Research facilities that are en tirely separate and apart from facili ties used for the preparation of licensed biological products will not be consid ered a part of the licensed premises for purposes of this section. (Approved by the Office of Management and Budget under control number 0579–0013) [30 FR 11848, Sept. 16, 1965, as amended at 48 FR 57473, Dec. 30, 1983; 56 FR 66783, Dec. 26, 1991] § 103.2 Disposition of animals adminis tered experimental biological prod ucts or live organisms. Safeguards as herein provided shall be established by the research investi gator or research sponsor to control disposition of all animals administered experimental biological products or live organisms. (a) Surviving test animals (including challenged control animals) shall not be removed from the premises on which the tests are conducted for at least 14 days after administration of an experi mental biological product or live orga nisms: Provided, however, That this holding period may be increased or de creased as permitted or requested by the Administrator following review of all relevant information or data avail able. (b) All animals administered experi mental biological products which are to be slaughtered at establishments subject to the Federal Meat Inspection Act, as amended and extended (21 U.S.C. 601 et. seq.) are subject to the ap plicable requirements of § 309.16 of this title (Meat Inspection Regulations). VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00685 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 676 9 CFR Ch. I (1–1–18 Edition) § 103.3 (c) Except as otherwise provided in this paragraph, the research investi gator or research sponsor shall main tain adequate records relative to the disposition of each animal adminis tered experimental biological products. These records shall be maintained for a minimum period of two years from the date that an experimental product was administered to such animal, and shall show the name and address of the owner; number, species, class and loca tion of the animals; and if sold, the name and address of the consignee, buyer, commission, firm or abattoir: Provided, however, That a research in vestigator or research sponsor may be exempted from these recordkeeping re quirements by the Administrator on the basis of acceptable data dem onstrating that use of the experimental biological product will not result in the presence of any unwholesome condition in the edible parts of animals subse quently presented for slaughter. (Approved by the Office of Management and Budget under control number 0579–0059) [30 FR 11848, Sept. 16, 1965, as amended at 48 FR 57473, Dec. 30, 1983; 56 FR 66783, Dec. 26, 1991; 66 FR 21063, Apr. 27, 2001] § 103.3 Shipment of experimental bio logical products. Except as provided in this section, no person shall ship or deliver for ship ment in or from the United States, the District of Columbia, or any Territory of the United States any unlicensed bi ological product for experimental use in animals. For the benefit of license applicants and to permit and encourage research, a person may be authorized by the Administrator to ship unli censed biological products for the pur pose of evaluating such experimental products by treating limited numbers of animals, Provided, that, the Admin istrator determines that the conditions under which the experiment is to be conducted are adequate to prevent the spread of disease and approves the pro cedures set forth in the request for such authorization. Special restric tions or tests may be imposed, espe cially in the case of products con taining live organisms, when they are deemed necessary or advisable by the Administrator. A request for author ization to ship an unlicensed biological product for experimental study and evaluation shall be accompanied by the following: (a) One copy of a permit or letter of permission from the proper State or foreign animal health authorities of each State or foreign country involved. (b) Two copies of a tentative list of the names of the proposed recipients and quantity of experimental product that is to be shipped to each individual. In the event of subsequent changes, ad ditional information shall be furnished when such facts are known; (c) Two copies of a description of the product, recommendations for use, and results of preliminary research work; (d) A copy of the labels or label sketches which show the name or iden tification of the product and bear the statement ‘‘Notice! For experimental use only-Not For Sale’’ or equivalent. Such statement shall appear on final container labels, except that it may ap pear on the carton in the case of very small final container labels and label ing for diagnostic test kits. The U.S. Veterinary License legend shall not ap pear on such labels; and (e) Two copies of a proposed general plan covering the methods and proce dures for evaluating the product and for maintaining records of the quan tities of experimental product pre pared, shipped and used. At the conclu sion of field studies, results shall be ob tained, summarized, and submitted to the Animal and Plant Health Inspec tion Service. (f) Data acceptable to the Adminis trator demonstrating that use of the experimental biological product in meat animals is not likely to result in the presence of any unwholesome con dition in the edible parts of animals subsequently presented for slaughter. (g) A statement from the research in vestigator or research sponsor agreeing to furnish, upon the Administrator’s request, additional information con cerning each group of meat animals in volved prior to movement of these ani mals from the premises where the test is to be conducted. Such information shall include the owner’s name and ad dress; number, species, class and loca tion of animals involved; date ship ment is anticipated; along with name VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00686 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 677 Animal and Plant Health Inspection Service, USDA § 104.3 and address of consignee, buyer, com mission firm or abattoir. (h) Any information the Adminis trator may require in order to assess the product’s impact on the environ ment. [26 FR 7726, Aug. 18, 1961, as amended at 30 FR 11848, Sept. 16, 1965; 52 FR 30131, Aug. 13, 1987; 56 FR 66783, Dec. 26, 1991; 75 FR 20772, Apr. 21, 2010; 81 FR 59433, Aug. 30, 2016] PART 104—PERMITS FOR BIOLOGICAL PRODUCTS Sec. 104.1 Permit required. 104.2 Permit authorized. 104.3 Permit application. 104.4 Products for research and evaluation. 104.5 Products for distribution and sale. 104.6 Products for transit shipment only. 104.7 Product permit. 104.8 Illegal shipments. AUTHORITY: 21 U.S.C. 151–159; 7 CFR 2.22, 2.80, and 371.4. SOURCE: 38 FR 32916, Nov. 29, 1973, unless otherwise noted. § 104.1 Permit required. Unless otherwise authorized or di rected by the Administrator, each per mit to import a biological product into the United States shall be issued in ac cordance with the regulations in this part. (a) No biological product shall be brought into the United States unless a permit has been issued for such prod uct. A separate U.S. Veterinary Bio logical Product Permit shall be re quired for each shipment of biological product to be imported: Provided, That, a permit shall also be required for each transit shipment of biological products moved through the United States. (b) Each person importing biological products shall hold an unexpired, un suspended, and unrevoked permit issued by Animal and Plant Health In spection Service. Such person shall re side within the United States, or oper ate a business establishment within the United States, or both. [38 FR 32916, Nov. 29, 1973, as amended at 56 FR 66783, Dec. 26, 1991; 56 FR 66783, Dec. 26, 1991] § 104.2 Permit authorized. (a) Animal and Plant Health Inspec tion Service is authorized to issue three types of permits for importing bi ological products. They shall be: (1) U.S. Veterinary Biological Prod uct Permit for Research and Evalua tion; (2) U.S. Veterinary Biological Prod uct Permit for Distribution and Sale; or (3) U.S. Veterinary Biological Prod uct Permit for Transit Shipment Only. (b) A permit shall not be issued for a biological product from countries known to have exotic diseases, includ ing but not limited to foot-and-mouth disease, rinderpest, highly pathogenic avian influenzaswine vesicular disease, Newcastle disease, and African swine fever, if in the opinion of the Adminis trator, such products may endanger the livestock or poultry of this coun try. (c) A permit shall not be issued until an inspector has determined the condi tion of the equipment and facilities of the producer, of the applicant, or of both if such a determination is consid ered necessary by the Administrator. (d) A permit shall not be issued for a biological product prepared in the United States, exported, and presented for reentry except as provided in § 104.4(d). [38 FR 32916, Nov. 29, 1973, as amended at 56 FR 66783, Dec. 26, 1991; 56 FR 66783, Dec. 26, 1991; 78 FR 19085, Mar. 29, 2013] § 104.3 Permit application. (a) Each person desiring to import a biological product shall make written application to Animal and Plant Health Inspection Service for a permit. Application forms are available on the Internet at (http://www.aphis.usda.gov/ animallhealth/ vetlbiologics/ vblforms.shtml) and application for a permit to import a veterinary biologic for research and evaluation or transit shipment may be made on the Internet at (http://www.aphis.usda.gov/ animallhealth/permits/ vetlbiolpermits.shtml). (b) The application shall specify the type of permit required, the port of entry at which the product shall be VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00687 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 678 9 CFR Ch. I (1–1–18 Edition) § 104.4 cleared through Customs, the esti mated quantity involved, and the an ticipated date on which the importa tion shall be made. (Approved by the Office of Management and Budget under control number 0579–0013) [38 FR 32916, Nov. 29, 1973, as amended at 48 FR 57473, Dec. 30, 1983; 56 FR 66783, Dec. 26, 1991; 75 FR 20772, Apr. 21, 2010] § 104.4 Products for research and eval uation. (a) An application for a U.S. Veteri nary Biological Product Permit to im port a biological product for research and evaluation shall be accompanied by a brief description of such product, methods of propagating antigens in cluding composition of medium, spe cies of animals or cell cultures in volved, degree of inactivation or at tenuation, recommendations for use, and the proposed plan of evaluation. The applicant shall also provide any in formation the Administrator may re quire in order to assess the product’s impact on the environment. (b)(1) A permit to import a biological product for research and evaluation shall not be issued unless the scientific capabilities of the investigator are de termined to be adequate to safeguard domestic animals and protect public health, interest, or safety from any deleterious effects which might result from use of such product. Special re strictions or tests may be specified as part of the permit when they are deemed necessary or advisable by the Administrator. (2) No person shall ship a product im ported under this section for research and evaluation anywhere in or from the United States unless authorized by the Administrator in accordance with the provisions of § 103.3 of this sub chapter. (c) A biological product shall not be imported for Research and Evaluation which is not packaged and labeled in accordance with § 112.9 of this sub chapter. (d) When a licensed product has been exported from the United States, a per mit may be issued to the producer for a small quantity of such product for in vitro Research and Evaluation tests: Provided, That, the importation of such product will not endanger the livestock or poultry of this country. (Approved by the Office of Management and Budget under control number 0579–0013) [38 FR 32916, Nov. 29, 1973, as amended at 48 FR 57473, Dec. 30, 1983; 52 FR 30131, Aug. 13, 1987; 56 FR 66783, Dec. 26, 1991] § 104.5 Products for distribution and sale. An application for a U.S. Veterinary Biological Product Permit to import a biological product for Distribution and Sale shall be accompanied by sup porting material necessary to satisfy the requirements provided in this sec tion. (a) A permit shall not be issued un less the conditions under which the bi ological product is to be prepared or the methods to be used are such as to reasonably insure that the product is pure, safe, potent, and efficacious. (1) Two copies of blueprints of the producing foreign establishment shall be submitted with the application un less satisfactory plans are on file with Animal and Plant Health Inspection Service from a previous application. The production facilities to be used for each product prepared at the establish ment shall be designated. (2) The manufacturer shall submit written authorization for properly ac credited inspectors to inspect without previous notification, and at such times as may be demanded by the aforesaid inspectors, all parts of the es tablishment in which biological prod ucts shall be prepared, all processes of preparation, and all records relative to such preparation. (3) The manufacturer shall furnish written assurance that a biological product to be imported for Distribution and Sale shall be prepared under the supervision of a person competent by education and experience to handle all matters pertaining to the preparation of such product and that each biologi cal product shall be prepared in accord ance with the regulations applicable to the product or in a manner acceptable to the Administrator so as to carry out the purposes of the Act. (4) The methods to be used in the preparation of each biological product VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00688 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 679 Animal and Plant Health Inspection Service, USDA § 104.8 shall be written into an approved Out line of Production prepared in accord ance with the applicable provisions of part 114 of this subchapter. Two copies of such Outlines of Production shall be submitted to Animal and Plant Health Inspection Service and be approved be fore the permit is issued. (5) Data shall be furnished by the ap plicant which establishes that the product involved complies with the provisions of the Act and the regula tions issued pursuant thereto. When deemed necessary to obtain required information, Animal and Plant Health Inspection Service may require that the product be tested under field condi tions within or outside the United States as the occasion demands. (b) The permittee shall furnish the following: (1) Adequate facilities for storing all imported biological products. An in spection of such facilities shall be made by inspectors before a permit is issued and additional inspections shall be made at any time subsequent to the importation of the biological products if deemed necessary by the Adminis trator; (2) Information regarding all claims to be made on labels and advertising matter used in connection with or re lated to the biological product to be imported; (3) Mounted copies of final container labels, carton labels, and enclosures to be used with the imported product as provided in part 112 of this subchapter; and (4) Samples of each serial from each shipment of biological products im ported or offered for importation. Such samples shall be collected, examined, and tested in a manner specified by the Administrator. The biological products being sampled shall not be further dis tributed by the permittee until re leased by Animal and Plant Health In spection Service. (Approved by the Office of Management and Budget under control number 0579–0013) [38 FR 32916, Nov. 29, 1973, as amended at 48 FR 57473, Dec. 30, 1983; 49 FR 21044, May 18, 1984; 56 FR 66783, Dec. 26, 1991; 75 FR 20772, Apr. 21, 2010] § 104.6 Products for transit shipment only. An application for a permit for Tran sit Shipment Only shall be required when a biological product is being shipped from one foreign country to another foreign country by way of the United States. The shipment shall move under a permit subject to the fol lowing restrictions: (a) The shipment shall be confined to the carrier at all times when such ship ment is to transit the United States on the same carrier on which it arrived. If the shipment is to be transferred to a carrier other than the one on which it shall arrive into the United States, a schedule of arrival and departure of each shipment shall be furnished by the permittee to Animal and Plant Health Inspection Service prior to ar rival in the United States. (b) The permittee shall be responsible to Animal and Plant Health Inspection Service for handling, storing, and for warding of the biological product. Ani mal and Plant Health Inspection Serv ice shall be notified of all shipments received and forwarded by the per mittee and an accurate accounting shall be made. (Approved by the Office of Management and Budget under control number 0579–0013) [38 FR 32916, Nov. 29, 1973, as amended at 48 FR 57473, Dec. 30, 1983; 56 FR 66784, Dec. 26, 1991; 61 FR 52873, Oct. 9, 1996] § 104.7 Product permit. (a) A permit shall be numbered and dated. (b) The purpose for which the product is imported shall be specified on the permit as for Research and Evaluation, Distribution and Sale, or Transit Ship ment Only. (c) A permit shall not be used after the date specified. [38 FR 32916, Nov. 29, 1973, as amended at 56 FR 66783, Dec. 26, 1991; 62 FR 13294, Mar. 20, 1997] § 104.8 Illegal shipments. (a) Biological products which are pre sented for importation without a per mit having been issued shall be re turned to the country of origin at the expense of the importer or in lieu VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00689 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 680 9 CFR Ch. I (1–1–18 Edition) Pt. 105 thereof, destroyed by Department per sonnel. (b) Biological products for Distribu tion and Sale presented for importa tion under a permit and found to be worthless, contaminated, dangerous, or harmful shall, within a period of 30 days after such finding, be returned to the country of origin at the expense of the importer or in lieu thereof, de stroyed by Department personnel: Pro vided, That such product shall not be returned to the country of origin while bearing a U.S. permit number on the label. PART 105—SUSPENSION, REVOCA TION, OR TERMINATION OF BIO LOGICAL LICENSES OR PERMITS Sec. 105.1 Suspension or revocation. 105.2 Notification of infractions. 105.3 Notices re: worthless, contaminated, dangerous, or harmful biological prod ucts. 105.4 Termination of licenses and permits for inactivity. AUTHORITY: 21 U.S.C. 151–159; 7 CFR 2.22, 2.80, and 371.4. § 105.1 Suspension or revocation. (a) An establishment license, product license, or permit issued under the Virus-Serum-Toxin Act may be for mally suspended or revoked after op portunity for hearing has been ac corded the licensee or permittee as pro vided in part 123 of this subchapter if the Secretary is satisfied that the li cense or permit is being used to facili tate or effect the preparation, sale, barter, exchange, shipment, or impor tation contrary to said Act of any worthless, contaminated, dangerous, or harmful biological product. Such use may be found to exist if: (1) The construction of the establish ment in which the biological product is prepared is defective, or the establish ment is not conducted as required by the regulations in parts 101 through 118 of this subchapter; (2) The methods of preparation of the product are faulty, or the product con tains impurities or lacks potency; (3) The product is so labeled or adver tised as to mislead or deceive the pur chaser in any particular; (4) The licensee, permittee, or the foreign manufacturer has failed to maintain and make available for in spection records in connection with the development and preparation of prod uct, has failed to provide complete and accurate information when requested, or has failed to provide complete and accurate information in the Outline of Production or in reports and records; (5) The licensee or permittee has vio lated or failed to comply with any pro vision of the Virus-Serum-Toxin Act or the regulations in this subchapter; (6) The license or permit is otherwise used to facilitate or effect the prepara tion, sale, barter, exchange, shipment, or importation, contrary to the Virus Serum-Toxin Act, of any worthless, contaminated, dangerous, or harmful biological product. (b) In case of willfulness or where the public health, interest, or safety so re quired the Secretary may, without hearing, informally suspend such es tablishment license, product license, or permit upon the grounds set forth in paragraph (a) of this section pending determination of formal proceedings under part 123 of this subchapter for suspension or revocation of the license or permit. [38 FR 23512, Aug. 31, 1973, as amended at 41 FR 44359, Oct. 8, 1976; 61 FR 52874, Oct. 9, 1996; 64 FR 43044, Aug. 9, 1999] § 105.2 Notification of infractions. If an infraction of a requirement of a product license is brought to the atten tion of the licensee by written notifica tion thereof by Animal and Plant Health Inspection Service, a subse quent violation of similar nature oc curring with the same licensed biologi cal product within 6 months of the said written notification shall be primafacie evidence of willful violation and the license for the product shall be subject to suspension or revocation under the provisions of § 105.1(b). [42 FR 31430, June 21, 1977, as amended at 56 FR 66783, Dec. 26, 1991] § 105.3 Notices re: worthless, contami nated, dangerous, or harmful bio logical products. (a) If at any time it appears that the preparation, sale, barter, exchange, shipment, or importation, as provided VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00690 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 681 Animal and Plant Health Inspection Service, USDA § 106.1 in the Virus-Serum-Toxin Act, of any biological product by any person hold ing a license or permit may be dan gerous in the treatment of domestic animals, the Secretary may without hearing notify the licensee or per mittee, and pending determination of formal proceedings instituted under part 123 of this subchapter for suspen sion or revocation of the license or per mit insofar as it authorizes the manu facture or importation of the par ticular product, no person so notified shall thereafter so prepare, sell, barter, exchange, ship, deliver for shipment, or import such product. (b) If a serial of biological product is found to be unsatisfactory according to applicable Standard Requirements, the Administrator may notify the licensee to stop distribution and sale of the se rial. (c) When notified to stop distribution and sale of a serial or subserial of a veterinary biological product under the provisions of paragraph (a) or (b) of this section, veterinary biologics li censees or permittees shall: (1) Stop the preparation, distribu tion, sale, barter, exchange, shipment, or importation of the affected serial(s) or subserial(s) of any veterinary bio logical product pending further in structions from APHIS. (2) Immediately, but no later than 2 days, send stop distribution and sale notifications to any wholesalers, job bers, dealers, foreign consignees, or other persons known to have any such veterinary biological product in their possession, which instruct them to stop the preparation, distribution, sale, bar ter, exchange, shipment, or importa tion of any such veterinary biological product. All notifications shall be doc umented in writing by the licensee or permittee. (3) Account for the remaining quan tity of each serial(s) or subserial(s) of any such veterinary biological product at each location in the distribution channel known to the manufacturer (li censee) or importer (permittee). (4) When required by the Adminis trator, submit complete and accurate reports of all notifications concerning stop distribution and sale actions to the Animal and Plant Health Inspec tion Service pursuant to § 116.5 of this subchapter. (Approved by the Office of Management and Budget under control number 0579–0318) [38 FR 23512, Aug. 31, 1973, as amended at 56 FR 66783, Dec. 26, 1991; 72 FR 17798, Apr. 10, 2007] § 105.4 Termination of licenses and permits for inactivity. (a) If a biological product has not been prepared by a licensee, or im ported by a permittee for a period of 5 years or more, the Administrator may require the licensee to show intent to resume production, or the permittee to show intent to resume importation, within 6 months of notification. If the licensee does not resume preparation, or the permittee does not resume im portation, within 6 months of notifica tion, or within a mutually agreeable period, the product license, or permit, may be terminated by the Adminis trator. (b) When a license or permit is termi nated, the licensee or permittee shall continue to be subject to the applicable records provisions of § 116.8. [61 FR 52874, Oct. 9, 1996] PART 106—EXEMPTION FOR BIO LOGICAL PRODUCTS USED IN DE PARTMENT PROGRAMS OR UNDER DEPARTMENT CONTROL OR SUPERVISION AUTHORITY: 21 U.S.C. 151–159; 7 CFR 2.22, 2.80, and 371.4. § 106.1 Biological products; exemption. The Administrator may exempt any biological product from one or more of the requirements of this subchapter if he determines that such product will be used by the Department or under the supervision or control of the De partment in the prevention, control or eradication of animal diseases in con nection with (a) an official USDA pro gram; or (b) an emergency animal dis ease situation, or (c) a USDA experi mental use of the product. [45 FR 65184, Oct. 2, 1980, as amended at 56 FR 66783, Dec. 26, 1991] VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00691 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 682 9 CFR Ch. I (1–1–18 Edition) Pt. 107 PART 107—EXEMPTIONS FROM PREPARATION PURSUANT TO AN UNSUSPENDED AND UNREVOKED LICENSE Sec. 107.1 Veterinary practitioners and animal owners. 107.2 Products under State license. AUTHORITY: 21 U.S.C. 151–159; 7 CFR 2.22, 2.80, and 371.4. § 107.1 Veterinary practitioners and animal owners. Products prepared as provided in paragraphs (a) and (b) of this section and facilities in which such products are prepared, shall be exempt from preparation pursuant to unsuspended and unrevoked establishment and prod uct licenses. Persons exempt from li censure under this part shipping prod ucts which contain live organisms shall provide any information the Adminis trator may require prior to shipment, or at any other time deemed necessary, in order to assess the products’ safety and effect on the environment. The shipment or delivery for shipment any where in or from the United States of any exempted product which is worth less, contaminated, dangerous, or harmful is prohibited, and any person shipping such product, or delivering such product for shipment, shall be subject to sanctions under the Act. (a)(1) Products prepared by a veteri nary practitioner (veterinarian) solely for administration to animals in the course of a State licensed professional practice of veterinary medicine by such veterinarian under a veterinarian client-patient relationship and facili ties in which such products are pre pared shall be exempt from licensing under the Act and regulations. Such a relationship is considered to exist when: (i) The veterinarian has assumed the responsibility for making medical judgments regarding the health of the animal(s) and the need for medical treatment, and the client (owner or other caretaker) has agreed to follow the instructions of the veterinarian; and when (ii) There is sufficient knowledge of the animal(s) by the veterinarian to initiate at least a general or prelimi nary diagnosis of the medical condition of the animal(s). This means that the veterinarian has recently seen and is personally acquainted with the keeping and care of the animal(s), and/or by medically appropriate and timely vis its to the premises where the animal(s) are kept; and when (iii) The practicing veterinarian is readily available for followup in case of adverse reactions or failure of the regi men. (2) All steps in the preparation of product being prepared under the ex emption in paragraph (a)(1) of this sec tion must be performed at the facilities that the veterinarian utilizes for the day-to-day activities associated with the treatment of animals in the course of his/her State-licensed professional practice of veterinary medicine. A vet erinary assistant employed by the vet erinary practitioner and working at the veterinary practice’s facility under the veterinarian’s direct supervision may perform the steps in the prepara tion of product. Such preparation may not be consigned to any other party or sub-contracted to a commercial labora tory/manufacturing facility. (3) Veterinarians preparing products subject to the exemption for products under this section shall maintain and make available for inspection by Ani mal and Plant Health Inspection Serv ice representatives or other Federal employees designated by the Secretary such records as are necessary to estab lish that a valid veterinarian-client-pa tient relationship exists and that there is a valid basis for the exemption under this section. (b) Products prepared by a person solely for administration to animals owned by that person shall be exempt from the requirement that preparation be pursuant to an unsuspended and unrevoked license. [52 FR 30131, Aug. 13, 1987, as amended at 56 FR 66783, Dec. 26, 1991; 80 FR 26821, May 11, 2015] § 107.2 Products under State license. (a) The Administrator shall exempt from the requirement of preparation pursuant to an unsuspended and unrevoked USDA establishment and product license, any biological product prepared solely for distribution within VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00692 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 683 Animal and Plant Health Inspection Service, USDA § 108.3 the State of production pursuant to a license granted by such State under a program determined by the Adminis trator to be consistent with the intent of the Act to prohibit the preparation, sale, barter, exchange, or shipment of worthless, contaminated, dangerous, or harmful biological products. (b) A request for exemption under this section must be made by the ap propriate State authority and shall in clude information demonstrating that: (1) The State has the authority to li cense viruses, serums, toxins, and anal ogous products and establishments that produce such products; and (2) The State has the authority to re view the purity, safety, potency, and efficacy of such products prior to re lease to the market; and (3) The State has the authority to re view product test results to assure compliance with applicable standards of purity, safety, and potency prior to release to the market; and (4) The State has the authority to deal effectively with violations of State law regulating viruses, serums, toxins, and analogous products; and (5) The State effectively exercises the authority specified in paragraphs (b)(1) through (4) of this section consistent with the intent of the Act prohibiting the preparation, sale, barter, exchange, or shipment of worthless, contami nated, dangerous, or harmful viruses, serums, toxins, or analogous products. (c) Each product to be exempted and each establishment preparing such product shall be identified by the State and the State shall give written notifi cation to the Administrator of each such product and establishment. The State shall also give written notice to the Administrator of each new license issued and of each license terminated. (d) In order to determine whether a State exercises its authority with re spect to biological products and estab lishments and whether its laws and regulations are being achieved, the Ad ministrator, in cooperation with proper State authorities, may conduct an on site evaluation of the State’s program which may include inspection of estab lishments and/or products to be in cluded under the exemptions in this section. [52 FR 30131, Aug. 13, 1987, as amended at 56 FR 66783, Dec. 26, 1991] PART 108—FACILITY REQUIREMENTS FOR LICENSED ESTABLISHMENTS Sec. 108.1 Applicability. 108.2 Plot plans, blueprints, and legends re quired. 108.3 Preparation of plot plans. 108.4 Preparation of blueprints. 108.5 Preparation of legends. 108.6 Revision of plot plans, blueprints, and legends. 108.7 Filing of plot plans, blueprints, and legends. 108.8 Construction of buildings. 108.9 Dressing rooms and other facilities. 108.10 Outer premises and stables. 108.11 Water quality requirements. AUTHORITY: 21 U.S.C. 151–159; 7 CFR 2.22, 2.80, and 371.4. SOURCE: 39 FR 16854, May 10, 1974, unless otherwise noted. § 108.1 Applicability. Unless otherwise authorized by the Administrator, all buildings, appur tenances, and equipment used in the preparation of biological products shall be in compliance with the regulations in this part. Each land area on which such buildings and appurtenances are located shall be identified by an ad dress which shall appear on the estab lishment license. [39 FR 16854, May 10, 1974, as amended at 56 FR 66783, Dec. 26, 1991] § 108.2 Plot plans, blueprints, and leg ends required. Each applicant for an establishment license shall prepare a plot plan show ing all buildings for each particular land area, blueprints for each building used in the preparation of biological products and legends containing a brief description of all activities in each room or area. § 108.3 Preparation of plot plans. Plot plans shall show all of the build ings on a particular land area, whether or not they are all used for the prepa ration and initial shipping of biological products: Provided, That, when a great VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00693 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 684 9 CFR Ch. I (1–1–18 Edition) § 108.4 number of buildings are on the same premises, only those surrounding the buildings used for preparation and ini tial shipping of biological products shall be shown. The presence of the re mainder of the buildings may be ac counted for by a single statement de noting the total number of such build ings not used for the preparation or shipping of biological products. (a) Reduce the entire premises to any standard scale on one sheet of paper which meets any of the American standard trimmed sizes. Indicate the scale used. (b) Clearly mark the boundaries of the licensed premises and indicate what marking denotes the boundaries. Such boundaries shall coincide with some readily apparent perimeter line. Identify all fences, walls, or streets. (c) Show buildings as reduced dimen sional drawings in the proper scale dis tance relationship with each other. (d) Number, letter, or otherwise iden tify all buildings so that they may be correlated with the respective blue prints and legends. (e) Describe on the plot plan the use of immediate adjacent properties such as, residential area, pasture, box fac tory, or the like. (f) Show compass points. (g) Show date of preparation. (h) Apply signature of responsible of ficial of the firm. § 108.4 Preparation of blueprints. (a) Blueprints, drawn to any suitable scale, on regular blueprint paper or a good grade of white paper of any one of the American standard trimmed sizes shall be acceptable: Provided, That the same scale shall be used for future revi sions unless the entire blueprint is re vised. Indicate the scale used. (b) Use a single sheet of paper for each floor of all buildings in which bio logical products are prepared. Illus trate in detail the areas in each build ing utilized for such preparation. (c) If only a portion of a floor is used in the preparation of a biological prod uct, the blueprint shall illustrate the entire floor in essentially the same de tail throughout. All functions or ac tivities performed in the remainder of the floor shall be indicated. (d) Identify the floors if the drawing is not for all floors in a multiple-story building and identify activities on each floor. (e) Identify all rooms by letters or numbers. (f) Show the location of important stationary equipment by a suitable code which will be further identified on legends. (g) Explain on the blueprint or on the legend, by a statement or listing, which rooms are equipped with water outlets, drains, and lighting. Show the location of doors and windows. (h) Show compass points. (i) Show building number. (j) Show date of preparation. (k) Apply signature of responsible of ficial of firm. § 108.5 Preparation of legends. A brief description of the activities performed in each room or area shall be prepared as provided in this section and shall be referred to as a legend. Legends shall be provided for each plot plan and each blueprint or drawing. All pages of the legends shall be numbered, identified with corresponding plot plan or blueprint, and submitted in booklet form either stapled together or clipped into a suitable folder. (a) Plot plan legends shall show the following: (1) Number of each building and the functions performed in each: Provided, That if it is a multiple-story building in which biological products are pre pared or handled, briefly describe func tions performed on each floor. (2) A practical and nontechnical de scription of construction materials used throughout those buildings used entirely or partially for production and handling of biological products. (b) Blueprint legends shall show the following: (1) A listing of all rooms by identi fying letters or numbers and the frac tions prepared in each. Exceptions may be listed for general purpose areas or rooms. Functions performed in each area and room shall be described, whether the licensed or unlicensed products. In rooms where products are exposed to the surroundings, a descrip tion of decontamination procedures VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00694 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 685 Animal and Plant Health Inspection Service, USDA § 108.8 and other precautions against cross contamination shall be included. (2) A listing of the coded stationary equipment. (3) A general listing of other essen tial biological equipment such as mills, centrifuges, mixing tanks, bottling and sealing equipment, and the like, which are not regarded as stationary but are maintained in certain rooms. [39 FR 16854, May 10, 1974, as amended at 40 FR 51413, Nov. 5, 1975; 50 FR 50764, Dec. 12, 1985] § 108.6 Revision of plot plans, blue prints, and legends. Preliminary drawings may be sub mitted to Animal and Plant Health In spection Service for comment prior to construction of new facilities or when remodeling is anticipated, old facilities are to be torn down, or other changes affecting the workflow are to be made. The licensee shall: (a) Prepare revised plot plans, blue prints, or legends and submit to Ani mal and Plant Health Inspection Serv ice for review and filing when changes have been completed. Also prepare a statement to accompany each revision to identify, by date of the superseded item, what is being superseded. (b) Prepare a drawing of the revised rooms, unit, or section to the same scale as the blueprint on file which shall be stamped and applied to the ex isting blueprint. If changes are numer ous, prepare a new blueprint. (c) Drawings of new buildings may be added to existing plot plans. Indicate the distance from surrounding build ings and boundary lines. (d) Any change prescribed in this sec tion shall necessitate a change in one or more pages of the respective leg ends. The revised pages shall carry the same numbers as superseded pages. [39 FR 16854, May 10, 1974, as amended at 56 FR 66783, Dec. 26, 1991] § 108.7 Filing of plot plans, blueprints, and legends. Two copies of all plot plans, blue prints, and legends, including revi sions, shall be submitted to Animal and Plant Health Inspection Service for review and filing. When the re viewer takes exception to a submitted item, such item shall be returned with appropriate comments for correction and resubmission. Acceptable submis sions shall be stamped as filed and the date noted. One stamped copy shall be returned and two copies retained for Animal and Plant Health Inspection Service files. [39 FR 16854, May 10, 1974, as amended at 56 FR 66783, Dec. 26, 1991; 75 FR 20772, Apr. 21, 2010] § 108.8 Construction of buildings. (a) The floors, walls, ceilings, parti tions, posts, doors, and all other parts of all structures, rooms, or facilities used for the preparation of biological products or ingredients of biological products at licensed establishments shall be of such material, construction, and finish as may be readily and thor oughly cleaned. (b) All rooms used in connection with the preparation of biological products shall be so constructed and arranged as to prevent cross-contamination of such biological products. Halls or walkways shall be provided for the movement of personnel or materials to each biologi cal products preparation area without going through another such area. (c) Rooms or compartments separate from the remainder of the establish ment shall be provided at licensed es tablishments for preparing, handling, and storing virulent or dangerous microorganisms and products. (d) All rooms and compartments at licensed establishments shall have an adequate air handling system to supply proper ventilation sufficient to insure sanitary and hygienic conditions for the protection of the products and per sonnel. (e) The supply of hot and cold water at licensed establishments shall be ample and clean. Adequate facilities shall be provided for the distribution of water in each establishment and for the washing of all containers, machin ery, instruments, other equipment, and animals used in the preparation of a bi ological product. (f) There shall be an efficient drain age and plumbing system for each li censed establishment and premises thereof, and all drains and gutters shall be properly installed with ap proved traps and vents. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00695 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 686 9 CFR Ch. I (1–1–18 Edition) § 108.9 § 108.9 Dressing rooms and other fa cilities. Each licensed establishment shall have dressing rooms, toilet facilities, and lavatory accommodations, includ ing hot and cold running water, soap, towels, and the like. They shall be in sufficient number, ample in size, con veniently located, properly ventilated, and meeting all requirements as to sanitary construction and equipment. (a) These rooms and facilities shall be separate from rooms or compart ments in which biological products are prepared, handled, or stored. (b) These rooms and facilities shall be so located in the establishment as to be readily accessible to all persons without having to enter or pass through biological products prepara tion areas. § 108.10 Outer premises and stables. (a) The outer premises of licensed es tablishments, embracing docks, drive ways, approaches, yards, pens, chutes, and alleys shall be drained properly and kept in a clean and orderly condi tion. No nuisance shall be allowed in any licensed establishment or on its premises. (b) Stables or other premises for ani mals used in the production or testing of biological products at licensed es tablishments shall be properly venti lated and lighted, appropriately drained and guttered, and kept in sani tary condition. (c) Every practical precaution shall be taken to keep licensed establish ments free of flies, rats, mice, and other vermin. The accumulation, on the premises of an establishment, of any material in which flies or other vermin may breed is forbidden. Suit able arrangements, in keeping with the local health practices, shall be made for the disposal of all refuse. § 108.11 Water quality requirements. A certification from the appropriate water pollution control agency, that the establishment is in compliance with applicable water quality control standards, pursuant to section 401 of the Federal Water Pollution Control Act, as amended (86 Stat. 877; 33 U.S.C. 1341), shall be filed with Animal and Plant Health Inspection Service for each licensed establishment. [39 FR 16854, May 10, 1974, as amended at 56 FR 66783, Dec. 26, 1991] PART 109—STERILIZATION AND PASTEURIZATION AT LICENSED ESTABLISHMENTS Sec. 109.1 Equipment and the like. 109.2 Sterilizers. 109.3 Pasteurizers. AUTHORITY: 21 U.S.C. 151–159; 7 CFR 2.22, 2.80, and 371.4. § 109.1 Equipment and the like. (a) All containers, instruments, and other apparatus and equipment, before being used in preparing, handling, or storing biological products, at a li censed establishment, except as other wise prescribed herein, shall be thor oughly sterilized by live steam at a temperature of at least 120 °C. for not less than one-half hour, or by dry heat at a temperature of at least 160 °C. for not less than one hour. If for any rea son such methods of sterilization are impracticable, then a process known to be equally efficacious in destroying microorganisms and their spores may be substituted after approval by the Administrator. (b) Instruments which are found to be damaged by exposure to the degree of heat prescribed in this section, after having been thoroughly cleaned, may be sterilized by boiling for not less than 15 minutes. [23 FR 10051, Dec. 23, 1958, as amended at 34 FR 18119, Nov. 11, 1969; 56 FR 66783, Dec. 26, 1991] § 109.2 Sterilizers. Steam and dry-heat sterilizers used in connection with the processing of bi ological products at licensed establish ments shall be equipped with auto matic temperature recording gauges: Provided, That other record keeping systems may be used when approved by the Administrator. When gauges are used, they shall be periodically stand ardized to assure accuracy. Charts and other temperature records made during production shall be available at all times charts and records shall be kept VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00696 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 687 Animal and Plant Health Inspection Service, USDA § 112.2 in accordance with part 116 of this chapter. [35 FR 16039, Oct. 13, 1970, as amended at 56 FR 66783, Dec. 26, 1991] § 109.3 Pasteurizers. All pasteurizing equipment shall meet the requirements in paragraphs (a), (b), and (c) of this section and be acceptable to Animal and Plant Health Inspection Service. (a) Metal serum containers shall be used in licensed establishments. During the heating process, each container shall be surrounded by a separate water jacket or equivalent so that the entire container, including its lid, is heated to the required temperature. Each serum container shall be equipped with a motor-driven agitator and a sep arate automatic recording thermom eter. (b) Each water bath shall have an automatic temperature control to limit the temperature of the water to a maximum of 62 °C., an automatic re cording thermometer, an indicating thermometer set in a fixed position, and circulating mechanism adequate to insure equal temperatures throughout the bath. The heating unit for the bath shall be separated from the serum con tainer and the water jacket. (c) Accurate thermometers at li censed establishments shall be used at frequent intervals to check tempera tures of the serum as registered by re cording thermometers. [35 FR 16039, Oct. 13, 1970, as amended at 56 FR 66783, Dec. 26, 1991] PART 112—PACKAGING AND LABELING Sec. 112.1 General. 112.2 Final container label, carton label, and enclosure. 112.3 Diluent labels. 112.4 Subsidiaries, divisions, distributors, and permittees. 112.5 Review and approval of labeling. 112.6 Packaging biological products. 112.7 Special additional requirements. 112.8 For export only. 112.9 Biological products imported for re search and evaluation. 112.10 Special packaging and labeling. AUTHORITY: 21 U.S.C. 151–159; 7 CFR 2.22, 2.80, and 371.4. SOURCE: 38 FR 12094, May 9, 1973, unless otherwise noted. § 112.1 General. (a) Unless otherwise authorized or di rected by the Administrator, each bio logical product prepared at a licensed establishment, or imported, shall be packaged and labeled as prescribed in this part before it is removed from the licensed establishment or presented for importation: Provided, That biological products to be imported for research and evaluation shall be subject to packaging and labeling requirements in § 112.9. Provided further, That, unless otherwise exempted, all preparation, including packaging and labeling, of bi ological products shall only be per formed in a licensed establishment under an approved Outline of Produc tion. (b) No person shall apply or affix to or include with, or cause to be applied or affixed to or included with, any car ton or final container of a biological product, any label, stamp, mark or statement that is false or misleading in any particular, is not in compliance with the regulations, or is not ap proved by APHIS. (c) No person shall alter, mark or re move any approved labeling affixed to or included with any biological product prior to selling or otherwise distrib uting such product. In addition, no per son shall mark any carton, other con tainer, or final container of a biologi cal product so as to falsify the labeling, make it misleading, or cause it to be il legible. (d) Labels that are stamped, printed or glued directly on cartons, other con tainers, or final containers shall be leg ible throughout the dating period. Bio logical products bearing labels, which have been altered, mutilated, de stroyed, obliterated or removed, shall be withheld from the market. [38 FR 12094, May 9, 1973, as amended at 59 FR 43445, Aug. 24, 1994] § 112.2 Final container label, carton label, and enclosure. (a) Unless otherwise provided, final container labels, carton labels, and en closures (inserts, circulars, or leaflets) VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00697 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 688 9 CFR Ch. I (1–1–18 Edition) § 112.2 shall include the information specified in this section. (1) The complete true name of the bi ological product which name shall be identical with that shown in the prod uct license under which such product is prepared or the permit under which it is imported, shall be prominently let tered and placed giving equal emphasis to each word composing it. Descriptive terms used in the true name on the product license or permit shall also ap pear. Abbreviations of the descriptive terms may be used on the final con tainer label if complete descriptive terms appear on the carton label and enclosure. The following exceptions are applicable to small final containers, and containers of interchangeable re agents included in diagnostic test kits: (i) For small final containers, an ab breviated true name of the biological product, which shall be identical with that shown in the product license under which the product is prepared or the permit under which it is imported, may be used: Provided, That the com plete true name of the product must appear on the carton label and enclo sures; (ii) In addition to the true name of the kit, the functional and/or chemical name of the reagent must appear on la beling for small final containers of re agents included in diagnostic kits: Pro vided, That the true name is not re quired on labeling for small final con tainers of interchangeable (non-crit ical) components of diagnostic kits. (2) For biological product prepared in the United States or in a foreign coun try, the name and address of the pro ducer (licensee, or subsidiary) or per mittee and of the foreign producer, and an appropriate consumer contact tele phone number: Provided, That in the case of a biological product exported from the United States in labeled final containers, a consumer contact tele phone number is not required; however, small single dose containers marketed in the United States must include con tact telephone information on carton and enclosures. (3) The United States Veterinary Bio logics Establishment License Number (VLN) or the United States Veterinary Biological Product Permit Number (VPN), and the Product Code Number (PCN) assigned by the Department, which shall be shown only as ‘‘VLN/ PCN’’ and ‘‘VPN/PCN,’’ respectively, except that: (i) Only the VLN or VPN is required on container labels of interchangeable (non-critical) components of diagnostic kits and container labels for individual products packaged together for co-ad ministration. (ii) The PCN may be used in lieu of the true name of the kit on small con tainer labels for critical components of diagnostic kits. (iii) Container labels for individually licensed biological products, when mar keted as components of combination packages, must include a statement re ferring the consumer to the carton or enclosure for the PCN of the combina tion package. (4) Storage temperature rec ommendation for the biological prod uct stated as 2 to 8 °C or 35 to 46 °F, or both. (5) Full instructions for the proper use of the product, including indica tions for use, target species, minimum age of administration, route of admin istration, vaccination schedule, prod uct license restriction(s) that bear on product use, warnings, cautions, and any other vital information for the product’s use; except that in the case of limited space on final container la bels, a statement as to where such in formation is to be found, such as ‘‘See enclosure for complete directions,’’ ‘‘Full directions on carton,’’ or com parable statement. (6) In the case of a multiple-dose final container, a warning to use entire contents when first opened: Provided, That a diagnostic or a desensitizing antigen packaged in a multiple-dose final container is exempt. (7) The following warning state ments, or equivalent statements, shall appear on the labeling as applicable: (i) Products other than diagnostic kits: ‘‘Do not mix with other products, except as specified on this label.’’ (ii) Injectable products and other products containing hazardous compo nents: ‘‘In case of human exposure, contact a physician.’’ (iii) Products containing viable orga nisms: ‘‘Inactivate unused contents be fore disposal.’’ VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00698 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 689 Animal and Plant Health Inspection Service, USDA § 112.2 (8) In the case of a biological product recommended for use in domestic ani mals, the edible portion of which may be used for food purposes, a with holding statement of not less than 21 days to read: ‘‘Do not vaccinate within (insert number) days before slaughter’’ or ‘‘Do not vaccinate food-producing animals within (insert number) days before slaughter’’: Provided, That longer periods shall be stated when deemed necessary by the Adminis trator. Very small final container la bels are exempted from this require ment. (9) The following information shall appear on the final container label and carton label, if any, but need not ap pear on the enclosure: (i) A permitted expiration date; (ii) The number of doses where appli cable; (iii) The recoverable quantity of the content of each final container stated in cubic centimeters (cc.) or milliliters (ml.) or units. (iv) A serial number by which the product can be identified with the man ufacturer’s records of preparation: Pro vided, That when a liquid antigenic fraction is to be used instead of a water diluent for one or more desiccated an tigenic fractions in a combination package, a hyphenated serial number composed of a serial number for the desiccated fraction and the serial num ber for the liquid fraction shall be used on the carton. (v) A statement similar to ‘‘For more information regarding efficacy and safety data, go to productdata.aphis.usda.gov. (10) In the case of a product that con tains a preservative that is added dur ing the production process and is not reduced to undetectable levels in the completed product through the produc tion process, the statement ‘‘Contains [name of preservative] as a preserva tive’’ or an equivalent statement must appear on cartons and enclosures, if used. If cartons are not used, such in formation must appear on the final container label. (11) The number of final containers of biological product and the number of doses in each final container shall be stated on each carton label for all car tons containing more than one final container of biological product. The number of final containers of diluent, if any, and the quantity in each shall also be stated on each carton label. (12) An indications statement to read, ‘‘This product has been shown to be effective for the vaccination of healthy (insert name of species) ll weeks of age or older against ll.’’ Provided, That in the case of very small final container labels or carton, a statement as to where such informa tion is to be found, such as ‘‘See enclo sure for complete directions,’’ ‘‘Full di rections on carton,’’ or comparable statement. (b) Labels may also include any other statement which is not false or mis leading and may include factual state ments regarding variable response of different animals when vaccinated as directed but may not include dis claimers of merchantability, fitness for the purpose offered, or responsibility for the product. (c) Labels of biological products pre pared at licensed establishments or im ported shall not include any statement, design, or device, which overshadows the true name of the product as li censed or which is false or misleading in any particular or which may other wise deceive the purchaser. (d) Carton labels and enclosures shall be subject to paragraph (d)(1), (d)(2), and (d)(3) of this section. (1) The statement, ‘‘Restricted to use by or under the direction of a veteri narian’’ or ‘‘Restricted to use by a vet erinarian,’’ shall be used on all carton labels and enclosures when such re striction is prescribed on the product license. (2) If the licensee states on the car ton labels and enclosures of a product that its sales are restricted to veteri narians, then the entire production of that particular product in the licensed establishment shall be so restricted by the licensee. (3) The statement ‘‘For use in ani mals only’’ may appear on the labeling as appropriate for a product to indicate that the product is recommended spe cifically for animals and not for hu mans. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00699 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 690 9 CFR Ch. I (1–1–18 Edition) § 112.3 (e) When label requirements of a for eign country differ from the require ments as prescribed in this part, spe cial labels may be approved by APHIS for use on biological products to be ex ported to such country upon receipt of written authorization, acceptable to APHIS, from regulatory officials of the importing country, provided that: (1) If the labeling contains claims or indications for use not supported by data on file with APHIS, the special la bels for export shall not bear the VLN. (2) All other labels for export shall bear the VLN unless the importing country provides documentation that the VLN is specifically prohibited. When laws, regulations, or other re quirements of foreign countries require exporters of biological products pre pared in a licensed establishment to furnish official certification that such products have been prepared in accord ance with the Virus-Serum-Toxin Act and regulations issued pursuant to the Act, such certification may be made by APHIS. (f) Multiple-dose final containers of liquid biological product and carton tray covers showing required labeling information are subject to the require ments in this paragraphs. (1) If a carton label or an enclosure is required to complete the labeling for a multiple-dose final container of liquid biological product, only one final con tainer, with a container of diluent if applicable, shall be packaged in each carton: Provided, That if the multiple dose final container is fully labeled without a carton label or enclosure, two or more final containers, and a corresponding number of diluent con tainers, may be packaged in a single carton which shall be considered a shipping box. Labels or stickers for shipping boxes shall not contain false or misleading information, but need not be submitted to APHIS for ap proval. (2) When required labeling informa tion is shown on a carton tray cover, it must be printed on the outside face of such tray cover where it may be read without opening the carton. The inside face of the tray cover may contain in formation suitable for an enclosure. (Approved by the Office of Management and Budget under control number 0579–0013) [38 FR 12094, May 9, 1973, as amended at 39 FR 16856, May 10, 1974; 41 FR 44359, Oct. 8, 1976; 42 FR 11825, Mar. 1, 1977; 42 FR 29854, June 10, 1977; 42 FR 41850, Aug. 19, 1977; 48 FR 57473, Dec. 30, 1983; 56 FR 66784, Dec. 26, 1991; 80 FR 39674, July 10, 2015; 81 FR 59433, Aug. 30, 2016; 81 FR 78500, Nov. 8, 2016] § 112.3 Diluent labels. Each final container of diluent, other than a liquid biological product, pack aged with desiccated biological prod ucts shall bear a label that includes the following: (a) The name—Sterile Diluent. (b) True name of the biological prod uct with which the diluent is packaged, except that when the firm packages all desiccated biological products with the same diluent, or two or more types of diluent are used, and the licensees’ methods of identification and storage insure that all products are packaged with the correct type of diluent, labels affixed to the containers of diluent are exempt from this provision. (c) The recoverable quantity of con tents in cubic centimeters (cc) or milli liters (ml). (d) A serial number by which the dil uent can be identified with the manu facturer’s records of preparation; (e) Name and address of the licensee or the permittee; (f) In the case of a diluent with which a desiccated biological product is to come in contact while the diluent is in its original container; and, (1) Is in a multiple-dose container, a positive warning that all of the biologi cal product shall be used at the time the container is first opened; and/or (2) The biological product is com posed of viable or dangerous organisms or viruses, the notice, ‘‘Inactivate un used contents before disposal.’’ (g) The establishment license number or the permit number, as the case may be, in one of the forms provided in § 112.2(a)(3). [38 FR 12094, May 9, 1973; 38 FR 13476, May 22, 1973, and amended at 39 FR 16856, May 10, 1974; 81 FR 59434, Aug. 30, 2016] VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00700 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 691 Animal and Plant Health Inspection Service, USDA § 112.5 § 112.4 Subsidiaries, divisions, dis tributors, and permittees. Labels used by subsidiaries, divi sions, distributors, and permittees shall be affixed by the licensee in a li censed establishment where the prod uct is produced. Such labels shall com ply with requirements for their review, approval, and filing as provided in the regulations. (a) Subsidiaries. Labels to be used on a licensed biological product prepared by a subsidiary operating in a licensed establishment shall be submitted in ac cordance with § 112.5. Only labels ap proved for use on such product shall be used by the subsidiary. (b) Divisions. Labels to be used on a licensed biological product prepared in a licensed establishment for distribu tion by a division or marketing unit of the licensee shall be submitted in ac cordance with § 112.5. The name, ad dress, and license number of the li censee shall be prominently placed on such labels. The relationship of the di vision or marketing unit to the li censee shall appear prominently on the label by use of the term ‘‘division of’’ or equivalent. (c) Distributors. The name and address of the distributor or any statement, de sign, or device shall not be placed on the labels or containers of a licensed biological product in a manner which could be false or misleading or which could indicate that the distributor is the manufacturer of such product or operating under the license number shown on the label. The manufacturer shall be identified by name, address, and license number with the term ‘‘manufactured by,’’ ‘‘produced by,’’ or an equivalent term prominently placed in connection therewith. The name and address of the distributor may be placed on labels or containers if the term ‘‘distributor,’’ or ‘‘distributed by,’’ or an equivalent term is promi nently placed in connection therewith. (d) Permittees. The name and address of the permittee and any statement, design, or device shall not be placed on the labels or containers of a biological product imported for sale and distribu tion in accordance with § 104.5 in a manner which could be false or mis leading or which could falsely indicate that the permittee is the manufacturer of such product. The manufacturer shall be identified by name and address with the term ‘‘manufactured by,’’ ‘‘produced by,’’ or an equivalent term prominently placed in connection therewith. Reference to the permittee shall be made by name, address, and permit number with the term ‘‘im ported by,’’ ‘‘produced for,’’ or an equivalent term prominently placed in connection therewith. [50 FR 46417, Nov. 8, 1985, as amended at 59 FR 43445, Aug. 24, 1994] § 112.5 Review and approval of label ing. Labels used with biological products prepared at licensed establishments or imported for general distribution and sale must be submitted to the Animal and Plant Health Inspection Service for review for compliance with the reg ulations and approval in writing prior to use, except as provided in paragraph (d) of this section and under the master label system provided in paragraph (e) of this section. (a) Transmittal forms, available on the APHIS Web page at http:// www.aphis.usda.gov/animalhealth/cvb/ forms, shall be used with each submis sion of sketches (including proofs) and labels. Separate forms shall be used for each biological product but only one copy of the form shall be used for all sketches and labels submitted at the same time for the same biological product. (b) A data summary, available on the Internet at productdata.aphis.usda.gov, shall be used with each submission of efficacy and safety data in support of a label claim. Manufacturers will submit the efficacy and safety data informa tion with either the efficacy and safety studies or at the time of label submis sion. This information will be posted at productdata.aphis.usda.gov to allow public disclosure of product perform ance. (c) Sketches may be submitted for comment to Animal and Plant Health Inspection Service by the licensee or permittee before preparing the finished label. Such sketches shall be returned to the licensee or permittee with com ments, if any. Failure of the reviewer to take exception to a sketch shall not VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00701 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 692 9 CFR Ch. I (1–1–18 Edition) § 112.5 constitute approval of a finished label subsequently prepared. (d)(1) Labels must be submitted to the Animal and Plant Health Inspec tion Service for review and written ap proval. Only labels which are approved as provided in paragraph (e) of this sec tion may be used. When changes are made in approved labels, the new labels shall be subject to review and approval before use: Provided, That certain minor changes may be made in labels for products with approved labels or master labels, and the revised labels may be used prior to review by APHIS, with the provision that a new label or master label bearing these changes is submitted to APHIS for review and written approval within 60 days of label use, and that such minor changes do not render the product mislabeled or the label false and misleading in any particular. (2) Minor label changes that may be made under the provision for products with approved labels or master labels are: (i) Changes in the physical dimen sions of the label provided that such change does not affect the legibility of the label; (ii) Changes in the color of label print or background, provided that such changes do not affect the legibility of the label; (iii) The addition or deletion of a Trade Mark (TM) or Registered (R) symbol; (iv) The correction of typographical errors; (v) Adding, changing, deleting, or repositioning label control numbers, universal product codes, or other in ventory control numbers; (vi) Revising or updating logos; (vii) Changing the telephone contact number; (viii) Adding, changing, or deleting an email and/or Web site address; (ix) Changing the establishment li cense or permit number assigned by APHIS, and/or changing the name and/ or address of the manufacturer or per mittee, provided that such changes are identical to information on the current establishment license or permit; and (x) Adding or changing the name and/ or address of a distributor. (e) Labels and sketches submitted shall be prepared in the number and manner prescribed in this paragraph. (1) Copies required: (i) For label sketches, submit two copies of each sketch of a final con tainer label, carton label, and enclo sure. Sketches must be legible, and must include all information specified in § 112.2. One copy of each sketch will be returned with applicable comments, and one copy will be held on file by APHIS for no more than one year after processing, until replaced by a finished label: Provided, That sketches sub mitted in support of an application for a license or permit shall be held as long as the application is considered active. (ii) For master label sketches, sub mit for each product two copies of each sketch of an enclosure, label for the smallest size final container, and car ton label; Provided, That labels for larger size containers and/or cartons that are identical, except for physical dimensions, need not be submitted. One copy of each master label sketch will be returned with applicable comments, and one copy will be held on file by APHIS for one year after processing, until replaced by a finished master label that is submitted according to paragraph (e)(1)(iii) of this section: Pro vided, That master label sketches sub mitted in support of an application for license or permit shall be held as long as the application is considered active. (iii) For finished labels, submit two copies of each finished final container label, carton label, and enclosure: Pro vided, That when an enclosure is to be used with more than one product, one extra copy shall be submitted for each additional product. One copy of each finished label will be retained by APHIS. One copy will be stamped and returned to the licensee or permittee. Labels to which exceptions are taken shall be marked as sketches and han dled under paragraph (e)(1)(i) of this section. (iv) For finished master labels, sub mit for each product two copies each of the enclosure and the labels for the smallest size final container and car ton. Labels for larger sizes of con tainers or cartons of the same product that are identical, except for physical dimensions, need not be submitted. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00702 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 693 Animal and Plant Health Inspection Service, USDA § 112.5 Such labels become eligible for use concurrent with the approval of the ap propriate finished master label, pro vided that the marketing of larger size final containers is approved in the filed Outline of Production, and the appro priate larger sizes of containers or car tons are identified on the label mount ing sheet. When a master label enclo sure is to be used with more than one product, one extra copy for each addi tional product shall be submitted. One copy of each finished master label will be retained by APHIS. One copy will be stamped and returned to the licensee or permittee. Master labels to which exception are taken will be marked as sketches and handled under paragraph (e)(1)(ii) of this section. (2) Mounting: (i) Each label or sketch shall be se curely fastened to a separate sheet of heavy bond paper (81⁄2″ × 11″) in such a manner that all information is avail able for review. (ii) Two-or three-part cartons, in cluding ‘‘sleeves,’’ shall be considered as one label. All parts shall be sub mitted together. (iii)(A) When two final containers are packaged together in a combination package, the labels for each shall be mounted on the same sheet of paper and shall be treated as one label. For diagnostic test kits, the labels for use on the individual reagent containers to be included in the kit shall be mounted together on a single sheet of paper, if possible; if necessary, a second sheet of paper may be used. The carton label and enclosure shall be mounted on sep arate individual sheets. (B) If either final container label is also used alone or in another combina tion package, sets of separate labels for each biological product with which it is used shall be submitted for review. (iv) When the same final container label is applied by different methods such as paper or screen printing, one of each shall be mounted on the same sheet of paper as one submission. (3) To appear on the top of each page: (i)(A) Name and product code number of the biological product as it appears on the product license or permit. (B) Extra copies of enclosures to be used with another product shall bear the name and code number of the prod uct affected. (ii)(A) Designation of the specimen as a label or master label: sketch, final container label, carton label, or enclo sure. (B) If two final container labels or multiple parts are on one sheet, each shall be named, and the label or part being revised shall be designated. (iii) Size of package (dose, ml., cc., or units) for which the labels or enclo sures are to be used. (4) To appear on the bottom of each page in the lower left hand corner, if applicable: (i) The dose size(s) to which the mas ter label applies. (ii) The APHIS assigned number for the label or sketch to be replaced. (iii) The APHIS assigned number for the label to be used as a reference for reviewing the submitted label. (f) Special requirements for foreign language labels: (1) An accurate English translation must accompany each foreign language label submitted for approval. A state ment affirming the accuracy of the translation must also be included. (2) Foreign language portion of a bi lingual label shall be a true translation of the English portion. Reference to ad ditional information on the enclosure shall not be made unless that enclosure is also bilingual. (g) When a request is received from Animal and Plant Health Inspection Service, the licensee or permittee shall submit a list of all approved labels cur rently being used. Each label listed shall be identified as to: (1) Name and product code number as it appears on the product license or permit for the product; and (2) Where applicable, the size of the package (doses, ml., cc., or units) on which the label shall be used; and (3) Label number and date assigned; and (4) Name of licensee or subsidiary ap pearing on the label as the producer. (h) At the time of an inspection, or when requested by APHIS, licensees or permittees shall make all labels and master labels, including labels ap proved for use but exempted from filing VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00703 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 694 9 CFR Ch. I (1–1–18 Edition) § 112.6 under the master label system, avail able for review by authorized inspec tors. Such labels shall be identical to the approved label or master label ex cept for physical dimensions, reference to recoverable volume or doses and/or certain minor differences permitted in accordance with paragraph (d) of this section. (Approved by the Office of Management and Budget under control number 0579–0013) [38 FR 12094, May 9, 1973, as amended at 48 FR 57473, Dec. 30, 1983; 49 FR 21044, May 18, 1984; 56 FR 66783, Dec. 26, 1991; 59 FR 43445, Aug. 24, 1994; 61 FR 29464, June 11, 1996; 61 FR 33175, June 26, 1996; 64 FR 43044, Aug. 9, 1999; 75 FR 20772, Apr. 21, 2010; 80 FR 39675, July 10, 2015; 81 FR 47005, July 20, 2016; 81 FR 59435, Aug. 30, 2016] § 112.6 Packaging biological products. (a) Multiple-dose final containers of a biological product with final container labeling including all information re quired under the regulations may be packaged one or more per carton with a container(s) of the proper volume of diluent, if required, for that dose as specified in the filed Outline of Produc tion: Provided, That cartons containing more than one final container of prod uct must comply with the conditions set forth in paragraphs (c)(1) through (4) of this section. Multiple-dose final containers of a product that require a carton or enclosure in order to provide all information required under the reg ulations shall be packaged one con tainer per carton with the proper vol ume of diluent, if required, for that dose as specified in the filed Outline of Production. (b) Single-dose final containers of a product need not be packaged one per carton. For single-dose products which require a diluent for administration, the number of containers of the proper amount of diluent specified in the filed Outline of Production for the number of doses contained in the carton shall be included in each carton. (c) Poultry products for mass admin istration (including but not limited to administration through drinking water and spray) and products used in auto matic vaccinating systems (including but not limited to pneumatic beak injectors and automated needle injectors) may be packaged in mul tiple-dose final containers as specified in the filed Outline of Production. Poultry products for manual adminis tration to individual birds shall not ex ceed 1,000 doses in each final container. Diluent need not be packaged with the final container(s) of the product, but the licensee shall provide the required number of containers of diluent as specified in the filed Outline of Produc tion. The following requirements apply to cartons containing more than one final container of poultry product: (1) They shall be sealed prior to leav ing the licensed establishment. (2) The contents may not be repack aged. (3) The contents of such cartons may not be sold in fractional units. (4) The following statement must ap pear in a prominent place on the car ton label: ‘‘Federal regulations pro hibit the repackaging or sale of the contents of this carton in fractional units. Do not accept if seal is broken.’’ (d) Diluent for the following products need not be packaged with the final container(s) of the product, but the li censee shall provide the consumer with the required number of containers of the proper amount of diluent as speci fied in the filed Outline of Production: (1) Marek’s Disease Vaccine. (2) Poultry vaccines administered to individual birds using automatic vacci nating equipment. (e) Final containers of biological product prepared at a licensed estab lishment, or imported, in cartons or other containers shall not be removed from such cartons or containers for sale or distribution, unless each final container bears, or is packaged in a carton with, complete and approved la beling which is affixed to or included with each container by the licensed es tablishment producing the product or by the producer in the case of imported product: Provided, That this paragraph is not intended to apply to licensed veterinary practitioners administering or dispensing biological products in the course of their practice under a veteri nary-client-patient-relationship as that term is used in § 107.1. (f) Labels which are affixed to or in cluded with a biological product shall VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00704 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 695 Animal and Plant Health Inspection Service, USDA § 112.7 not be removed or altered in any man ner. [47 FR 8761, Mar. 2, 1982, as amended at 48 FR 12691, Mar. 28, 1983; 59 FR 43445, Aug. 24, 1994; 64 FR 43044, Aug. 9, 1999; 81 FR 59435, Aug. 30, 2016] § 112.7 Special additional require ments. The label requirements in this sec tion are additional to those prescribed elsewhere in this part. (a) In the case of biological products containing live Newcastle Disease virus, a caution statement indicating that Newcastle Disease can cause in flammation of the eyelids of humans, and a warning to the user to avoid in fecting his eyes shall be included on the enclosure. (b) In the case of a biological product containing infectious bronchitis virus, all labels shall show the infectious bronchitis virus type or types used in the product. Abbreviation is permitted. (c) In the case of a biological product containing inactivated rabies virus, carton labels, enclosures, and all but very small final container labels shall include a warning against freezing and the recommendations provided in this paragraph. (1) That vaccine be administered to animals at 3 months of age or older, with a repeat dose 1 year later. (2) Subsequent revaccination as de termined from the results of duration of immunity studies conducted as pre scribed in § 113.209, paragraph (b) or (c), or both. (d) In the case of a biological product containing modified live rabies virus, the carton labels, enclosures, and all but very small final container labels shall include the recommendations provided in this paragraph. (1) For low egg-passage (below the 180th egg-passage level) the statement ‘‘For Use in Dogs Only! Not For Use in Any Other Animal!’’ (2) For other vaccines containing modified live rabies virus, the state ment ‘‘For Use In (designate animal(s)) Only! Not For Use In Any Other Ani mal!’’ (3) Intramuscular injection at one site in the thigh shall be recommended. (4) The statement ‘‘In event of acci dental exposure to the vaccine virus, the possible hazard to human health should be considered and State Public Health Officials should be consulted for specific recommendations’’ shall be prominently placed on all carton labels and on enclosures, if used. (5) That vaccine be administered to animals at 3 months of age or older, with a repeat dose 1 year later. (6) Subsequent revaccination as de termined from the results of duration of immunity studies conducted as pre scribed in § 113.312, paragraph (b) or (c), or both. (e) Labeling for all products for use in mammals must bear an appropriate statement concerning use in pregnant animals. (1) For bovine rhinotracheitis vaccine or bovine virus diarrhea vaccine con taining modified live virus, all labeling except small final container labels shall bear the following statement: ‘‘Do not use in pregnant cows or in calves nursing pregnant cows.’’: Pro vided, That such vaccines which have been shown to be safe for use in preg nant cows may be excepted from this label requirement by the Adminis trator. (2) For other modified live and inac tivated vaccine, labeling shall bear a statement appropriate to the level of safety that has been demonstrated in pregnant animals. (i) Products known to be unsafe in pregnant animals shall include state ments such as ‘‘Do not use in pregnant animals,’’ or ‘‘Unsafe for use in preg nant animals,’’ or an equivalent state ment acceptable to APHIS. (ii) Products without safety docu mentation acceptable to APHIS, but not known to be unsafe, labeling shall include the statement ‘‘This product has not been tested in pregnant ani mals’’ or an equivalent statement ac ceptable to APHIS. (3) For modified live vaccines con taining agents with potential reproduc tive effects but having acceptable preg nant animal safety data on file with APHIS, labeling still must bear the fol lowing statement concerning residual risk: ‘‘Fetal health risks associated with the vaccination of pregnant ani mals with this vaccine cannot be un equivocally determined during clinical VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00705 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 696 9 CFR Ch. I (1–1–18 Edition) § 112.8 trials conducted for licensure. Appro priate strategies to address the risks associated with vaccine use in preg nant animals should be discussed with a veterinarian.’’ (f) For biological products recom mending annual booster vaccinations, such recommendations must be sup ported by data acceptable to APHIS. In the absence of data that establish the need for booster vaccination, labeling must bear the following statement: ‘‘The need for annual booster vaccina tions has not been established for this product; consultation with a veteri narian is recommended.’’ (g) In the case of a liquid product au thorized in a filed Outline of Produc tion to be used as a diluent in a com bination package, the carton labels and enclosures used for serials which are ei ther not tested for bactericidal or viri cidal activity or have been found un satisfactory by such test shall contain the statement: ‘‘CAUTION: DO NOT USE AS DILUENT FOR LIVE VAC CINES.’’ (h) In the case of wart vaccine, rec ommendations shall be limited to use in cattle. Indications for use shall be for prophylactic use only, as an aid in the control of viral papillomas (warts). All labels shall include a dosage rec ommendation of at least 10 ml to be given subcutaneously and the dose re peated in 3 to 5 weeks. (i) All but very small final container labels for feline panleukopenia vac cines shall contain the following rec ommendations for use: (1) Killed virus vaccines. Vaccinate healthy cats with one dose, except that if the animal is less than 12 weeks of age, a second dose should be given no earlier than 16 weeks of age. (2) Modified live virus vaccines. Vac cinate healthy cats with one dose, ex cept that if the animal is less than 12 weeks of age, a second dose should be given no earlier than16 weeks of age. (j) In the case of normal serum, anti serum, or antiserum derivatives, the type of preservative used shall be indi cated on all labels. (k) Unless acceptable data has been filed with Animal and Plant Health In spection Service, to show that develop ment of corneal opacity is not associ ated with the product, carton labels and enclosures used with biological products containing modified live ca nine hepatitis virus or modified live ca nine adenovirus Type 2 shall bear the following statement: ‘‘Occasionally, transient corneal opacity may occur following the administration of this product.’’ (l) All labels for autogenous biologics must specify the name of the micro organism(s) or antigen(s) that they contain, and shall bear the following statement: ‘‘Potency and efficacy of autogenous biologics have not been es tablished. This product is prepared for use only by or under the direction of a veterinarian or approved specialist.’’ (m) In the case of biological products containing Marek’s disease virus, all labels shall specify the Marek’s disease virus serotype(s) used in the product. (n) All labels for conditionally li censed products shall bear the fol lowing statement: ‘‘This product li cense is conditional; efficacy and po tency have not been fully dem onstrated.’’ (Approved by the Office of Management and Budget under control number 0579–0013) [38 FR 12094, May 9, 1973] EDITORIAL NOTE: For FEDERAL REGISTER ci tations affecting § 112.7, see the List of CFR Sections Affected, which appears in the Finding Aids section of the printed volume and at www.fdsys.gov. § 112.8 For export only. The applicable regulations for pack aging and labeling a biological product produced in the United States shall apply to such biological product if ex ported from the United States except as otherwise provided in this section. Only labels approved as provided in § 112.5 shall be used. (a) Biological products which have been packaged and labeled for export or which have been exported, shall be subject to the applicable provisions in this paragraph. (1) After leaving the licensed estab lishment, a biological product shall not be bottled, repackaged, relabeled, or otherwise altered in any way while in the United States; and (2) An exported biological product shall not be returned to the United States: Provided, That, in the case of a biological product exported in labeled VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00706 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 697 Animal and Plant Health Inspection Service, USDA Pt. 113 final containers, the Administrator may authorize by permit the importa tion of a limited number for research and evaluation by the producing li censee; and (3) An exported biological product which is bottled, rebottled, or altered in any way in a foreign country shall not bear a label which indicates by es tablishment license number that it has been prepared in the United States. (b) Desiccated and frozen liquid prod ucts, packaged and labeled as for do mestic use, may be exported without the diluent required for rehydration or dilution, as the case may be, if the la beling includes adequate instructions for preparing the product for use and the words ‘‘For Export Only’’. (c) Final containers of products, la beled or unlabeled, may be exported in sealed shipping boxes, adequately iden tified as to contents with an approved label, and plainly marked ‘‘For Export Only’’: Provided, That such products shall not be diverted to domestic use. (d) Completed inactivated liquid products, antiserums, and antitoxins, may be exported in large multiple-dose containers identified with an approved label that contains the words ‘‘For Ex port Only’’ prominently displayed. (e) Concentrated inactivated liquid product, completed except for dilution to the proper strength for use, may be exported in large multiple-dose con tainers identified with an approved label that contains the words ‘‘For Ex port Only’’ prominently displayed. [38 FR 12094, May 9, 1973, as amended at 39 FR 19202, May 31, 1974; 40 FR 46093, Oct. 6, 1975; 43 FR 11145, Mar. 17, 1978; 56 FR 66784, Dec. 26, 1991] § 112.9 Biological products imported for research and evaluation. A biological product imported for re search and evaluation under a permit issued in accordance with § 104.4, with the exception of products imported under § 104.4(d), shall be labeled as pro vided in this section. (a) The label shall identify the prod uct and the name and address of the manufacturer and shall provide in structions for proper use of the prod uct, including all warnings and cau tions needed by the permittee to safely use the product. (b) Labels on each product to be fur ther distributed in accordance with § 103.3 shall bear the statement ‘‘No tice! For Experimental Use Only—Not for Sale!’’ (c) The labeling shall contain any other information deemed necessary by the Administrator and specified on the permit. [50 FR 46417, Nov. 8, 1985, as amended at 56 FR 66784, Dec. 26, 1991] § 112.10 Special packaging and label ing. A biological product, which requires special packaging and/or labeling not provided for in this part, shall be pack aged and/or labeled in accordance with requirements written into the approved outline for such product. PART 113—STANDARD REQUIREMENTS APPLICABILITY Sec. 113.1 Compliance. 113.2 Testing aids. 113.3 Sampling of biological products. 113.4 Exemptions to tests. 113.5 General testing. 113.6 Animal and Plant Health Inspection Service testing. 113.7 Multiple fractions. 113.8 In vitro tests for serial release. 113.9 New potency test. 113.10 Testing of bulk material for export or for further manufacture. STANDARD PROCEDURES 113.25 Culture media for detection of bac teria and fungi. 113.26 Detection of viable bacteria and fungi except in live vaccine. 113.27 Detection of extraneous viable bac teria and fungi in live vaccines. 113.28 Detection of mycoplasma contamina tion. 113.29 Determination of moisture content in desiccated biological products. 113.30 Detection of Salmonella contamina tion. 113.31 Detection of avian lymphoid leukosis. 113.32 Detection of Brucella contamination. 113.33 Mouse safety tests. 113.34 Detection of hemagglutinating vi ruses. 113.35 Detection of viricidal activity. 113.36 Detection of pathogens by the chick en inoculation test. 113.37 Detection of pathogens by the chick en embryo inoculation test. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00707 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 698 9 CFR Ch. I (1–1–18 Edition) Pt. 113 113.38 Guinea pig safety test. 113.39 Cat safety tests. 113.40 Dog safety tests. 113.41 Calf safety test. 113.42 Detection of lymphocytic chorio meningitis contamination. 113.43 Detection of chlamydial agents. 113.44 Swine safety test. 113.45 Sheep safety test. 113.46 Detection of cytopathogenic and/or hemadsorbing agents. 113.47 Detection of extraneous viruses by the fluorescent antibody technique. INGREDIENT REQUIREMENTS 113.50 Ingredients of biological products. 113.51 Requirements for primary cells used for production of biologics. 113.52 Requirements for cell lines used for production of biologics. 113.53 Requirements for ingredients of ani mal origin used for production of bio logics. 113.54 Sterile diluent. 113.55 Detection of extraneous agents in Master Seed Virus. LIVE BACTERIAL VACCINES 113.64 General requirements for live bac terial vaccines. 113.65 Brucella Abortus Vaccine. 113.66 Anthrax Spore Vaccine—Nonencap sulated. 113.67 Erysipelothrix Rhusiopathiae Vac cine. 113.68 Pasteurella Haemolytica Vaccine, Bovine. 113.69 Pasteurella Multocida Vaccine, Bo vine. 113.70 Pasteurella Multocida Vaccine, Avian Isolate. 113.71 Chlamydia Psittaci Vaccine (Feline Pneumonitis), Live Chlamydia. INACTIVATED BACTERIAL PRODUCTS 113.100 General requirements for inac tivated bacterial products. 113.101 Leptospira Pomona Bacterin. 113.102 Leptospira Icterohaemorrhagiae Bacterin. 113.103 Leptospira Canicola Bacterin. 113.104 Leptospira Grippotyphosa Bacterin. 113.105 Leptospira Hardjo Bacterin. 113.106 Clostridium Chauvoei Bacterin. 113.107 Clostridium Haemolyticum Bacterin. 113.108 Clostridium Novyi Bacterin-Toxoid. 113.109 Clostridium Sordellii Bacterin-Tox oid. 113.110 Clostridium Botulinum Type C Bacterin-Toxoid. 113.111 Clostridium Perfringens Type C Tox oid and Bacterin-Toxoid. 113.112 Clostridium Perfringens Type D Tox oid and Bacterin-Toxoid. 113.113 Autogenous biologics. 113.114 Tetanus Toxoid. 113.115 Staphylococcus Aureus Bacterin Toxoid. 113.116 Pasteurella Multocida Bacterin, Avian Isolate, Type 4. 113.117 Pasteurella Multocida Bacterin, Avian Isolate, Type 1. 113.118 Pasteurella Multocida Bacterin, Avian Isolate, Type 3. 113.119 Erysipelothrix Rhusiopathiae Bacterin. 113.120 Salmonella Typhimurium Bacterin. 113.121 Pasteurella Multocida Bacterin. 113.122 Salmonella Choleraesuis Bacterin. 113.123 Salmonella Dublin Bacterin. KILLED VIRUS VACCINES 113.200 General requirements for killed virus vaccines. 113.201–203 [Reserved] 113.204 Mink Enteritis Vaccine, Killed Virus. 113.205 Newcastle Disease Vaccine, Killed Virus. 113.206 Wart Vaccine, Killed Virus. 113.207 Encephalomyelitis Vaccine, Eastern, Western, and Venezuelan, Killed Virus. 113.208 Avian Encephalomyelitis Vaccine, Killed Virus. 113.209 Rabies Vaccine, Killed Virus. 113.210 Feline Calicivirus Vaccine, Killed Virus. 113.211 [Reserved] 113.212 Bursal Disease Vaccine, Killed Virus. 113.213–113.214 [Reserved] 113.215 Bovine Virus Diarrhea Vaccine, Killed Virus. 113.216 Bovine Rhinotracheitis Vaccine, Killed Virus. LIVE VIRUS VACCINES 113.300 General requirements for live virus vaccines. 113.301 Ovine Ecthyma Vaccine. 113.302 Distemper Vaccine—Mink. 113.303 Bluetongue Vaccine. 113.304 Feline Panleukopenia Vaccine. 113.305 Canine Hepatitis and Canine Adenovirus Type 2 Vaccine. 113.306 Canine Distemper Vaccine. 113.308 Encephalomyelitis Vaccine, Ven ezuelan. 113.309 Bovine Parainfluenza3 Vaccine. 113.310 Bovine Rhinotracheitis Vaccine. 113.311 Bovine Virus Diarrhea Vaccine. 113.312 Rabies Vaccine, Live Virus. 113.313 Measles Vaccine. 113.314 Feline Calicivirus Vaccine. 113.315 Feline Rhinotracheitis Vaccine. 113.316 Canine Parainfluenza Vaccine. 113.317 Parvovirus Vaccine (Canine). 113.318 Pseudorabies Vaccine. 113.319–113.324 [Reserved] 113.325 Avian Encephalomyelitis Vaccine. 113.326 Avian Pox Vaccine. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00708 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 699 Animal and Plant Health Inspection Service, USDA § 113.3 113.327 Bronchitis Vaccine. 113.328 Fowl Laryngotracheitis Vaccine. 113.329 Newcastle Disease Vaccine. 113.330 Marek’s Disease Vaccines. 113.331 Bursal Disease Vaccine. 113.332 Tenosynovitis Vaccine. DIAGNOSTICS AND REAGENTS 113.400–113.405 [Reserved] 113.406 Tuberculin, Intradermic. 113.407 Pullorum antigen. 113.408 Avian mycoplasma antigen. 113.409 Tuberculin—PPD Bovis, Intradermic. ANTIBODY PRODUCTS 113.450 General requirements for antibody products. 113.451 Tetanus Antitoxin. 113.452 Erysipelothrix Rhusiopathiae Anti body. 113.453 [Reserved] 113.454 Clostridium Perfringens Type C Antitoxin. 113.455 Clostridium Perfringens Type D Antitoxin. 113.456–113.498 [Reserved] 113.499 Products for treatment of failure of passive transfer. AUTHORITY: 21 U.S.C. 151–159; 7 CFR 2.22, 2.80, and 371.4. SOURCE: 34 FR 18004, Nov. 7, 1969, unless otherwise noted. EDITORIAL NOTE: Nomenclature changes to part 113 appear at 79 FR 55969, Sept. 18, 2014. APPLICABILITY § 113.1 Compliance. The regulations in this part apply to each serial or subserial of a licensed bi ological product manufactured in a li censed establishment and to each serial or subserial of a biological product in each shipment imported for distribu tion and sale. § 113.2 Testing aids. To better ensure consistent and re producible test results when Standard Requirement tests prescribed in the regulations are conducted, National Veterinary Services Laboratories, U.S. Department of Agriculture, may pro vide testing aids, when available, to li censees, permittees, and applicants for licenses and permits. Such aids shall be as follows: (a) Supplemental Assay Method (SAM) is a technical bulletin con taining detailed instructions for con ducting a test. Such instructions shall be in accordance with the procedures currently being followed at National Veterinary Services Laboratories and as improved, proven procedures are de veloped, shall be revised and reissued prior to application. (b) Standard Reference Preparation is a serum, virus, bacterial culture, or antigen to be used in test systems for direct comparison with serials of bio logical products under test. (c) Standard Test Reagent is a serum, antitoxin, fluorescent antibody con jugate, toxin, virus, bacterial cultural, or antigen to be used in test systems but not for direct comparison with se rials of biological products under test. (d) Seed cultures are small quantities of standard organisms to be propagated by the recipient to establish a supply for use. (e) Test Code Number is a number as signed by Animal and Plant Health In spection Service to each test procedure specified in the Standard Requirements and in each filed Outline of Production where such test is conducted to support a request for release of a serial or sub serial. [39 FR 21041, June 18, 1974, as amended at 40 FR 758, Jan. 3, 1975; 50 FR 21799, May 29, 1985; 56 FR 66784, Dec. 26, 1991] § 113.3 Sampling of biological prod ucts. Each licensee and permittee shall furnish representative samples of each serial or subserial of a biological prod uct manufactured in the United States or imported into the United States as prescribed in this section. Additional samples may be purchased in the open market by a Animal and Plant Health Inspection Service representative. (a) Either an employee of the Depart ment of Agriculture, of the licensee, or of the permittee, as designated by the Administrator shall select prerelease samples of biological product in the number prescribed in paragraph (b) of this section. Each sample shall be marked for identification by the person making the selection after which they shall be packaged by the licensee or permittee, as the case may be, and for warded to National Veterinary Serv ices Laboratories; except that an em ployee of the Department may forward VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00709 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 700 9 CFR Ch. I (1–1–18 Edition) § 113.3 or deliver the samples to National Vet erinary Services Laboratories if such action deemed advisable by the Admin istrator. (1) Selection shall be made as fol lows: (i) Nonviable liquid biological prod ucts—either bulk or final container samples of completed product shall be selected for purity, safety, or potency tests. Biological product in final con tainer shall be selected to test for via ble bacteria and fungi. (ii) Viable liquid biological products; samples shall be in final containers and shall be randomly selected at the end of the filling operation. Bulk con tainers of completed product may be sampled when authorized by the Ad ministrator. (iii) Desiccated biological products; samples shall be in final containers and shall be randomly selected if des iccated in the final container. Biologi cal products desiccated in bulk shall be sampled at the end of the filling oper ation. (iv) Representative samples of each serial or subserial in each shipment of imported biological products shall be selected. (2) Comparable samples shall be used by Animal and Plant Health Inspection Service, the licensee, and the per mittee for similar tests. (3) When bulk samples of completed product in liquid form are to be tested as prescribed in paragraph (a)(1) of this section, the number of such samples from each serial and the minimum quantity of product to be provided in each sample shall be stated in the filed Outline of Production. (b) Unless otherwise prescribed by the Administrator, the number of final container samples to be selected from each serial and subserial shall be: (1) Vaccines: (i) Six multiple-dose samples of Brucella Abortus Vaccine; (ii) Twelve samples of all other live bacterial vaccines; (iii) Two samples of Coccidiosis Vac cine; (iv) Eighteen samples of Rabies Vac cine, Modified Live Virus; (v) Sixteen samples of all other vac cines consisting of live microorga nisms; (vi) Thirty single-dose or 14 multiple dose samples of Equine Encephalomyelitis Vaccine, Killed Virus; (vii) Twenty-two single-dose or 14 multiple-dose samples of Rabies Vac cine, Killed Virus; (viii) Sixteen single-dose or 12 mul tiple-dose samples of all other vaccines consisting of killed microorganisms. (2) Bacterins and bacterin-toxoids: (i) Twelve samples of single-fraction products; (ii) Thirteen samples of two-fraction products; (iii) Fourteen samples of products consisting of 3 or more fractions. (3) Antiserums: Twelve samples of antiserum recommended for large ani mals or 14 samples of antiserum rec ommended for small animals or the number of reagent serum samples pre scribed in the filed Outline of Produc tion for the product. (4) Antitoxins: (i) Fourteen single-dose or 12 mul tiple dose samples of Tetanus Anti toxin; (ii) Twelve samples of all other antitoxins. (5) Toxoids: (i) Eighteen single-dose or 12 mul tiple dose samples of all toxoids. (6) Antigens: Twelve samples of poul try antigens or 20 samples of tuber culin or four samples of all other diag nostic antigens. (7) Diagnostic test kits: Two samples of diagnostic test kits. The licensee or permittee will hold one of these se lected samples at the storage tempera ture recommended on the label while awaiting a request by the animal and Plant Health Inspection Service to sub mit the additional sample. If submis sion is not requested by the Animal and Plant Health Inspection Service, the additional sample may be returned to the serial inventory after the serial is released. In the case of diagnostic test kits in which final packaging con sists of multiple microtiter test plates or strips, the licensee or permittee may submit a specified number of test plates or strips along with all other test reagents as prescribed in a filed Outline of Production and retain a similar amount as a second sample for VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00710 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 701 Animal and Plant Health Inspection Service, USDA § 113.3 submission upon request. When the ini tial sample is not representative of final packaging by the licensee of per mittee, e.g., does not consist of all the microtiter test plates or strips, the sec ond sample is not eligible to be re turned to serial inventory after the se rial is released. (8) Autogenous biologics: With the ex ception of the first serial or subserial, 10 samples must be selected and sub mitted to the Animal and Plant Health Inspection Service from each serial or subserial of an autogenous biologic eli gible to be shipped that consists of more than 50 containers. For first seri als or subserials eligible for shipment consisting of more than 50 containers, 10 samples from each serial or subserial must be selected and held for submis sion to the Animal and Plant Health Inspection Service upon request in ac cordance with paragraph (e)(4) of this section. For serials or subserials of au togenous biologic with 50 or fewer con tainers, no samples, other than those required by paragraph (e) of this sec tion, are required. (9) Miscellaneous: The number of sam ples from products not in the cat egories provided for in paragraphs (b)(1) through (b)(8) of this section shall be prescribed in the filed Outline of Production for the product. (c) Prelicensing and Outline of Pro duction changes: Samples needed to support a license application or a change in the Outline of Production for a licensed product shall be submitted only upon request from the animal and Plant Health Inspection Service. Ex cept for miscellaneous products speci fied in paragraph (b)(9) of this section, the number of such samples shall be at least one and one-half times the num ber prescribed for such product in para graph (b) of this section. Samples of Master Seeds and Master Cell Stocks with a minimum individual volume of 1 ml shall be submitted as follows: (1) Ten samples of Bacterial Master Seeds. (2) Thirteen samples of viral Master Seeds or nonviral Master Seeds requir ing cell culture propagation. For Mas ter Seeds isolated or passed in a cell line different from the species of in tended use, an additional 2 samples are required for each additional species. For Master Seeds grown in cell culture and intended for use in more than one species, an additional 2 samples are re quired for each additional species. (3) Thirty-six samples of at least 1 ml each or six samples of at least 1 ml each, one sample of at least 20 ml, and one sample of at least 10 ml of Master Cell Stocks. In the case of Master Cell Stocks which are persistently infected with a virus, an additional four sam ples of at least 1 ml each are required. If these persistently infected cell stocks are intended for use in more than one species, an additional two samples of at least 1 ml each are re quired for each additional species. (4) Four samples of the Master Cell Stock + n (highest passage) cells. (d) Sterile diluent: A sample of Ster ile Diluent shall accompany each sam ple of product, other than Marek’s Dis ease Vaccine, if such diluent is re quired to rehydrate or dilute the prod uct before use. The volume of diluent shall be an appropriate amount to re hydrate or dilute the product. Samples of Sterile Diluent prepared for use with Marek’s Disease Vaccine shall be sub mitted upon request from the Animal and Plant Health Inspection Service. (e) Reserve samples shall be selected from each serial and subserial of bio logical product. Such samples shall be selected at random from final con tainers of completed product by an em ployee of the Department, of the li censee, or of the permittee, as des ignated by the administrator. Each sample shall: (1) Consist of 5 single-dose packages, 2 multiple-dose packages, or 2 diag nostic test kits, except that, in the case of diagnostic test kits in which final packaging consists of multiple microtiter test plates or strips, a sam ple may consist of a specified number of test plates or strips along with all other test reagents as prescribed in a filed Outline of Production; (2) Be adequate in quantity for appro priate examination and testing; (3) Be truly representative and in final containers; (4) Be held in a special compartment set aside by the licensee or permittee for holding these samples under refrig eration at the storage temperature rec ommended on the labels for 6 months VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00711 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 702 9 CFR Ch. I (1–1–18 Edition) § 113.4 after the expiration date stated on the labels. The samples that are stored in this manner shall be delivered to the Animal and Plant Health Inspection Service upon request. (Approved by the Office of Management and Budget under control number 0579–0013) [38 FR 29886, Oct. 30, 1973, as amended at 40 FR 758, Jan. 3, 1975; 40 FR 49768, Oct. 24, 1975; 41 FR 56627, Dec. 29, 1976; 48 FR 9506, Mar. 7, 1983; 48 FR 57473, Dec. 30, 1983; 50 FR 21799, May 29, 1985; 56 FR 66784, Dec. 26, 1991; 60 FR 14356, Mar. 17, 1995; 67 FR 15713, Apr. 3, 2002] § 113.4 Exemptions to tests. (a) The test methods and procedures contained in all applicable Standard Requirements shall be complied with unless otherwise exempted by the Ad ministrator and provided that such ex emption is noted in the filed Outline of Production for the product. (b) Test methods and procedures by which the biological products shall be evaluated shall be designated in the Outline of Production for such prod ucts. [38 FR 29887, Oct. 30, 1973, as amended at 56 FR 66784, Dec. 26, 1991] § 113.5 General testing. (a) No biological product shall be re leased prior to the completion of tests prescribed in a filed Outline of Produc tion or Standard Requirements for the product to establish the product to be pure, safe, potent, and efficacious. (b) Tests of biological products shall be observed by a competent employee of the manufacturer during all critical periods. A critical period shall be the time when certain specified reactions must occur in required tests to prop erly evaluate the results. (c) Records of all tests shall be kept in accordance with part 116 of this chapter. Results of all required tests prescribed in the filed Outline of Pro duction or the Standard Requirements for the product shall be submitted to Animal and Plant Health Inspection Service. Blank forms shall be furnished upon request to Animal and Plant Health Inspection Service. (d) When the initial or any subse quent test is declared a No Test, the reasons shall be reported in the test records, the results shall not be consid ered as final, and the test may be re peated. When a test is declared satis factory, the test designation is consid ered to be a final conclusion. When a test is declared unsatisfactory, the test designation is considered to be a final conclusion. When the initial or any subsequent test is declared inconclu sive, the reasons shall be reported in the test records, the result shall not be considered as final, and the test may be repeated as established in the filed Outline of Production or Standard Re quirement. If a test is designated in conclusive or No Test and the biologi cal product is not further tested, the test designation of unsatisfactory is the final conclusion. (e) When new test methods are devel oped and approved by Animal and Plant Health Inspection Service, bio logical products tested thereafter shall be evaluated by such methods, and if not found to be satisfactory when so tested shall not be released. (Approved by the Office of Management and Budget under control number 0579–0059) [34 FR 18004, Nov. 4, 1969, as amended at 39 FR 25463, July 11, 1974; 40 FR 45420, Oct. 2, 1975; 40 FR 46093, Oct. 6, 1975; 41 FR 6751, Feb. 13, 1976; 48 FR 57473, Dec. 30, 1983; 56 FR 66784, Dec. 26, 1991; 79 FR 55969, Sept. 18, 2014] § 113.6 Animal and Plant Health In spection Service testing. A biological product shall with rea sonable certainty yield the results in tended when used as recommended or suggested in its labeling or proposed la beling prior to the expiration date. (a) The Administrator is authorized to cause a biological product, manufac tured in the United States or imported into the United States, to be examined and tested for purity, safety, potency, or efficacy; in which case, the licensee or permittee shall withhold such prod uct from the market until a determina tion has been made. (b) The final results of each test con ducted by the licensee and Animal and Plant Health Inspection Service shall be considered in evaluating a biological product. A serial or subserial which has been found unsatisfactory by a re quired test prescribed in a filed Outline VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00712 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 703 Animal and Plant Health Inspection Service, USDA § 113.8 of Production or Standard Require ment is not in compliance with the reg ulations and shall not be released for market. [34 FR 18004, Nov. 7, 1969, as amended at 40 FR 45420, Oct. 2, 1975; 40 FR 53378, Nov. 18, 1975; 41 FR 6751, Feb. 13, 1976; 56 FR 66784, Dec. 26, 1991] § 113.7 Multiple fractions. (a) When a biological product con tains more than one immunogenic frac tion, the completed product shall be evaluated by tests applicable to each fraction. (b) When similar potency tests are re quired for more than one fraction of a combination biological product, dif ferent animals must be used to evalu ate each fraction except when written Standard Requirements or outlines of production make provisions and set forth conditions for use of the same animals for testing different fractions. (c) When the same safety test is re quired for more than one fraction, re quirements are fulfilled by satisfactory results from one test of the completed product. (d) When an inactivated fraction(s) is used as a diluent for a live virus frac tion(s), the inactivated fraction(s) may be tested separately and the live virus fraction(s) may be tested separately: Provided, That, the viricidal test re quirements prescribed in § 113.100 are complied with. (e) Virus titrations for a multivirus product shall be conducted by methods which will quantitate each virus. [34 FR 18004, Nov. 7, 1969, as amended at 40 FR 46093, Oct. 6, 1975; 56 FR 66785, Dec. 26, 1991] § 113.8 In vitro tests for serial release. (a) Master Seed which has been es tablished as pure, safe, and immunogenic shall be used for pre paring seed for production as specified in the Standard Requirements or in the filed Outline of Production. The Ad ministrator may exempt a product from a required animal potency test for release when an evaluation can, with reasonable certainty, be made by: (1) Subjecting the master seed to the applicable requirements prescribed in §§ 113.64, 113.100, 113.200, and 113.300; (2) Testing the Master Seed for immunogenicity in a manner accept able to the Animal and Plant Health Inspection Service (APHIS); (3) Establishing satisfactory potency for the product in accordance with the following provisions: (i) Potency for live products may be determined by log10 virus titer or de termining the live bacterial count based on the protective dose used in the Master Seed immunogenicity test plus an adequate overage for adverse conditions and test error; and (ii) Potency for inactivated products may be determined using tests for rel ative antigen content by comparing the antigen content of the test serial to a reference preparation using a par allel line immunoassay or equivalent method which measures linearity, specificity, and reproducibility in a manner acceptable to APHIS. (b) In the case of live products, each serial and subserial of desiccated prod uct derived from an approved Master Seed and bulk or final container sam ples of each serial of completed liquid product derived from an approved Mas ter Seed shall be evaluated by a test procedure acceptable to APHIS. On the basis of the results of the test, as com pared with the required minimum po tency, each serial and subserial shall either be released to the firm for mar keting or withheld from the market. The evaluation of such products shall be made in accordance with the fol lowing criteria: (1) If the initial test shows the count or titer to equal or exceed the required minimum, the serial or subserial is sat isfactory without additional testing. (2) If the initial test shows the count or titer to be lower than the required minimum, the serial or subserial may be retested, using double the number of samples. The average counts or titers obtained in the retests shall be deter mined. If the average is less than the required minimum, the serial or sub serial is unsatisfactory without further consideration. (3) If the average is equal to or great er than the required minimum, the fol lowing shall apply to live virus vac cines: (i) If the difference between the aver age titer obtained in the retests and VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00713 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 704 9 CFR Ch. I (1–1–18 Edition) § 113.8 the titer obtained in the initial test is 10 0.7 or greater, the initial titer may be considered a result of test system error and the serial or subserial considered satisfactory for virus titer. (ii) If the difference between the av erage titer obtained in the retests and the titer obtained in the initial test is less than 10 0.7, a new average shall be determined using the titers obtained in all tests. If the new average is below the required minimum, the serial or subserial is unsatisfactory. (4) If the average is equal to or great er than the required minimum, the fol lowing shall apply to bacterial vac cines: (i) If the average count obtained in the retests is at least three times the count obtained in the initial test, the initial count may be considered a re sult of test system error and the serial or subserial considered satisfactory for bacterial count. (ii) If the average count obtained in the retests is less than three times the count obtained in the initial test, a new average shall be determined using the counts obtained in all tests. If the new average count is below the re quired minimum, the serial or sub serial is unsatisfactory. (5) Exceptions. When a product is eval uated in terms other than log10 virus titer or organism count, an appropriate difference between the average potency value obtained in the retests and the potency value obtained in the initial test shall be established for use in paragraphs (b)(3) and (b)(4) of this sec tion to evaluate such products and shall be specified in the product Stand ard Requirement or filed Outline of Production. (c) In the case of inactivated prod ucts, bulk or final container samples of completed product from each serial de rived from an approved Master Seed, shall be evaluated for relative antigen content (potency) as compared with an unexpired reference by a parallel line immunoassay or other procedure ac ceptable to APHIS. Firms currently using immunoassays which do not sat isfy this requirement shall have 2 years from the effective date of the final rule to update their filed Outlines of Pro duction to be in compliance with this requirement unless granted an exten sion by the Administrator based on a showing by the firm seeking the exten sion that they have made a good faith effort with due diligence to achieve compliance. On the basis of the results of such test procedures, each serial that meets the required minimum po tency shall be released to the firm for marketing; each serial not meeting the required minimum potency shall be withheld from the market. The evalua tion of such products shall be made in accordance with the following criteria: (1) A test that results in no valid lines is considered a ‘‘no test’’ and may be repeated. (2) An initial test (test 1) that results in valid lines that are not parallel is considered a valid equivocal test. Re lease of the serial may not be based on such test since the result cannot be termed ‘‘satisfactory’’ or ‘‘unsatisfac tory.’’ (3) If the initial test (test 1) shows that potency equals or exceeds the re quired minimum potency, the serial is satisfactory without additional test ing. (4) If the initial test (test 1) is an equivocal test due to lack of par allelism, the serial may be retested up to three times (tests 2, 3, and 4) with disposition to be as specified in para graphs (c)(4)(i) and (ii) of this section; Provided, That, if the serial is not re tested or the other provisions of this section are not satisfied, the serial shall be deemed unsatisfactory. (i) If: The first retest (test 2) fol lowing an initial equivocal test; the second retest (test 3) following two consecutive equivocal tests (tests 1 and 2); or the third retest (test 4) following three consecutive equivocal tests (tests 1, 2, and 3) shows that the potency equals or exceeds the required min imum potency, the serial is satisfac tory. (ii) If the first retest (test 2) fol lowing an initial equivocal test shows that potency is less than the required minimum potency, disposition of the serial will be based on the outcome of retests 2 and 3 (tests 3 and 4) as follows: if either retest (test 3 or 4) shows that potency is less than the required min imum potency, the serial is unsatisfac tory. If either retest 2 or retest 3 (tests 3 or 4) is an equivocal test, or in the VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00714 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 705 Animal and Plant Health Inspection Service, USDA § 113.8 event that each retest (tests 2, 3, and 4) following an initial equivocal test is also an equivocal test, the accumulated test results shall be considered indic ative of a lack of potency and release of the serial withheld. In which case, the licensee may submit data con firming the continued validity of the test system to APHIS for review and approval. If the data are acceptable to APHIS, the potency test may be re peated by the firm, subject to the pro visions specified in paragraphs (i) and (ii) and confirmatory testing by APHIS. (5) If the initial test (test 1) shows that potency is less than the required minimum potency, the serial may be retested a minimum of two times (tests 2 and 3) but not more than three times (tests 2, 3, and 4) with disposition as specified in paragraphs (c)(5) (i) and (ii) of this section; Provided, That, if the serial is not retested or the other pro visions of this section are not satisfied, the serial shall be deemed unsatisfac tory. (i) If two consecutive retests (tests 2 and 3) show that potency of the serial equals or exceeds the required min imum potency, the serial is satisfac tory. If one of the two retests (test 2 or 3) shows that the potency is less than the required minimum potency, the se rial is unsatisfactory. (ii) If one of the retests (tests 2 or 3) shows that the potency equals or ex ceeds the required minimum potency and the other retest (test 2 or 3) is an equivocal test, a third retest (test 4) may be performed. If the third retest (test 4) shows that the potency of the serial equals or exceeds the required minimum potency, the serial is deemed satisfactory. If both retests (tests 2 and 3) or if the third retest (test 4) is an equivocal test, the accumulated test results shall be considered indicative of a lack of potency and release of the se rial withheld, in which case the li censee may submit data confirming the continued validity of the test system to APHIS for review and approval. If the data are acceptable to APHIS, the potency test may be repeated by the firm, subject to the provisions specified in paragraphs (c)(4) (i) and (ii) and (c)(5) (i) and (ii) of this section, and confirmatory testing by APHIS. (d) Extending the dating of a reference. All determinations of relative antigen content using parallel line immunoassays or equivalent methods shall be conducted with an unexpired reference. The lot of reference used to determine antigenic content shall have an initial dating period equal to the dating of the product or as supported by data acceptable to APHIS, except that frozen references may have an ini tial dating of up to 5 years, Provided, That the request for dating of the fro zen references beyond the dating of the product is supported by preliminary data acceptable to APHIS and includes provisions for monitoring the stability of the reference to determine when the potency starts to decline and for tak ing the appropriate steps to requalify a reference with declining potency either by testing a Qualifying Serial in host animals or by providing other evidence of immunogenicity, e.g., antibody titers or laboratory animal test data previously correlated to host animal protection in a manner acceptable to APHIS. Prior to the expiration date, such reference may be granted an ex tension of dating, Provided, That its immunogenicity has been confirmed using a Qualifying Serial of product in a manner acceptable to APHIS. The dating period of the Master Reference and Working Reference may be ex tended by data acceptable to APHIS if the minimum potency of the Master Reference is determined to be ade quately above the minimum level need ed to provide protection in the host animal. If a new Master Reference is established, it shall be allowed an ini tial dating period equal to the dating of the product or as supported by data acceptable to APHIS, except that fro zen references may have an initial dat ing period of 5 years, or as supported by data acceptable to APHIS. Prior to the expiration date, such reference may be granted an extension of dating by confirming its immunogenicity using a Qualifying Serial of product. (e) Final container samples of com pleted product derived from Master Seed found immunogenic in accordance with paragraph (a) of this section and found satisfactory in accordance with paragraphs (b) and (c) of this section VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00715 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 706 9 CFR Ch. I (1–1–18 Edition) § 113.9 may also be subjected to an animal po tency test by Animal and Plant Health Inspection Service as provided in this paragraph. Products shall be used ac cording to label directions including dose(s) and route of administration. (1) A one stage test using 20 vac cinates and 5 controls or a two stage test using 10 vaccinates and 5 controls for each stage shall be used. The cri teria used for judging the specific re sponse in the controls and vaccinates shall be in accordance with the test protocol used in the Master Seed immunogenicity test. (2) If at least 80 percent of the con trols do not show specific responses to challenge, the test is inconclusive and may be repeated. If a vaccinate shows the specific responses to challenge ex pected in the controls, the vaccinate shall be listed as a failure. (3) The results of the testing shall be evaluated according to the following table: CUMULATIVE TOTALS Stage Num ber of ani mals Failures for satisfactory serials Failures for unsatisfac tory serials 1 ............................. 10 1 or less ...... 3 or more. 2 (or 1) ................... 20 4 or less ...... 5 or more. (4) When a serial has been found un satisfactory for potency by the test provided in paragraphs (e)(1), (2), and (3) of this section, the serial shall be withheld from the market and the fol lowing actions taken: (i) The Administrator shall require that at least two additional serials pre pared with the same Master Seed be subjected to similar animal potency tests by Animal and Plant Health In spection Service or the licensee or both. (ii) If another serial is found unsatis factory for potency, the product shall be removed from the market while a reevaluation of the product is made and the problem is resolved. [49 FR 22625, May 31, 1984, as amended at 56 FR 66784, 66786, Dec. 26, 1991; 62 FR 19038, Apr. 18, 1997; 72 FR 72564, Dec. 21, 2007; 79 FR 31021, May 30, 2014] § 113.9 New potency test. A potency test written into the filed Outline of Production for a product shall be considered confidential infor mation by Animal and Plant Health In spection Service until at least two ad ditional product licenses are issued for the product or unless use of the test is authorized by the licensee, in which case, such potency test may be pub lished as part of the Standard Require ment for the product. (a) Until a potency test is published as part of the Standard Requirement for the product, reference to such a test shall be made in the filed Outline of Production and the test shall be con ducted. (b) When a potency test has been pub lished as part of the Standard Require ment, such test shall be conducted un less the product is specifically exempt ed as provided in § 113.4. [40 FR 14084, Mar. 28, 1975, as amended at 56 FR 66784, Dec. 26, 1991] § 113.10 Testing of bulk material for export or for further manufacture. When a product is prepared in a li censed establishment for export in large multiple-dose containers as pro vided in § 112.8(d) or (e) of this sub chapter or for further manufacturing purposes as provided in § 114.3(d) of this subchapter, samples of the bulk mate rial shall be subjected to all required tests prescribed in the filed Outline of Production or Standard Requirements for the product. Samples of con centrated liquid product shall be di luted to a volume equal to the contents of the sample times the concentration factor prior to initiating potency tests. [49 FR 45846, Nov. 21, 1984] STANDARD PROCEDURES § 113.25 Culture media for detection of bacteria and fungi. (a) Ingredients for which standards are prescribed in the United States Pharmacopeia, or elsewhere in this part, shall conform to such standards. In lieu of preparing the media from the individual ingredients, they may be made from dehydrated mixtures which, when rehydrated with purified water, VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00716 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 707 Animal and Plant Health Inspection Service, USDA § 113.26 have the same or equivalent composi tion as such media and have growth promoting buffering, and oxygen ten sion-controlling properties equal to or better than such media. The formulas for the composition of the culture media prescribed in §§ 113.26 and 113.27 are set forth in the United States Phar macopeia, 19th Edition. (b) The licensee shall test each quan tity of medium prepared at one time from individual ingredients and the first quantity prepared from each lot of commercial dehydrated medium for growth-promoting qualities. If any por tion of a lot of commercial dehydrated medium is held for 90 days or longer after being so tested, it shall be re tested before use. Two or more strains of micro-organisms that are exacting in their nutritive requirements shall be used. More than one dilution shall be used to demonstrate the adequacy of the medium to support the growth of a minimum number of micro-organisms. (c) The sterility of the medium shall be confirmed by incubating an ade quate number of test vessels and exam ining each for growth. Additional con trol may be used by incubation of rep resentative uninoculated test vessels for the required incubation period dur ing each test. (d) A determination shall be made by the licensee for each biological product of the ratio of inoculum to medium which shall result in sufficient dilution of such product to prevent bacteriostatic and fungistatic activity. The determination may be made by tests on a representative biological product for each group of comparable products containing identical preserva tives at equal or lower concentrations. Inhibitors or neutralizers of preserva tives, approved by the Administrator, may be considered in determining the proper ratio. [35 FR 16039, Oct. 13, 1970, as amended at 37 FR 2430, Feb. 1, 1972; 41 FR 27715, July 6, 1976; 56 FR 66784, Dec. 26, 1991] § 113.26 Detection of viable bacteria and fungi except in live vaccine. Each serial and subserial of biologi cal product except live vaccines shall be tested as prescribed in this section unless otherwise specified by the Ad ministrator. When cell lines, primary cells, or ingredients of animal origin used in the preparation of a biological product are required to be free of via ble bacteria and fungi, they shall also be tested as prescribed in this section. (a) The media to be used shall be as follows: (1) Fluid Thioglycollate Medium with 0.5 percent beef extract shall be used to test for bacteria in biological products containing clostridial toxoids, bacterins, and bacterin-toxoids. (2) Fluid Thioglycollate Medium with or without 0.5 percent beef extract shall be used to test for bacteria in bio logical products other than clostridial toxoids, bacterins, and bacterin-tox oids. (3) Soybean-Casein Digest Medium shall be used to test biological prod ucts for fungi; provided, that Fluid Thioglycollate Medium without beef extract shall be substituted when test ing biological products containing mer curial preservatives. (b) Test procedure: (1) Ten test vessels shall be used for each of two media selected in accord ance with paragraph (a)(1), (a)(2), or (a)(3) of this section. Each test vessel shall contain sufficient medium to ne gate the bacteriostatic or fungistatic activity in the inoculum as determined in § 113.25(d). (2) Inoculum: (i) When completed product is tested, 10 final container samples from each serial and each subserial shall be test ed. One ml from each sample shall be inoculated into a corresponding indi vidual test vessel of culture medium: Provided, That, if each final container sample contains less than 2 ml, one half of the contents shall be used as inoculum for each test vessel. (ii) When cell lines, primary cells, or ingredients of animal origin are tested, at least a 20 ml test sample from each lot shall be tested. One ml shall be in oculated into each test vessel of me dium. (3) Incubation shall be for an observa tion period of 14 days at 30 °to 35 °C. to test for bacteria and 14 days at 20 °to 25 °C. to test for fungi. (4) If the inoculum renders the me dium turbid so that the absence of growth cannot be determined by visual examination, subcultures shall be made VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00717 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 708 9 CFR Ch. I (1–1–18 Edition) § 113.27 on the seventh to eleventh day from bi ological products prepared from clostridial toxoids, bacterins, and bacterin-toxoids and the third to sev enth day for other biological products. Portions of the turbid medium in amounts of not less than 1.0 ml. shall be transferred to 20 to 25 ml. of fresh medium, and incubated the balance of the 14-day period. (c) Examine the contents of all test vessels for macroscopic microbial growth during the incubation period. When demonstrated by adequate con trols to be invalid, the test may be re peated. For each set of test vessels rep resenting a serial or subserial in a valid test, the following rules shall apply: (1) If no growth is found in any test vessel, the serial or subserial meets the requirements of the test. (2) If growth is found in any test ves sel, one retest to rule out faulty tech nique may be conducted using 20 un opened final container samples. (3) If growth is found in any test ves sel of the final test, the serial, sub serial, or ingredients to be used in the preparation of a biological product, as the case may be, is unsatisfactory. [35 FR 16039, Oct. 13, 1970, as amended at 37 FR 2430, Feb. 1, 1972; 39 FR 21042, June 18, 1974; 40 FR 758, Jan. 3, 1975; 40 FR 14084, Mar. 28, 1975; 56 FR 66784, Dec. 26, 1991] § 113.27 Detection of extraneous viable bacteria and fungi in live vaccines. Unless otherwise specified by the Ad ministrator or elsewhere exempted in this part, each serial and subserial of live vaccine and each lot of Master Seed Virus and Master Seed Bacteria shall be tested for extraneous viable bacteria and fungi as prescribed in this section. A Master Seed found unsatis factory shall not be used in vaccine production and a serial found unsatis factory shall not be released. (a) Live viral vaccines. Each serial and subserial of live viral vaccine shall be tested for purity as prescribed in this paragraph. However, products of chick en embryo origin recommended for ad ministration other than by parenteral injection may be tested as provided in paragraph (e) of this section. (1) Soybean Casein Digest Medium shall be used. (2) Ten final container samples from each serial and subserial shall be test ed. (3) Immediately prior to starting the test, frozen liquid vaccine shall be thawed, and desiccated vaccine shall be rehydrated as recommended on the label with accompanying diluent or with sterile purified water. (4) To test for bacteria, place 0.2 ml of vaccine from each final container into a corresponding individual vessel containing at least 120 ml of Soybean Casein Digest Medium. Additional me dium shall be used if the determination required in § 113.25(d) indicates the need for a greater dilution of the product. Incubation shall be at 30 °to 35 °C for 14 days. (5) To test for fungi, place 0.2 ml of vaccine from each final container sam ple into a corresponding individual ves sel containing at least 40 ml of Soy bean Casein Digest Medium. Additional medium shall be used if the determina tion required in § 113.25(d) indicates the need for a greater dilution of the prod uct. Incubation shall be at 20 °to 25 °C for 14 days. (6) Examine the contents of all test vessels macroscopically for microbial growth at the end of the incubation pe riod. If growth in a vessel cannot be re liably determined by visual examina tion, judgment shall be confirmed by subcultures, microscopic examination, or both. (7) For each set of test vessels rep resenting a serial or subserial tested according to these procedures, the fol lowing rules shall apply: (i) If growth is found in 2 or 3 test vessels of the initial test, 1 retest to rule out faulty technique may be con ducted using 20 unopened final con tainer samples. (ii) If no growth is found in 9 or 10 of the test vessels in the initial test, or 19 or 20 vessels in the retest, the serial or subserial meets the requirements of the test. (iii) If growth is found in four or more test vessels in the initial test, or two or more in a retest, the serial or subserial is unsatisfactory. (b) Live bacterial vaccines. Each serial or subserial of live bacterial vaccine shall be tested for purity as prescribed in this paragraph. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00718 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 709 Animal and Plant Health Inspection Service, USDA § 113.27 (1) Soybean Casein Digest Medium and Fluid Thioglycollate Medium shall be used. (2) Ten final container samples from each serial and subserial shall be test ed. (3) Immediately prior to starting the test, frozen liquid vaccine shall be thawed, and desiccated vaccine shall be rehydrated as recommended on the label with accompanying diluent or with sterile purified water. Product recommended for mass vaccination shall be rehydrated at the rate of 30 ml sterile purified water per 1,000 doses. (4) To test for extraneous bacteria, place 0.2 ml of vaccine from each final container into a corresponding indi vidual vessel containing at least 40 ml of Fluid Thioglycollate Medium. Addi tional medium shall be used if the de termination required in § 113.25(d) indi cates the need for a greater dilution of the product. Incubation shall be at 30 °to 35 °C for 14 days. (5) To test for extraneous fungi, place 0.2 ml of vaccine from each final con tainer into a corresponding individual vessel containing at least 40 ml of Soy bean Casein Digest Medium. Additional medium shall be used if the determina tion required in § 113.25(d) indicates the need for a greater dilution of the prod uct. Incubation shall be at 20 °to 25 °C for 14 days. (6) Examine the contents of all test vessels macroscopically for atypical microbial growth at the end of the in cubation period. If growth of extra neous microorganisms cannot be reli ably determined by visual examina tion, judgment shall be confirmed by subculturing, microscopic examina tion, or both. (7) For each set of test vessels rep resenting a serial or subserial tested according to these procedures, the fol lowing rules shall apply: (i) If extraneous growth is found in 2 or 3 test vessels of the initial test, 1 retest to rule out faulty technique may be conducted using 20 unopened final container samples. (ii) If no extraneous growth is found in 9 or 10 test vessels in the initial test, or 19 or 20 vessels in the retest, the se rial or subserial meets the require ments of the test. (iii) If extraneous growth is found in 4 or more test vessels in the initial test, or 2 or more in a retest, the serial or subserial is unsatisfactory. (c) Master Seed Virus. Not less than 4 ml of each lot of Master Seed Virus shall be tested. Frozen liquid Master Seed Virus shall be thawed, and des iccated Master Seed Virus shall be re hydrated with Soybean Casein Digest Medium immediately prior to starting the test. (1) To test for bacteria, place 0.2 ml of the sample of Master Seed Virus into 10 individual vessels each containing at least 120 ml of Soybean Casein Digest Medium. Incubation shall be at 30 °to 35 °C for 14 days. (2) To test for fungi, place 0.2 ml of the sample of Master Seed Virus into 10 individual vessels each containing at least 40 ml of Soybean Casein Digest Medium. Incubation shall be at 20 °to 25 °C for 14 days. (3) Examine the contents of all test vessels macroscopically for microbial growth at the end of the incubation pe riod. If growth in a vessel cannot be re liably determined by visual examina tion, judgment shall be confirmed by subcultures, microscopic examination, or both. (4) For each set of test vessels rep resenting a lot of Master Seed Virus tested according to these procedures, the following rules shall apply: (i) If growth is found in any test ves sel of the initial test, one retest to rule out faulty technique may be conducted using a new sample of Master Seed Virus. (ii) If growth is found in any test ves sel of the final test, the lot of Master Seed Virus is unsatisfactory. (d) Master Seed Bacteria. Not less than 4 ml of each lot of Master Seed Bac teria shall be tested. Frozen liquid Master Seed Bacteria shall be thawed, and desiccated Master Seed Bacteria shall be rehydrated with sterile puri fied water immediately prior to start ing the test. (1) To test for extraneous bacteria, place 0.2 ml of the sample of Master Seed Bacteria into 10 individual vessels each containing at least 40 ml of Fluid Thioglycollate Medium. Incubation shall be at 30 °to 35 °C for 14 days. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00719 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 710 9 CFR Ch. I (1–1–18 Edition) § 113.28 (2) To test for extraneous fungi, place 0.2 ml of the sample of Master Seed Bacteria into 10 individual vessels each containing at least 40 ml of Soybean Casein Digest Medium. Incubation shall be at 20 °to 25 °C for 14 days. (3) Examine the contents of all test vessels macroscopically for atypical microbial growth at the end of the in cubation period. If growth of extra neous microorganisms cannot be reli ably determined by visual examina tion, judgment shall be confirmed by subcultures, microscopic examination, or both. (4) For each set of test vessels rep resenting a lot of Master Seed Bacteria tested according to these procedures, the following rules shall apply: (i) If extraneous growth is found in any test vessel of the initial test, one retest to rule out faulty technique may be conducted using a new sample of Master Seed Bacteria. (ii) If extraneous growth is found in any test vessel of the final test, the lot of Master Seed Bacteria is unsatisfac tory. (e) Live viral vaccines of chicken em bryo origin recommended for adminis tration other than by parenteral injec tion, which were not tested or have not been found free of bacteria and fungi by the procedures prescribed in paragraph (a) of this section, may be tested ac cording to the procedures prescribed in this paragraph. (1) Brain Heart Infusion Agar shall be used with 500 Kinetic (Kersey) units of penicillinase per ml of medium added just prior to pouring the plates. (2) Ten final containers from each se rial and each subserial shall be tested. (3) Immediately prior to starting the test, frozen liquid vaccine shall be thawed, and lyophilized vaccine shall be rehydrated to the quantity rec ommended on the label using the ac companying sterile diluent or sterile purified water. Product recommended for mass vaccination shall be re hydrated at the rate of 30 ml sterile pu rified water per 1,000 doses. (4) From each container sample, each of 2 plates shall be inoculated with vac cine equal to 10 doses if the vaccine is recommended for poultry or 1 dose if the vaccine is recommended for other animals. Twenty ml of medium shall be added to each plate. One plate shall be incubated at 30 °to 35 °for 7 days and the other plate shall be incubated at 20 °to 25 °C for 14 days. (5) Colony counts shall be made for each plate at the end of the incubation period. An average colony count for the 10 samples representing the serial or subserial shall be made for each incu bation condition. (6) For each set of test vessels rep resenting a serial or subserial tested according to these procedures, the fol lowing rules shall apply: (i) If the average count at either in cubation condition exceeds 1 colony per dose for vaccines recommended for poultry, or 10 colonies per dose for vac cines recommended for other animals in the initial test, 1 retest to rule out faulty technique may be conducted using 20 unopened final containers. (ii) If the average count at either in cubation condition of the final test for a serial or subserial exceeds 1 colony per dose for vaccines recommended for poultry, or 10 colonies per dose for vac cines recommended for other animals, the serial or subserial is unsatisfac tory. [48 FR 28430, June 22, 1983, as amended at 56 FR 66784, Dec. 26, 1991] § 113.28 Detection of mycoplasma con tamination. The heart infusion test, using heart infusion broth and heart infusion agar, provided in this section shall be con ducted when a test for mycoplasma contamination is prescribed in an ap plicable Standard Requirement or in the filed Outline of Production for the product. (a) Media additives provided in this paragraph shall be prepared as follows: (1) DPN-Cysteine Solution: (i) Use Nicotinamide adenine dinucleotide (oxidized) and L-Cysteine hydrochloride. (ii) Prepare 1 gram/100 milliliters (ml) purified water (1 percent solution) of each. Mix the solutions together; the cysteine reduces the DPN. Filter steri lize, dispense in appropriate amounts and store frozen at ¥20 degrees centi grade. (2) Inactivated horse serum—horse serum which has been inactivated at 56 °C for 30 minutes. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00720 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 711 Animal and Plant Health Inspection Service, USDA § 113.29 (b) Heart infusion broth shall be pre pared as provided in this paragraph. (1) Dissolve in 970 ml of purified water, 25 grams of heart infusion broth, 10 grams of proteose peptone No. 3, and either 5 grams of yeast autolysate or 5 ml of fresh yeast extract. (2) Add the following: 1 percent tetrazolium chloride (ml) ........................ 5.5 1 percent thallium acetate (ml) .............................. 25 Penicillin (units) ...................................................... 500,000 Inactivated horse serum (ml) ................................. 100 (3) Adjust pH to 7.9 with NaOH, filter sterilize, and dispense 100 ml aliquots into 125 ml flasks and store until need ed. (4) Add 2 ml of DPN-Cysteine solu tion to each 100 ml of broth on day of use. (c) Heart Infusion Agar shall be pre pared as provided in this paragraph. (1) Dissolve in 900 ml of purified water by boiling the following: Heart infusion agar (g) ........................................... 25 Heart-infusion broth (g) .......................................... 10 Proteose peptone No. 3 (g) ................................... 10 1 pct thallium acetate (ml) ..................................... 25 (2) Cool the medium and adjust pH to 7.9 with NaOH. (3) Autoclave the medium. (4) Cool the medium 30 minutes in a 56 °C waterbath. (5) Dissolve 5 grams of yeast autoly sate in 100 ml of distilled water, filter sterilize, and add to the medium. (6) Add to the medium: 126 ml of inactivated horse serum 21 ml of DPN-Cysteine solution 525,000 units of Penicillin. Dispense 10 ml aliquots into 60 × 15 mm dis posable culture dishes or petri dishes. (d) The test procedure provided in this paragraph shall be followed when conducting the mycoplasma detection test. (1) Preparation of inoculum. Imme diately prior to starting the test, fro zen liquid vaccine shall be thawed, and lyophilized vaccine shall be rehydrated to the volume recommended on the label with mycoplasma medium. In the case of a lyophilized biological product, e.g., 1,000 dose vial of poultry vaccine to be administered via the drinking water, the vaccine shall be rehydrated to 30 ml with mycoplasma medium. In the case of a cell line or a sample of primary cells, the inoculum shall con sist of the resuspended cells. Control tests shall be established as provided in paragraph (d)(4) of this section. (2) Inoculation of plate. Plate 0.1 ml of inoculum on an agar plate and make a short, continuous streak across the plate with a pipet. Tilt the plate to allow the inoculum to flow over the surface. (3) Inoculation of flask of medium. Transfer 1 ml of the inoculum into a flask containing 100 ml mycoplasma medium and mix thoroughly. Incubate the flask at 33 to 37 °C for 14 days dur ing which time, one of four agar plates shall be streaked with 0.1 ml of mate rial from the incubating flask of inocu lated medium on the 3d day, one on the 7th day, one on the 10th day, and one on the 14th day post-inoculation. (4) Control tests shall be conducted simultaneously with the detection test using techniques provided in para graphs (d)(2) and (3) of this section, ex cept the inoculum for the positive con trol test shall be selected mycoplasma cultures and the negative control test shall be uninoculated medium from the same lot used in the detection test. (5) All plates shall be incubated in a high humidity, 4–6 percent CO2 atmos phere at 33 °to 37 °C for 10–14 days and examined with a stereoscopic micro scope at 35x to 100x or with a regular microscope at 100x. (e) Interpretation of test results. (1) If growth appears on at least one of the plates in the positive control test and does not appear on any of the plates in the negative control test, the test is valid. (2) If mycoplasma colonies are found on any of the plates inoculated with material being tested, the results are positive for mycoplasma contamina tion. [38 FR 29887, Oct. 30, 1973, as amended at 41 FR 6752, Feb. 13, 1976; 41 FR 32882, Aug. 6, 1976] § 113.29 Determination of moisture content in desiccated biological products. Methods provided in this section must be used when a determination of moisture content in desiccated biologi cal products is prescribed in an appli cable Standard Requirement or in the filed Outline of Production for the VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00721 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 712 9 CFR Ch. I (1–1–18 Edition) § 113.30 product. Firms currently using meth ods other than those provided in this section for determining the moisture content in desiccated biological prod ucts have until November 5, 2004 to up date their Outlines of Production to be in compliance with this requirement. (a) Final container samples of com pleted product shall be tested. The weight loss of the sample due to drying in a vacuum oven shall be determined. All procedures should be performed in an environment with a relative humid ity less than 45 percent. The equipment necessary to perform the test is as fol lows: (1) Cylindrical weighing bottles with airtight glass stoppers. (2) Vacuum oven equipped with vali dated thermometer and thermostat. A suitable air-drying device should be at tached to the inlet valve. (3) Balance, accurate to 0.1 mg (rated precision ±0.01mg). (4) Desiccator jar equipped with phos phorous pentoxide, silica gel, or equiv alent. (5) Desiccated vaccine in original sealed vial. Sample and control should be kept at room temperature in their original airtight containers until use. (b) Test procedure: (1) Thoroughly cleaned and labeled sample-weighing bottles with stoppers should be allowed to dry at 60 ±3 °C under vacuum at less than 2.5 kPa. (i) Transfer hot bottles and stoppers into the desiccator and allow to cool to room temperature. (ii) After bottles have cooled, insert stoppers and weigh and record the weights of the bottles as ‘‘A.’’ (iii) Return weighing bottles to the desiccator. (2) Remove the sample container seal. (i) Using a spatula, break up the sam ple plug and transfer the required amount of sample to the previously tared weighing bottle. (ii) Insert the stopper and weigh and record the weights of the weighing bot tles as ‘‘B.’’ (3) Place the weighing bottle with the stopper at an angle in the vacuum oven. Set the vacuum to <2.5 kPa and the temperature to 60 ±3 °C. (4) After a minimum of 3 hours of drying time, turn off the vacuum pump and allow dry air to bleed into the oven until the pressure inside the oven is equalized with the prevailing atmos pheric pressure. (5) While the bottle is still warm, re place the stopper in its normal position and transfer the weighing bottle to the desiccator. (i) Allow a minimum of 2 hours for the weighing bottle to cool to room temperature or for its weight to reach equilibrium. (ii) Weigh, and record the weight as ‘‘C.’’ (6) Calculate the percentage of mois ture in the original sample as follows: (B¥C)/(B¥A) × (100) = Percentage of re sidual moisture, where: A = tare weight of weighing bottle B¥A = weight of sample before drying B¥C = weight of sample after drying (7) The results are considered satis factory if the percentage of residual moisture is less than or equal to the manufacturer’s specification. [68 FR 57608, Oct. 6, 2003] § 113.30 Detection of Salmonella con tamination. The test for detection of Salmonella contamination provided in this section shall be conducted when such a test is prescribed in an applicable Standard Requirement or in the filed Outline of Production for the product. (a) Samples shall be collected from the bulk suspension before bacteriostatic or bactericidal agents have been added. When tissue culture products are to be tested, 1 ml of tissue extract used as the source of cells or 1 ml of the minced tissue per se shall be tested. (b) Five ml of the liquid vaccine sus pension shall be used to inoculate each 100 ml of liquid broth medium (tryptose and either selenite F or tetrathionate). The inoculated media shall be incu bated 18–24 hours at 35–37 °C. (c) Transfers shall be made to either MacConkey agar or Salmonella Shigella agar, incubated for 18–24 hours and examined. (d) If no growth typical of Salmonella is noted, the plates shall be incubated an additional 18–24 hours and again ex amined. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00722 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 713 Animal and Plant Health Inspection Service, USDA § 113.31 (e) If suspicious colonies are ob served, further subculture on suitable media shall be made for positive identi fication. If Salmonella is found, the bulk suspension is unsatisfactory. [38 FR 29888, Oct. 30, 1973] § 113.31 Detection of avian lymphoid leukosis. The complement-fixation test for de tection of avian lymphoid leukosis pro vided in this section shall be conducted on all biological products containing virus which has been propagated in substrates of chicken origin: Provided, An inactivated viral product shall be exempt from this requirement if the li censee can demonstrate to Animal and Plant Health Inspection Service that the agent used to inactivate the vac cine virus would also inactivate lymph oid leukosis virus. (a) Propagation of contaminating lymphoid leukosis viruses, if present, shall be done in chick embryo cell cul tures. (1) Each vaccine virus, cytopathic to chick embryo fibroblast cells, shall be effectively neutralized, inactivated, or separated so that minimal amounts of lymphoid leukosis virus can be propa gated on cell culture during the 21-day growth period. If a vaccine virus can not be effectively neutralized, inac tivated, or separated, a sample of an other vaccine prepared the same week from material harvested from each source flock (or other sampling proce dure acceptable to Animal and Plant Health Inspection Service) used for the preparation of the questionable vaccine virus that cannot be neutralized, inac tivated, or separated shall be tested each week during the preparation of such questionable vaccine. (2) When cell cultures are tested, 5 ml of the final cell suspension as prepared for seeding of production cell cultures shall be used as inoculum. When vac cines are tested, the equivalent of 200 doses of Newcastle disease vaccine or 500 doses of other vaccines for use in poultry, or one dose of vaccine for use in other animals shall be used as inoculum. Control cultures shall be prepared from the same cell suspension as the cultures for testing the vaccine. (3) Uninoculated chick embryo fibro blast cell cultures shall act as negative controls. One set of chick fibroblast cultures inoculated with subgroup A virus and another set inoculated with subgroup B virus shall act as positive controls, A and B respectively. (4) The cell cultures shall be propa gated at 35–37 °C for at least 21 days. They shall be passed when necessary to maintain viability and samples har vested from each passage shall be test ed for group specific antigen. (b) The microtiter complement-fixa tion test shall be performed using ei ther the 50 percent or the 100 percent hemolytic end point technique to de termine complement unitage. Five 50 percent hemolytic units or two 100 per cent hemolytic units of complement shall be used for each test. (1) All test materials, including posi tive and negative controls, shall be stored at ¥60 °C or colder until used in the test. Before use, each sample shall be thawed and frozen three times to disrupt intact cells and release the group specific antigen. (2) The antiserum used in the microtiter complement-fixation test shall be a standard reagent supplied or approved by the Animal and Plant Health Inspection Service. Four units of antiserum shall be used for each test. (3) Presence of complement-fixing ac tivity in the harvested samples (from passages) at the 1:4 or higher dilution, in the absence of anticomplementary activity, shall be considered a positive test unless the activity can definitely be established to be caused by some thing other than lymphoid leukosis virus, subgroups A and/or B. Activity at the 1:2 dilution shall be considered suspicious and the sample further sub cultured to determine presence or ab sence of the group specific antigen. (4) Biological products or primary cells which are found contaminated with lymphoid leukosis viruses are un satisfactory. Source flocks from which contaminated material was obtained are also unsatisfactory. [38 FR 29888, Oct. 30, 1973, as amended at 38 FR 32917, Nov. 29, 1973; 39 FR 21042, June 18, 1974; 56 FR 66784, Dec. 26, 1991] VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00723 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 714 9 CFR Ch. I (1–1–18 Edition) § 113.32 § 113.32 Detection of Brucella contami nation. The test for detection of Brucella contamination provided in this section shall be conducted when such a test is prescribed in an applicable Standard Requirement or in a filed Outline of Production for the product. (a) One ml of the minced tissue used as the source of cells or 1 ml of the ex tract of the tissue prior to the addition of antibiotics, diluent and stabilizer, shall be inoculated onto each of three tryptose agar plates and incubated in a 10 percent CO2 atmosphere at a tem perature of 35–37 °C for at least 7 days. (b) If colonies are identified as Brucella, the biological product is un satisfactory. (c) If colonies suspicious of Brucella are observed but cannot be identified as a Brucella species, either (1) The biological product shall be re garded as unsatisfactory and de stroyed; or (2) Further subculture or other proce dures shall be carried out until a posi tive identification can be made. [38 FR 29888, Oct. 30, 1973] § 113.33 Mouse safety tests. One of the mouse safety tests pro vided in this section shall be conducted when such test is prescribed in a Standard Requirement or in the filed Outline of Production for a biological product recommended for animals other than poultry: Provided, That if the inherent nature of one or more in gredients makes the biological product lethal or toxic for mice but not lethal or toxic for the animals for which it is recommended, the licensee shall dem onstrate the safety of such product by an acceptable test written into such Outline of Production. (a) Final container samples of com pleted product from live virus vaccines shall be tested for safety using young adult mice in accordance with the test provided in this paragraph. (1) Vaccine prepared for use as rec ommended on the label shall be tested by inoculating eight mice intraperitoneally or subcutaneously with 0.5 mL (the inoculation volume may be divided among more than one injection site), and the animals ob served for 7 days. (2) If unfavorable reactions attrib utable to the product occur in any of the mice during the observation period, the serial or subserial is unsatisfac tory. If unfavorable reactions which are not attributable to the product occur, the test shall be declared a No Test and may be repeated: Provided, That, if the test is not repeated, the se rial or subserial shall be declared un satisfactory. (b) Bulk or final container samples of completed product from liquid prod ucts, such as but not limited to antiserums and bacterins, shall be test ed for safety in accordance with the test provided in this paragraph. (1) Unless otherwise prescribed in the Standard Requirement or approved in a filed Outline of Production for the product, a 0.5 ml dose shall be injected intraperitoneally or subcutaneously into eight mice and the animals ob served for 7 days. (2) If unfavorable reactions attrib utable to the product occur in any of the mice during the observation period, the serial or subserial is unsatisfac tory. If unfavorable reactions which are not attributable to the product occur, the test shall be declared a No Test and may be repeated: Provided, That, if the test is not repeated, the se rial or subserial shall be declared un satisfactory. [38 FR 34727, Dec. 18, 1973, as amended at 39 FR 16857, May 10, 1974; 72 FR 72564, Dec. 21, 2007] § 113.34 Detection of hemagglutinating viruses. The test for detection of hemagglutinating viruses provided in this section shall be conducted when such a test is prescribed in an applica ble Standard Requirement or in the filed Outline of Production for the product. (a) Final container samples of com pleted product rehydrated as rec ommended on the label shall be used as inoculum: Provided, That poultry vac cines distributed without diluent shall be rehydrated with 30 ml of sterile dis tilled water per 1,000 doses and used as inoculum. When one or more fractions are to be used in combination with VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00724 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 715 Animal and Plant Health Inspection Service, USDA § 113.36 Newcastle Disease Vaccine, test sam ples shall be collected from bulk sus pensions of each prior to mixing with the Newcastle Disease Vaccine. (b) Each of ten 9- to 10-day-old embryonating eggs from Newcastle dis ease susceptible flocks shall be inocu lated into the allantoic cavity with 0.2 ml of the undiluted inoculum. (1) Test five uninoculated embryos of the same age and from the same flock as those used for the test as negative controls. (2) Test an allantoic fluid sample of Newcastle disease virus as a positive control. (c) Three to five days post-inocula tion, a sample of allantoic fluid from each egg shall be tested separately by a rapid plate test for hemagglutinating activity using a 0.5 percent suspension of fresh chicken red blood cells. (d) If the results are inconclusive, one or two blind passages shall be made using fluids from each of the original test eggs. Fluids from dead and live embryos may be pooled separately for inoculum in these passages. (e) If hemagglutinating activity at tributable to the product is observed, the serial is unsatisfactory. [38 FR 29889, Oct. 30, 1973] § 113.35 Detection of viricidal activity. The test for detection of viricidal ac tivity provided in this section shall be conducted when such a test is pre scribed in an applicable standard re quirement or in the filed Outline of Production for each inactivated liquid biological product used as diluent for a desiccated live virus vaccine in a com bination package. (a) Bulk or final container samples of completed product from each serial shall be tested. (b) The product shall be tested with each virus fraction for which it is to be used as a diluent. If the vaccine to be rehydrated contains more than one virus fraction, the test shall be con ducted with each fraction after neu tralization of the other fraction(s), and/ or dilution of the vaccine beyond the titer range of the other fraction(s), or the test shall be conducted using rep resentative single-fraction desiccated vaccines which are prepared by the li censee and which are licensed. Pro vided, That the Administrator may au thorize licensees to prepare and use un licensed single-fraction vaccines for this purpose. (c) Test procedure: (1) Rehydrate at least two vials of the vaccine with the liquid product under test according to label recommendations and pool the contents. (2) Rehydrate at least two vials of the vaccine with the same volume of sterile purified water and pool the con tents. (3) Neutralize to remove other frac tions, if necessary. (4) Hold the two pools of vaccine at room temperature (20 °to 25 °C) for 2 hours. The holding period shall begin when rehydration is completed. (5) Titrate the virus(es) in each pool of vaccine as provided in the filed Out line of Production or an applicable standard requirement. (6) Compare respective titers. (d) If the titer of the vaccine virus(es) rehydrated with the product under test is more than 0.7 log10 below the titer of the vaccine virus(es) rehydrated with sterile purified water, the product is unsatisfactory for use as diluent. (e) If the product is unsatisfactory in the first test, one retest to rule out faulty techniques may be conducted using four vials of the vaccine for each pool and the acceptability of the prod uct judged by the results of the second test. (f) Liquid products found to be unsat isfactory for use as diluent by this test are not prohibited from release as sepa rate licensed products if labeled as pre scribed in § 112.7(g). [44 FR 25412, May 1, 1979, as amended at 56 FR 66784, Dec. 26, 1991; 64 FR 43044, Aug. 9, 1999] § 113.36 Detection of pathogens by the chicken inoculation test. The test for detection of extraneous pathogens provided in this section shall be conducted when such a test is prescribed in an applicable Standard Requirement or in the filed Outline of Production for the product. (a) The biological product to be test ed shall be prepared for use as rec ommended on the label, or in the case of desiccated vaccine to be used in VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00725 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 716 9 CFR Ch. I (1–1–18 Edition) § 113.37 poultry, rehydrated with sterile dis tilled water at the rate of 30 ml per 1,000 doses. (b) At least 25 healthy susceptible young chickens, properly identified and obtained from the same source and hatch, shall be immunized at least 14 days prior to being put on test. The im munizing agent shall be the same as the product to be tested but from a se rial previously tested and found satis factory. (c) At least 20 of the previously im munized birds shall be inoculated with 10 label doses of the vaccine being test ed by each of the following routes: Sub cutaneous, intratracheal, eye-drop, and comb scarification (1 cm 2). Twenty birds may be used for each route or combination of routes. (d) At least five birds shall be iso lated as control birds. (e) All birds shall be observed for 21 days for signs of septicemic diseases, respiratory diseases, or other pathologic conditions. (f) If the controls remain healthy and unfavorable reactions attributable to the product occur in the vaccinates, the serial or subserial tested is unsatis factory. If the controls do not remain healthy or if unfavorable reactions not attributable to the product occur in the vaccinates, or both, the test shall be declared a No Test and may be re peated: Provided, That, if the test is not repeated, the serial of subserial tested shall be considered unsatisfactory. [38 FR 29889, Oct. 30, 1973, as amended at 39 FR 21042, June 18, 1974; 43 FR 7610, Feb. 24, 1978] § 113.37 Detection of pathogens by the chicken embryo inoculation test. The test for detection of extraneous pathogens provided in this section shall be conducted when such a test is prescribed in an applicable Standard Requirement or in the filed Outline of Production for the product. (a) The biological product to be test ed shall be prepared for use as rec ommended on the label, or in the case of desiccated vaccine to be used in poultry, rehydrated with sterile dis tilled water at the rate of 30 ml per 1,000 doses. (b) One volume of the prepared vac cine shall be mixed with up to nine vol umes of sterile heat-inactivated spe cific antiserum to neutralize the vac cine virus in the product. Each lot of antiserum shall be demonstrated by virus neutralization tests not to in hibit other viruses known to be pos sible contaminants. (c) After neutralization, 0.2 ml of the vaccine-serum mixture shall be inocu lated into each of at least 20 fully sus ceptible chicken embryos. (1) Twenty embryos, 9 to 11 days old, shall be inoculated on the chorio allantoic membrane (CAM) with 0.1 ml, and in the allantoic sac with 0.1 ml. (2) Eggs shall be candled daily for 7 days. Deaths occurring during the first 24 hours shall be disregarded but at least 18 viable embryos shall survive 24 hours post-inoculation for a valid test. Examine all embryos and CAM’s from embryos which die after the first day. When necessary, embryo subcultures shall be made to determine the cause of a death. The test shall be concluded on the seventh day post-inoculation and the surviving embryos (including CAM’s) examined. (d) If death and/or abnormality at tributable to the inoculum occur, the serial is unsatisfactory: Provided, That, if there is a vaccine virus override, the test may be repeated, using a higher titered antiserum. [38 FR 29889, Oct. 30, 1973, as amended at 39 FR 21042, June 18, 1974] § 113.38 Guinea pig safety test. The guinea pig safety test provided in this section shall be conducted when prescribed in a Standard Requirement or approved Outline of Production for a biological product. When desiccated products are tested, final container samples of completed product prepared for administration in the manner rec ommended on the label shall be used. When liquid products are tested, either bulk or final container samples of com pleted product shall be used. (a) Unless otherwise specified in the Standard Requirement or approved Outline of Production for the product, a 2 ml dose shall be injected either intramuscularly or subcutaneously into each of two guinea pigs and the animals observed for 7 days. (b) If unfavorable reactions attrib utable to the product occur in either of VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00726 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 717 Animal and Plant Health Inspection Service, USDA § 113.40 the guinea pigs during the observation period, the serial or subserial is unsat isfactory. If unfavorable reactions which are not attributable to the prod uct occur, the test shall be declared a No Test and may be repeated: Provided, That, if the test is not repeated, the se rial or subserial shall be declared un satisfactory. [39 FR 16857, May 10, 1974; 39 FR 20368, June 10, 1974] § 113.39 Cat safety tests. The safety tests provided in this sec tion shall be conducted when pre scribed in a standard requirement or in the filed Outline of Production for a bi ological product recommended for use in cats. (a) The cat safety test provided in this paragraph shall be used when the Master Seed Virus is tested for safety. (1) The test animals shall be deter mined to be susceptible to the virus under test as follows: (i) Throat swabs shall be collected from each cat and individually tested on susceptible cell cultures for the presence of the virus. Blood samples shall also be drawn and individual serum samples tested for antibody to the virus. (ii) The cats shall be considered sus ceptible if swabs are negative for virus isolation and the serums are free of virus antibody at the 1:2 final dilution in a 50 percent plaque reduction test or other serum-neutralization test of equal sensitivity. (iii) When determining susceptibility to a virus which does not lend itself to the methods in paragraphs (a)(1)(i) and (ii) of this section, a method acceptable to Animal and Plant Health Inspection Service shall be used. (2) Each of at least 10 susceptible cats shall be administered a sample of the Master Seed Virus equivalent to the amount of virus to be used in one cat dose of the vaccine, by the method to be recommended on the label, and the cats observed each day for 14 days. (3) If unfavorable reactions attrib utable to the virus occur in any of the cats during the observation period, the Master Seed Virus is unsatisfactory. If unfavorable reactions occur which are not attributable to the Master Seed Virus, the test shall be declared a No Test and repeated: Provided, That, if not repeated, the Master Seed Virus shall be unsatisfactory. (b) The cat safety test provided in this paragraph shall be used when a se rial of vaccine is tested for safety be fore release. (1) Each of two healthy cats shall be administered 10 cat doses by the meth od recommended on the label and the cats observed each day for 14 days. (2) If unfavorable reactions attrib utable to the biological product occur during the observation period, the se rial is unsatisfactory. If unfavorable reactions occur which are not attrib utable to the product, the test shall be declared a No Test and repeated: Pro vided, That, if not repeated, the serial shall be unsatisfactory. [44 FR 58898, Oct. 12, 1979, as amended at 56 FR 66784, Dec. 26, 1991] § 113.40 Dog safety tests. The safety tests provided in this sec tion shall be conducted when pre scribed in a Standard Requirement or in the filed Outline of Production for a biological product recommended for use in dogs. Serials which are not found to be satisfactory when tested pursuant to the procedures in this sec tion may not be released for shipment. (a) The dog safety test provided in this paragraph shall be used when the Master Seed Virus is tested for safety. (1) The test animals shall be deter mined to be susceptible to the virus under test by a method acceptable to the Animal and Plant Health Inspec tion Service. (2) Each of at least 10 susceptible dogs shall be administered a sample of the Master Seed Virus equivalent to the amount of virus to be used in one dog dose of the vaccine, by the method recommended on the label, and the dog shall be observed each day for 14 days. (3) If unfavorable reactions attrib utable to the virus occur in any of the dogs during the observation period, the Master Seed Virus is unsatisfactory. If unfavorable reactions occur which are not attributable to the Master Seed Virus, the test shall be declared a No Test and may be repeated: Provided: That, if the test is not repeated, the Master Seed Virus shall be considered unsatisfactory. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00727 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 718 9 CFR Ch. I (1–1–18 Edition) § 113.41 (b) The dog safety test provided in this paragraph shall be used when a se rial of vaccine is tested for safety be fore release. (1) Each of two healthy dogs shall be administered 10 dog doses by the meth od recommended on the label and the dogs shall be observed each day for 14 days. (2) If unfavorable reactions attrib utable to the biological product occur during the observation period, the se rial is unsatisfactory. If unfavorable reactions occur which are not attrib utable to the biological product, the test shall be declared a No Test and may be repeated: Provided, That, if the test is not repeated, the serial shall be considered unsatisfactory. [60 FR 14358, Mar. 17, 1995] § 113.41 Calf safety test. The calf safety test provided in this section shall be conducted when pre scribed in a Standard Requirement or in the filed Outline of Production for a product. (a) Test procedure. Each of two calves shall be injected with the equivalent of 10 doses of vaccine administered in the manner recommended on the label and observed each day for 21 days. (b) Interpretation. If unfavorable reac tions attributable to the product occur in either of the calves during the obser vation period, the serial or subserial is unsatisfactory. If unfavorable reac tions which are not attributable to the product occur, the test shall be de clared a No Test and may be repeated: Provided, That, if the test is not re peated, the serial or subserial shall be declared unsatisfactory. [39 FR 27428, July 29, 1974] § 113.42 Detection of lymphocytic cho riomeningitis contamination. The test for detection of lymphocytic choriomeningitis (LCM) virus provided in this section shall be conducted when such a test is prescribed in an applica ble Standard Requirement or in a filed Outline of Production. Vaccine virus may be neutralized with specific anti serum when necessary. (a) Each of at least 10 mice obtained from a source free of LCM shall be in jected in the footpad of a hindfoot with 0.02 ml of the material being tested and observed each day for 21 days. (b) If any of the mice show swelling in the injected footpad or if more than one becomes systemically abnormal, the material being tested is unsatisfac tory. [42 FR 6794, Feb. 4, 1977] § 113.43 Detection of chlamydial agents. The test for chlamydial agents pro vided in this section shall be conducted when such a test is prescribed in an ap plicable standard requirement or in a filed Outline of Production. (a) The yolk sac of 6-day-old chicken embryos shall be injected. Three groups of 10 embryos shall be used se quentially. (1) The inoculum for each embryo in the first group shall consist of 0.5 ml of a mixture of equal parts of the seed virus with phosphate buffered saline that may contain Streptomycin, Vancomycin, Kanamycin, or a com bination thereof. Not more than 2 mg/ ml of each antibiotic shall be used. (2) On the 10th day postinoculation, the yolk sac of viable embryos shall be harvested, pooled, homogenized as a 20 percent suspension in phosphate buffered saline antibiotic diluent, and 0.5 ml of the mixture injected into the second group of chicken embryos. This process shall be repeated for the injec tion of the third group of embryos using the yolk sacs of viable embryos from the second group. (3) For each of the three passages, embryo deaths occurring within 48 hours of injection shall be disregarded, except that if more than three such deaths occur at any passage, that pas sage shall be repeated. (b) If one or more embryo deaths occur at any passage after 48 hours postinjection, the yolk sacs from each of the dead embryos shall be subcul tured into 10 additional embryos. If one or more embryo deaths again occur due to chlamydial agents, the Master Seed Virus is unsatisfactory for use to produce vaccine. [44 FR 58899, Oct. 12, 1979] VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00728 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 719 Animal and Plant Health Inspection Service, USDA § 113.46 § 113.44 Swine safety test. The swine safety test provided in this section shall be conducted when pre scribed in a Standard Requirement or in the filed Outline of Production for a product. (a) Test procedure. (1) Inject each of two swine of the minimum age for which the product is recommended with the equivalent of two doses of bac terial vaccine or 10 doses of viral vac cine. (2) Administer vaccine in the manner recommended on the label. (3) Observe swine each day for 21 days. (b) Interpretation. If unfavorable reac tions attributable to the product occur in either of the swine during the obser vation period, the serial or subserial is unsatisfactory. If unfavorable reac tions which are not attributable to the product occur, the test shall be de clared a No Test and may be repeated; Provided, That, if the test is not re peated, the serial or subserial shall be declared unsatisfactory. [48 FR 33476, July 22, 1983] § 113.45 Sheep safety test. The sheep safety test provided in this section shall be conducted when pre scribed in a Standard Requirement or in the filed Outline of Production for a product. (a) Test procedure. (1) Inject each of two sheep of the minimum age for which the product is recommended with the equivalent of two doses of bac terial vaccine or 10 doses of viral vac cine. (2) Administer vaccine in the manner recommended on the label. (3) Observe sheep each day for 21 days. (b) Interpretation. If unfavorable reac tions attributable to the product occur in either of the sheep during the obser vation period, the serial or subserial is unsatisfactory. If unfavorable reac tions which are not attributable to the product occur, the test shall be de clared a No Test and may be repeated; Provided, That, if the test is not re peated, the serial or subserial shall be declared unsatisfactory. [48 FR 33476, July 22, 1983] § 113.46 Detection of cytopathogenic and/or hemadsorbing agents. The tests for detection of cytopathogenic and/or hemadsorbing agents provided in this section shall be conducted when prescribed in an appli cable Standard Requirement or in the filed Outline of Production for a prod uct. (a) Test for cytopathogenic agents. One or more monolayers that are at least 6 cm 2 and at least 7 days from the last subculture shall be tested as provided in this paragraph. (1) Stain each monolayer with a suit able cytological stain. (2) Examine the entire area of each stained monolayer for evidence of in clusion bodies, abnormal number of giant cells, or other cytopathology in dicative of cell abnormalities attrib utable to an extraneous agent. (b) Test for hemadsorbing agents. One or more monolayers that are at least 6 cm 2 and at least 7 days from the last subculture shall be tested as provided in this paragraph. (1) Wash the monolayer with several changes of phosphate buffered saline. (2) Add an appropriate volume of a 0.2 percent red blood cell suspension to uniformly cover the surface of the monolayer of cultured cells. Suspen sions of washed guinea pig and chicken red blood cells shall be used. These sus pensions may be mixed before addition to the monolayer or they may be added separately to individual monolayers. (3) Incubate the monolayer at 4 °C for 30 minutes, wash with phosphate buffered saline, and examine for hemadsorption. (4) If no hemadsorption is apparent, repeat step (b)(2) of this section and in cubate the monolayers at 20–25 °C for 30 minutes, wash with phosphate buffered saline, and examine again for hemadsorption. If desired, separate monolayers may be used for each incu bation temperature. (c) If specific cytopathology or hemadsorption attributable to an ex traneous agent is found, the material under test is unsatisfactory and shall not be used to prepare biological prod ucts. If an extraneous agent is sus pected because of cytopathology or hemadsorption and cannot be elimi nated as a possibility by additional VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00729 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 720 9 CFR Ch. I (1–1–18 Edition) § 113.47 testing, the material under test is un satisfactory. [50 FR 441, Jan. 4, 1985, as amended at 58 FR 50252, Sept. 27, 1993] § 113.47 Detection of extraneous vi ruses by the fluorescent antibody technique. The test for detection of extraneous viruses by the fluorescent antibody technique provided in this section shall be conducted when prescribed in an ap plicable Standard Requirement or in a filed Outline of Production for a prod uct. (a) Monolayer cultures of cells (monolayers), at least 7 days after the last subculturing, shall be processed and stained with the appropriate antiviral fluorochrome-conjugated antibody as specified in paragraph (b) of this section. (1) Three groups of one or more monolayers shall be required for each specific virus prescribed in paragraph (b) of this section. (i) At the time of the last subcul turing, one group of test monolayers shall be inoculated with approximately 100–300 FAID50 of the specific virus being tested for as positive controls. (ii) One group of monolayers shall be the ‘‘material under test.’’ (iii) One group of monolayers, that are of the same type of cells as the test monolayers and that have been tested as prescribed in §§ 113.51 or 113.52 (whichever is applicable), shall be pre pared as negative controls. (2) Each group of monolayers shall have a total area of at least 6 cm 2. (3) Positive control monolayers may be fixed (processed so as to arrest growth and assure attachment of the monolayer to the surface of the vessel in which they are grown) before 7 days after subculturing if fluorescence is en hanced by doing so, Provided, That a monolayer of the material under test is also fixed at the same time as the posi tive control and a monolayer of the material under test is also fixed at least seven days after subculturing. Monolayers that are fixed before 7 days after subculturing shall be stained at the same time as the test monolayers and negative controls fixed at least 7 days after subculturing. (b) The antiviral fluorochrome-con jugated antibodies to be used shall de pend on the type of cells required to be tested for extraneous viruses as speci fied in an applicable Standard Require ment or in a filed Outline of Produc tion. Antiviral fluorochrome-con jugated antibodies specific for the ex traneous viruses shall be applied to each respective type of cell in accord ance with the following list. Under cer tain circumstances, additional tests may need to be conducted, as deter mined by the Administrator. When a specific antiviral fluorochrome-con jugated antibody is used in testing for the listed extraneous viruses specified in more than one cell type, it need only be applied to the most susceptible cell type. (1) All cells shall be tested for: (i) Bovine virus diarrhea virus; (ii) Reovirus; and (iii) Rabies virus. (2) Bovine, caprine, and ovine cells shall, in addition, be tested for: (i) Bluetongue virus; (ii) Bovine adenoviruses; (iii) Bovine parvovirus; and (iv) Bovine respiratory syncytial virus. (3) Canine cells shall, in addition, be tested for: (i) Canine coronavirus; (ii) Canine distemper virus; and (iii) Canine parvovirus. (4) Equine cells shall, in addition, be tested for: (i) Equine herpesvirus; and (ii) Equine viral arteritis virus. (5) Feline cells shall, in addition, be tested for: (i) Feline infectious peritonitis virus; and (ii) Feline panleukopenia virus. (6) Porcine cells shall, in addition, be tested for: (i) Porcine adenovirus; (ii) Porcine parvovirus; (iii) transmissible gastroenteritis virus; and (iv) Porcine hemagglutinating en cephalitis virus. (7) Firms that do not have rabies virus on premises either for research or production purposes are exempt from having to produce positive rabies virus control monolayers. Fixed positive ra bies virus control monolayers will be VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00730 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 721 Animal and Plant Health Inspection Service, USDA § 113.51 provided by the National Veterinary Services Laboratories. (c) After staining, each group of monolayers shall be examined for the presence of specific fluorescence attrib utable to the presence of extraneous vi ruses. (1) If the material under test shows any evidence of specific viral fluores cence, it is unsatisfactory and may not be used; Provided, That, if specific fluo rescence attributable to the virus being tested for is absent in the positive con trol monolayers, the test is a No Test and may be repeated. (2) If the fluorescence of the monolayers inoculated with the spe cific virus as positive controls is equiv ocal, or if the negative monolayers show equivocal fluorescence indicating possible viral contamination, or both, the test shall be declared a No Test, and may be repeated; Provided, That, if the test is not repeated, the material under test shall be regarded as unsatis factory for use in the production of bio logics. [60 FR 24548, May 9, 1995] INGREDIENT REQUIREMENTS § 113.50 Ingredients of biological prod ucts. All ingredients used in a licensed bio logical product shall meet accepted standards of purity and quality; shall be sufficiently nontoxic so that the amount present in the recommended dose of the product shall not be toxic to the recipient; and in the combina tions used shall not denature the spe cific substances in the product below the minimum acceptable potency with in the dating period when stored at the recommended temperature. [38 FR 29889, Oct. 30, 1973] § 113.51 Requirements for primary cells used for production of bio logics. Primary cells used to prepare bio logical products shall be derived from normal tissue of healthy animals. When prescribed in an applicable Standard Requirement or in the filed Outline of Production, each batch of primary cells used to prepare a biologi cal product shall be tested as pre scribed in this section. A batch of pri mary cells found unsatisfactory by any prescribed test shall not be used. A se rial of biological product shall not be released if produced from primary cells that are found unsatisfactory by any prescribed test. (a) Final container samples of com pleted product or samples of the final pool of harvested material or samples of each subculture of cells used to pre pare the biological product shall be shown free of mycoplasma as pre scribed in § 113.28. The sample for test ing shall consist of at least 75 cm 2 of actively growing cells or the equiva lent in harvest fluids; Provided, That all sources of cells in the batch of pri mary cells are represented. (b) Final container samples of com pleted product or samples of the final pool of harvested material or samples of each subculture of cells used to pre pare the biological product shall be shown free of bacteria and fungi as pre scribed in § 113.26 or § 113.27 (whichever is applicable). (c) A monolayer at least 75 cm 2 from each batch of primary cells or each subculture of primary cells used to pre pare a biological product shall be shown free of extraneous agents as pre scribed in this paragraph. (1) The test monolayer shall be main tained using the medium (with addi tives) and under conditions similar to those used to prepare biological prod ucts. (i) Monolayers of avian origin shall be maintained for at least 14 days and shall be subcultured at least once dur ing the maintenance period. All but the last subculture shall result in a new monolayer of at least 75 cm 2. The last subculture shall meet the minimum area requirement specified in §§ 113.46 and 113.47. (ii) Monolayers not of avian origin shall be maintained for at least 28 days and shall be subcultured at least twice during the maintenance period. All but the last subculture shall result in a new monolayer of at least 75 cm 2. The last subculture shall meet the min imum area requirement specified in §§ 113.46 and 113.47. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00731 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 722 9 CFR Ch. I (1–1–18 Edition) § 113.52 (2) Monolayers shall be examined reg ularly throughout the required mainte nance period for evidence of the pres ence of cytopathogenic agents. If evi dence of a cytopathogenic agent is found, the batch of primary cells is un satisfactory. (3) At the conclusion of the required maintenance period, monolayers shall be tested for: (i) Cytopathogenic and/or hemadsorbing agents as prescribed in § 113.46; (ii) Extraneous viruses by the fluo rescent antibody technique as pre scribed in § 113.47. [50 FR 442, Jan. 4, 1985, as amended at 60 FR 24549, May 9, 1995] § 113.52 Requirements for cell lines used for production of biologics. When prescribed in an applicable Standard Requirement or in a filed Outline of Production each cell line used to prepare a biological product shall be tested as prescribed in this sec tion. A cell line found unsatisfactory by any prescribed test shall not be used. A serial of biological product shall not be released if produced from a cell line that is found unsatisfactory by any prescribed test. (a) General requirements. (1) A com plete record of the cell line shall be kept, such as, but not limited to, the source, passage history, and medium used for propagation. (2) A Master Cell Stock (MCS) shall be established at a specified passage level for each cell line. The passage level and identity of the MCS and the highest passage level (MCS + n) in tended for use in the preparation of a biological product shall be specified in the Outline of Production for the prod uct. (3) Sufficient 1.0 ml or larger aliquots of MCS and MCS + n shall be prepared, kept in a frozen state, and made avail able to Animal and Plant Health In spection Service (APHIS) upon request for performing the tests prescribed in this section. (4) Each lot of cells shall be mon itored for the characteristics deter mined to be normal for the cell line, such as, but not limited to, micro scopic appearance, growth rate, acid production, or other observable fea tures. (b) The MCS shall be shown to be of the same species of origin as that re ported in paragraph (a)(1) of this sec tion by the following method: (1) At least four monolayers with a total area of at least 6 cm 2 shall be grown to at least 80 percent confluency. (2) The monolayers shall be removed from their media, processed, stained, and examined. (i) At least two monolayers shall be stained with an antispecies fluorchrome-conjugated antibody unre lated to the species of origin of the MCS. (ii) At least two monolayers shall be stained with an antispecies fluorochrome-conjugated antibody spe cific to the species of origin of the MCS. (iii) All monolayers shall be exam ined for evidence of specific fluores cence. (3) If specific fluorescence is not found in the monolayers stained with the conjugate specific to the species of origin of the MCS, the cell line is un satisfactory and shall not be used for vaccine production. (4) If nonspecific fluorescence is found in the monolayers stained with conjugate from an unrelated species of origin or other results make the test results equivocal, the procedure shall be repeated until either specific fluo rescence is found only in the monolayers stained with conjugate spe cific to the species of origin of the MCS and not in the control monolayers or specific fluorescence cannot be identi fied and the MCS is declared unsatis factory. (5) Alternate tests to determine the species of origin of the MCS may be used if approved by APHIS. (c) The MCS and either each subcul ture of cells used to prepare a biologi cal product or the final pool of har vested material (with or without the stabilizer) or final container samples of completed product for each serial of such product shall be shown to be free of mycoplasma as prescribed in § 113.28. The sample for testing shall consist of at least 75 cm 2 of actively growing cells or the equivalent, in harvest VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00732 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 723 Animal and Plant Health Inspection Service, USDA § 113.52 fluids. The cells shall represent all sources of cells in the batch. (d) The MCS and either each subcul ture used to prepare a biological prod uct or the final pool of harvested mate rial for each serial of such product or final container samples of completed product for each serial of such product shall be tested for bacteria and fungi as prescribed in § 113.26 or § 113.27 (which ever is applicable). If bacteria or fungi are found in the MCS, the MCS shall not be used. If bacteria or fungi are found in a subculture, the subculture shall not be used. (e) A monolayer at least 75 cm 2 from each MCS shall be shown free of extra neous agents as prescribed in this para graph. (1) The test monolayer shall be main tained for at least 21 days using the medium (with additives) intended for growth and maintenance and under conditions similar to those used to pre pare biological products. (2) Cells shall be subcultured at least two times during the maintenance pe riod. All but the last subculture shall result in at least one new monolayer of at least 75 cm 2. The last subculture shall meet the minimum area require ment specified in §§ 113.46 and 113.47 and paragraph (f) of this section. (3) Monolayers shall be examined reg ularly throughout the 21-day mainte nance period for evidence of the pres ence of cytopathogenic agents. If evi dence of a cytopathogenic agent is found, the MCS is unsatisfactory. (4) At the conclusion of the 21-day maintenance period, monolayers shall be tested for: (i) Cytopathogenic and/or hemadsorbing agents as prescribed in § 113.46; and (ii) Extraneous agents by the fluores cent antibody technique as prescribed in § 113.47. (f) At the conclusion of the 21-day maintenance period provided in para graph (e) of this section, at least one monolayer of at least 75 cm 2 shall also be shown free of extraneous agents as prescribed in this paragraph. (1) Alternately freeze and thaw the monolayer(s) three times. Centrifuge the disrupted cells at no greater than 2,000 × g for no more than 15 minutes to remove cellular debris. Divide the su pernatant into equal aliquots and dis pense 1.0 ml onto each of at least one monolayer (at least 75 cm 2) of: (i) Vero (African green monkey kid ney) cell line; (ii) Embryonic cells, neonatal cells, or a cell line of the same species of ori gin as the MCS if different than pro vided in paragraph (f)(1)(i) of this sec tion; (iii) Embryonic cells, neonatal cells, or a cell line of the species for which the vaccine is recommended if different than provided in paragraph (f)(1)(ii) of this section; and (iv) Embryonic cells, neonatal cells, or a cell line of bovine origin if not specified in paragraphs (f)(1)(ii), and (iii) of this section. (2) The monolayers of cells specified in paragraphs (f)(1)(i), (ii), (iii), and (iv) of this section shall be maintained for at least 14 days after inoculation with the aliquot of disrupted MCS. Monolayers shall be subcultured at least once during the maintenance pe riod. All but the last subculture shall result in a new monolayer of at least 75 cm 2. The last subculture shall meet the minimum area requirement speci fied in §§ 113.46 and 113.47. (3) Monolayers shall be examined reg ularly throughout the 14-day mainte nance period for evidence of the pres ence of cytopathogenic agents. If evi dence of a cytopathogenic agent is found, the MCS is unsatisfactory. (4) At the conclusion of the 14-day maintenance period, monolayers shall be tested for: (i) Cytopathogenic and/or hemadsorbing agents as prescribed in § 113.46; and (ii) Extraneous viruses by the fluo rescent antibody technique as pre scribed in § 113.47. (g) The karyology of cells lines used in the production of biologics shall be examined as follows. A minimum of 50 mitotic cells shall be examined at both the MCS and MCS + n. The modal num ber in the MCS + n shall not exceed plus or minus 15 percent of the modal number of the MCS. Any marker chro mosomes present in the MCS shall per sist at the MCS + n. If the modal num ber exceeds the limits and/or the mark er chromosomes do not persist (through the MCS + n passage level), VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00733 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 724 9 CFR Ch. I (1–1–18 Edition) § 113.53 the cell line shall not be used for vac cine production. (h) If direct or indirect evidence ex ists that a cell line which is intended for use in the preparation of a vaccine may induce malignancies in the species for which the product is intended, that cell line shall be tested for tumorigenicity/oncogenicity by a method acceptable to APHIS. [50 FR 442, Jan. 4, 1985; 50 FR 3316, Jan. 24, 1985, as amended at 56 FR 66784, Dec. 26, 1991; 60 FR 24549, May 9, 1995] § 113.53 Requirements for ingredients of animal origin used for produc tion of biologics. Each lot of ingredient of animal ori gin which is not subjected to heat ster ilization or other sterilization methods acceptable to Animal and Plant Health Inspection Service (APHIS), such as, but not limited to serum and albumin, used to prepare a biological product shall be tested as prescribed in this sec tion by the licensee or a laboratory ac ceptable to VS. Results of all tests shall be recorded by the testing labora tory and made a part of the licensee’s records. A lot of ingredient found un satisfactory by any prescribed test shall not be used to prepare a biologi cal product. A serial of biological prod uct shall not be released if produced using an ingredient that is found un satisfactory by any prescribed test. (a) Samples of each lot of ingredient of animal origin which is not subjected to heat sterilization, used to prepare a biological product shall be shown free of mycoplasma by the method pre scribed in § 113.28. (b) Samples of each lot of ingredient or animal origin which is not subjected to heat sterilization of other steriliza tion methods acceptable to APHIS used to prepare a biological product shall be shown free of bacteria and fungi as pre scribed in § 113.26. (c) Samples of each lot of ingredient of animal origin, except porcine trypsin, which is not subjected to heat sterilization or other viricidal proce dure acceptable to APHIS used in the preparation of biological products shall be tested as prescribed in this para graph; (1) Monolayers at least 75 cm 2 of Vero (African green monkey kidney) cell line and of primary cells or a cell line of the same species of origin as the ingredient shall be used in the test. Cell lines used shall have been found satisfactory when tested as prescribed in § 113.52 and primary cells used shall have been found satisfactory when tested as prescribed in § 113.51. (2) At least 3.75 ml or 15 percent of the ingredient shall be used in the growth medium for the preparation of at least 75 cm 2 test monolayers. The ingredient shall also be used in the growth medium when monolayers are subcultured. If the ingredient being tested is cytotoxic when tested in this manner, other procedures may be used if approved by APHIS. (3) The test monolayers shall be maintained for at least 21 days. (4) Cells shall be subcultured at least two times during the maintenance pe riod. All but the last subculture shall result in at least one new monolayer of at least 75 cm 2. The last subculture shall meet the minimum area require ments specified in §§ 113.46 and 113.47. (5) Monolayers shall be examined reg ularly throughout the 21-day mainte nance period for evidence of cytopathogenic agents. If evidence of a cytopathogenic agent is found, the in gredient is unsatisfactory. (6) At the conclusion of the 21-day maintenance period, monolayers shall be tested for: (i) Cytopathogenic and/or hemadsorbing agents as prescribed in § 113.46; and (ii) Extraneous viruses by the fluo rescent antibody technique as pre scribed in § 113.47. (d) Each lot of porcine trypsin which has not been treated to inactivate por cine parvovirus (PPV) in a manner ac ceptable to VS shall be tested for PPV as prescribed in this paragraph. (1) Not less than 5.0 grams of trypsin shall be dissolved in a volume of suit able diluent sufficient to fill a cen trifuge angle head. After centrifuging for 1 hour at 80,000 × g, the pellet mate rial shall be reconstituted in distilled water and inoculated into a flask con taining 75 cm 2 of a 30 to 50 percent con fluent monolayer culture of primary porcine cells or a porcine cell line of proven equal PPV susceptibility. An VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00734 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 725 Animal and Plant Health Inspection Service, USDA § 113.55 additional flask of cells shall be held as a negative control. (2) The test and control monolayers shall be maintained for at least 14 days and subcultured at least once during the maintenance period. (3) At the end of the 14-day mainte nance period, and 4 to 7 days after the last subculturing, monolayers shall be tested for the presence of porcine parvovirus by the fluorescent antibody technique as prescribed in § 113.47(c). (e) A sample of serum from each donor horse used to produce a lot of equine serum used in the preparation of biological products recommended for use in horses shall be tested at a lab oratory approved by Animal and Plant Health Inspection Service using the Coggins test for equine infectious ane mia antibodies. If antibodies to equine infectious anemia are found, the lot of serum is unsatisfactory. [50 FR 442, Jan. 4, 1985; 50 FR 3316, Jan. 24, 1985, as amended at 56 FR 66784, Dec. 26, 1991; 60 FR 24549, May 9, 1995] § 113.54 Sterile diluent. Sterile Diluent shall be supplied in a final container by the licensee when such diluent is required for rehydration or dilution of the vaccine. (a) Sterile Diluent may be distilled or deionized water or it may be a spe cial liquid solution formulated in ac cordance with an acceptable outline on file with Animal and Plant Health In spection Service. (b) Each quantity prepared at one time in a single container and bottled into final containers shall be des ignated as a serial. Each serial shall be given a number which shall be used in records, test reports, and on the final container label. (c) Final container samples from each serial shall be tested for bacteria and fungi in accordance with the test provided in § 113.26. Any serial found to be unsatisfactory shall not be released. [39 FR 27428, July 29, 1974, as amended at 56 FR 66784, Dec. 26, 1991] § 113.55 Detection of extraneous agents in Master Seed Virus. Unless otherwise prescribed in a Standard Requirement or in a filed Outline of Production, each Master Seed Virus (MSV) shall be tested as prescribed in this section. A MSV found unsatisfactory by any prescribed test shall not be used. A serial of bio logical product shall not be released if produced from a MSV that is found un satisfactory by any prescribed test. (a) At least a 1.0 ml aliquot per cell culture of MSV shall be dispensed onto monolayers (at least 75 cm 2 in area) of: (1) Vero (African green monkey kid ney) cell line; (2) Embryonic cells, neonatal cells, or a cell line of the species for which the vaccine is recommended; and (3) Embryonic cells, neonatal cells, or a cell line of the species of cells in which the MSV is presently being prop agated if different than prescribed in paragraphs (a)(1) and (a)(2) of this sec tion. Cell lines used shall have been found satisfactory when tested as pre scribed in § 113.52 and primary cells used shall have been found satisfactory when tested as prescribed in § 113.51. If the MSV is cytopathic for or causes hemadsorption in the cells in which it is to be tested, the MSV shall be neu tralized with monospecific antiserum supplied or approved by Animal and Plant Health Inspection Service (APHIS) or counteracted by a method approved by APHIS. (b) At least one monolayer of each cell type used in the test shall be main tained as an uninoculated control. (c) Each monolayer shall be main tained at least 14 days. (d) Cells shall be subcultured at least once during the maintenance period. All but the last subculture shall result in at least one new monolayer at least 75 cm 2. The last subculture shall meet the minimum area requirement speci fied in §§ 113.46 and 113.47. (e) Monolayers shall be examined regularly throughout the 14-day main tenance period for evidence of cytopathogenic agents. If evidence of a cytopathogenic agent is found, the MSV is unsatisfactory. (f) At the conclusion of the 14-day maintenance period, monolayers shall be tested for: (1) Cytopathogenic and/or hemadsorbing agents as prescribed in § 113.46; VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00735 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 726 9 CFR Ch. I (1–1–18 Edition) § 113.64 (2) Extraneous agents by the fluores cent antibody technique as prescribed in § 113.47. [50 FR 444, Jan. 4, 1985, as amended at 56 FR 66784, Dec. 26, 1991] LIVE BACTERIAL VACCINES § 113.64 General requirements for live bacterial vaccines. When prescribed in an applicable Standard Requirement or in the filed Outline of Production, a live bacterial vaccine shall meet the requirements in this section. (a) Purity test. Final container sam ples of completed product from each se rial and subserial, and samples of each lot of Master Seed Bacteria shall be tested for the presence of extraneous viable bacteria and fungi in accordance with the test provided in § 113.27(b). (b) Safety tests. (1) Samples of com pleted product from each serial or first subserial and samples of each lot of Master Seed Bacteria shall be tested for safety in young adult mice in ac cordance with the test provided in § 113.33(b) unless: (i) The bacteria or agents in the vac cine are inherently lethal for mice. (ii) The vaccine is recommended for poultry. (2) Samples of completed product from each serial or first subserial of live bacterial vaccine shall be tested for safety in one of the species for which the product is recommended as follows: (i) Live bacterial vaccine rec ommended for use in dogs shall be test ed as provided in § 113.40, except that dogs shall be injected with the equiva lent of two doses of vaccine adminis tered as recommended on the label. (ii) Live bacterial vaccine rec ommended for use in cattle shall be tested as provided in § 113.41, except that calves shall be injected with the equivalent of two doses of vaccine ad ministered as recommended on the label. (iii) Live bacterial vaccine rec ommended for use in sheep shall be tested as provided in § 113.45. (iv) Live bacterial vaccine rec ommended for use in swine shall be tested as provided in § 113.44. (c) Identity test. At least one of the identity tests provided in this para graph shall be conducted for the Mas ter Seed Bacteria and final container samples from each serial or first sub serial of completed biological product. A known positive control (reference) provided or approved by Animal and Plant Health Inspection Service shall be included in such tests. (1) Fluorescent antibody test. The di rect fluorescent antibody staining technique shall be conducted using suitable smears of the vaccine bac teria. Fluorescence typical for the bac teria concerned shall be demonstrated. Fluorescence shall not occur in control smears treated with specific antiserum. (2) Tube agglutination test. A tube ag glutination test shall be conducted with a suitable suspension of the vac cine bacteria using the constant anti gen decreasing serum method with spe cific antiserum. Agglutination typical for the bacteria shall be demonstrated. Agglutination shall not occur with negative serum used as a control in this test. (3) Slide agglutination test. The rapid plate (slide) agglutination test shall be conducted with suitable suspensions of the vaccine bacteria using the hanging drop, slide or plate method, with spe cific antiserum. Agglutination typical for the bacteria shall be demonstrated by microscopic or macroscopic obser vation. Agglutination shall not occur with negative serum used as a control in this test. (4) Characterization tests. Applicable biochemical and cultural characteris tics shall be demonstrated as specified in the filed Outline of Production. (d) Ingredient requirements. Ingredi ents used for the growth and prepara tion of Master Seed Bacteria and of live bacterial vaccine shall meet the requirements provided in § 113.50. Ingre dients of animal origin shall meet the applicable requirements provided in § 113.53. (e) Moisture content. The maximum percent moisture in desiccated vac cines shall be stated in the filed Out line of Production and shall be estab lished by the licensee as follows: (1) Prelicensing. Data obtained by con ducting accelerated stability tests and VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00736 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 727 Animal and Plant Health Inspection Service, USDA § 113.65 bacterial counts shall be acceptable on a temporary basis. (2) Licensed products. Data shall be obtained by determining the percent moisture and bacterial count at release and expiration on a minimum of 10 con secutive released serials. (3) Final container samples of com pleted product from each serial and subserial must be tested for moisture content in accordance with the test provided in § 113.29. [48 FR 33476, July 22, 1983, as amended at 54 FR 19352, May 5, 1989; 56 FR 66784, Dec. 26, 1991; 68 FR 57608, Oct. 6, 2003] § 113.65 Brucella Abortus Vaccine. Brucella Abortus Vaccine shall be prepared as a desiccated live culture bacterial vaccine from smooth colonial forms of the Brucella abortus organism, identified as Strain 19. Each serial and subserial shall be tested for purity, po tency, and moisture content. A serial or subserial found unsatisfactory by a prescribed test shall not be released. (a) Purity tests. Each serial and sub serial shall be tested for purity as pro vided in this paragraph. (1) Macroscopic and microscopic ex amination shall be made on bulk sam ples from production containers. If or ganisms not typical of Brucella abortus organisms are evident, the serial or subserial is unsatisfactory. (2) Two final container vials of com pleted product shall be tested by inoculating one tube of Dextrose Andrades broth with gas tube and one tube of thioglycollate broth from each vial. The inoculated media shall be in cubated at 35 to 37 °C for 96 hours. If growth not typical of Brucella abortus organisms is evident, the serial or sub serial is unsatisfactory. (3) Bacterial dissociation test. Final container samples of completed prod uct from each serial and subserial shall be tested for bacterial dissociation. Smooth colonies are the desired form. Rough colonies are undesirable ter minal dissociation forms. Intermediate and intermediate-to-rough are also un desirable. (i) The sample container shall be re hydrated and streaked on one potato agar plate in such a manner as to produce confluent colonies. Artificial reflected light shall be used so that the rays pass through the plate at a 45 °angle. (ii) If the vaccine contains more than 5 percent rough colonies or more than 15 percent total undesirable colonies, the serial or subserial is unsatisfac tory. If organisms or growth not char acteristic of Brucella abortus are found, the serial or subserial is unsatisfac tory. The test may be repeated one time using double the number of sam ples: Provided, That, if the test is not repeated, the serial or subserial is un satisfactory. (b) Bacterial count requirements for re duced dose vaccine. Each serial and each subserial shall be tested for potency. (1) Two final container vials of com pleted product shall be tested for the number of viable organisms per dose of rehydrated vaccine. A bacterial count per vial shall be made on tryptose agar plates from suitable dilutions using 1 percent peptone as a diluent. The in oculated media shall be incubated at 35 to 37 °C for 96 hours. (2) If the average count of the two final container samples of freshly pre pared vaccine contains less than 3.0 or more than 10.0 billion organisms per dose, the serial or subserial is unsatis factory. (3) If the average count on the initial test is less than the minimum or great er than the maximum required in para graph (b)(2) of this section, the serial or subserial may be retested one time using four additional final container vials. The average count of the retest is determined. If the average count of the four vials retested is less than the required minimum or greater than the required maximum, the serial or sub serial is unsatisfactory. If the average count of the four vials retested is with in the required limits described in paragraph (b)(2) of this section, the fol lowing shall apply: (i) If the average count obtained in the initial test is less than one-third or more than three times the average count obtained on the retest, the aver age count of the initial test shall be considered the result of test system error and the serial or subserial is sat isfactory. (ii) If the average count obtained in the initial test is one-third or more than the average retest count or three VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00737 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 728 9 CFR Ch. I (1–1–18 Edition) § 113.66 times or less than the average retest count, a new average count shall be de termined from the counts of all six vials. If the new average is less than the minimum or greater than the max imum required in paragraph (b)(2) of this section, the serial or subserial is unsatisfactory. (4) If tested at any time within the expiration period, each dose of re hydrated vaccine must contain at least 3.0 billion viable organisms per dose. (c) Bacterial count requirements for standard vaccine. Each serial and sub serial shall be tested for potency. (1) Two final container samples shall be tested for the number of viable orga nisms per milliliter of rehydrated vac cine. One bacterial count per vial shall be made on tryptose agar plates from suitable dilutions using 1 percent pep tone as a diluent. The inoculated media shall be incubated at 35 to 37 °C for 96 hours. (2) If the average count of the two final container samples of freshly pre pared vaccine does not contain at least 10 billion viable organisms per milli liter, the serial or subserial is unsatis factory. (3) If the initial bacterial count is less than 10 billion organisms per milli liter, the serial or subserial may be re tested one time using four samples. If the average count of the four vials re tested is less than the required min imum, the serial or subserial is unsat isfactory. (4) If tested at any time within the expiration period, each milliliter of re hydrated vaccine does not contain at least 5 billion viable organisms per milliliter, the serial or subserial is un satisfactory. [39 FR 16857, May 10, 1974. Redesignated at 39 FR 25463, July 11, 1974, and amended at 40 FR 758, Jan. 3, 1975; 50 FR 23794, Jan. 6, 1985] § 113.66 Anthrax Spore Vaccine—Non encapsulated. Anthrax Spore Vaccine—Nonencap sulated shall be a live spore suspension prepared from nonencapsulated variants of Bacillus anthracis. Only Master Seed which has been estab lished as pure, safe, and immunogenic shall be used for production. All serials of vaccine shall be prepared from the first through the fifth passage from the Master Seed. (a) The Master Seed shall meet the applicable general requirements pre scribed in § 113.64 and the requirements in this section. (b) Each lot of Master Seed shall be tested for immunogenicity as follows: (1) Forty-two susceptible guinea pigs from the same source each weighing 400 to 500 grams, shall be used as test ani mals (30 vaccinates and 12 controls). (2) An arithmetic mean spore count of vaccine produced from the highest passage of the Master Seed shall be es tablished before the immunogenicity test is conducted. The guinea pigs used as vaccinates shall be injected as rec ommended on the label with a pre determined number of vaccine spores. To confirm the dosage, five replicate spore counts shall be conducted on a sample of the vaccine dilution used. (3) Fourteen to fifteen days postvaccination the vaccinates and controls shall each be challenged with not less than 4,500 guinea pig LD50 of a virulent suspension of Bacillus anthracis furnished or approved by Ani mal and Plant Health Inspection Serv ice and observed for 10 days. (4) If at least 10 of the 12 controls do not die from Bacillus anthracis within the 10-day postchallenge observation period the test is invalid and may be repeated. (5) If at least 27 of 30 of the vac cinates do not survive the 10-day postchallenge observation period, the Master Seed is unsatisfactory. (6) An Outline of Production change shall be made before authority for use of a new lot of Master Seed shall be granted by Animal and Plant Health Inspection Service. (c) Test Requirements for Release. Each serial and subserial shall meet the ap plicable general requirements pre scribed in 9 CFR 113.64 and the require ments in this paragraph. Any serial or subserial found unsatisfactory by a prescribed test shall not be released. (1) Safety test. Samples of completed product from each serial or first sub serial shall be tested for safety in sheep or goats by the methods described in 9 CFR 113.45(a). VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00738 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 729 Animal and Plant Health Inspection Service, USDA § 113.67 (2) Spore Count Requirements. Final container samples of completed prod uct shall be tested for spore count. Samples shall be diluted in tenfold steps. Each dilution expected to yield 30 to 300 colonies per plate shall be plated in triplicate on tryptose agar, inverted, and incubated at 35 to 70 °C for 24 hours to 28 hours. Each plate having uniformly distributed colonies shall be counted and an average count determined. To be eligible for release, each serial and each subserial shall have a spore count sufficiently greater than that of the vaccine used in the immunogenicity test to assure that when tested at any time within the ex piration period, each serial and sub serial shall have a spore count of at least twice that used in the immunogenicity test but not less than 2,000,000 spores per dose. [50 FR 23794, June 6, 1985, as amended at 56 FR 66784, Dec. 26, 1991; 72 FR 72564, Dec. 21, 2007] § 113.67 Erysipelothrix Rhusiopathiae Vaccine. Erysipelothrix Rhusiopathiae Vac cine shall be prepared as a desiccated live culture of an avirulent or modified strain of Erysipelothrix rhusiopathiae. Only Master Seed which has been es tablished as pure, safe, and immunogenic shall be used for vaccine production. (a) The Master Seed shall meet the applicable requirements prescribed in § 113.64 and the requirements in this section. (b) Each lot of Master Seed used for vaccine production shall be tested for immunogenicity. The selected bac terial count from the lot of Master Seed shall be established as follows: (1) Thirty Erysipelothrix rhusiopathiae susceptible swine shall be used as test animals (20 vaccinates and 10 controls) for each route of administration rec ommended on the label. (2) An arithmetic mean count of the colony forming units from vaccine pro duced from the highest passage of the Master Seed shall be established before the immunogenicity test is conducted. The 20 swine to be used as vaccinates shall be injected as recommended on the label with a predetermined quan tity of vaccine bacteria. The 10 control swine shall be held separately from the vaccinates. To confirm the dosage cal culation, an arithmetic mean count shall be established by conducting five replicate titrations on a sample of the bacterial vaccine dilution used. Only plates containing between 30 and 300 colonies shall be considered in a valid test. (3) The vaccinates and controls shall be examined and their average body temperature determined prior to chal lenge. Fourteen to twenty-one days postvaccination, the vaccinates and controls shall be challenged with a vir ulent Erysipelothrix rhusiopathiae cul ture and observed for 7 days. The chal lenge culture and instructions for prep aration and use shall be obtained from Animal and Plant Health Inspection Service. (4) A satisfactory challenge shall be evidenced in the controls by a high body temperature or clinical signs in cluding, but not limited to acute ill ness with hyperemia of the abdomen and ears, possibly terminating in sud den death; moribundity, with or with out metastatic skin lesions; depression with anorexia, stiffness, and/or joint involvement; or any combination of these symptoms and lesions. (5) If at least 80 percent of the con trols do not show characteristic signs during the observation period includ ing, but not limited to a body tempera ture of 105.6 °F or higher on at least 2 consecutive days, the test shall be con sidered a No Test: Provided, That con trol pigs which meet the criteria re quirements for susceptibility except for high body temperature shall be con sidered susceptible if sacrificed and or ganisms identified as Erysipelothrix rhusiopathiae can be isolated from the blood, spleen, or other organs. (6) To demonstrate immunity after challenge, the vaccinates shall remain free of clinical signs and the body tem perature shall not exceed 104.6 °F on 2 or more consecutive days. If at least 90 percent of the vaccinates do not re main free from clinical signs and high body temperature throughout the ob servation period, the Master Seed is unsatisfactory. (7) An Outline of Production change shall be made before authority for use of a new Master Seed shall be granted VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00739 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 730 9 CFR Ch. I (1–1–18 Edition) § 113.68 by Animal and Plant Health Inspection Service. (c) Test requirements for release. Each serial and subserial shall meet the ap plicable requirements in § 113.64 and the requirements in this paragraph. Any serial or subserial found unsatisfactory by a prescribed test shall not be re leased. (1) Safety test. Samples of completed product from each serial or first sub serial shall be tested for safety in young adult mice as prescribed in § 113.33(b) and in swine as prescribed in § 113.44. (2) Bacterial count requirements. Final container samples of completed prod uct from each serial and each subserial shall be tested for bacterial count using the method used in paragraph (b)(2) of this section. Two replicate ti trations shall be conducted on each sample. To be eligible for release, each serial and subserial shall have a bac terial count sufficiently greater than that of the vaccine used in the immunogenicity test to assure that, when tested at any time within the ex piration period, each serial and sub serial shall have a bacterial count two times greater than that used in such immunogenicity test. [50 FR 23795, June 6, 1985, as amended at 56 FR 66784, Dec. 26, 1991; 72 FR 72564, Dec. 21, 2007] § 113.68 Pasteurella Haemolytica Vac cine, Bovine. Pasteurella Haemolytica Vaccine, Bovine, shall be prepared as a des iccated live culture bacterial vaccine of an avirulent or modified strain of Pasteurella haemolytica, identified as serotype 1. Only Master Seed which has been established as pure, safe, and immunogenic shall be used for vaccine production. All serials of vaccine shall be prepared from the first through the fifth passage from the Master Seed. (a) The Master Seed shall meet the applicable general requirements pre scribed in § 113.64 and the requirements in this section. (b) Each lot of Master Seed used for vaccine production shall be tested for immunogenicity. The immunogenicity of a selected bacterial count from the lot of Master Seed shall be established as follows: (1) Fifteen Pasteurella haemolytica susceptible calves shall be used as test animals (10 vaccinates and 5 controls) for each route of administration rec ommended on the label. (2) An arithmetic mean count of the colony forming units from vaccine pro duced from the highest passage of the Master Seed shall be established before the immunogenicity test is conducted. The 10 calves to be used as vaccinates shall be injected as recommended on the label with a predetermined quan tity of vaccine bacteria. The five con trol calves shall be held separately from the vaccinates. To confirm the dosage calculation, five replicate titra tions on a sample of the bacterial vac cine used. Only plates containing be tween 30 and 300 colonies shall be con sidered a valid test. (3) The vaccinates and controls shall be examined and their average body temperature determined prior to chal lenge. Fourteen to twenty-one days post vaccination, the vaccinates and controls shall each be challenged by the respiratory route with a (virulent) pneumonia producing Pasteurella haemolytica culture and observed for 4 to 7 days. The challenge culture and in structions for preparation for use shall be furnished or approved by the Animal and Plant Health Inspection Service. (4) A satisfactory challenge shall be evidenced in the controls by progres sion of clinical signs consistent with respiratory system infection following challenge, including but not limited to lacrimation, mucoid nasal exudates, expiratory dyspnea, tachypnea, pul monary rales, and cough possibly ter minating in death; moribundity, de pression with anorexia, diarrhea with substantial weight loss; or any com bination of these symptoms. (5) Lung lesion response to challenge will be assessed in all calves. Lung le sions will be assessed at necropsy in calves that succumb to challenge. Sur viving calves will be euthanized on day 4 to 7 following challenge and lung le sions assessed at necropsy. Lung lesion scores will be used in the assessment of the response to challenge exposure. If a significant difference in lung lesion scores cannot be demonstrated between vaccinates and controls using a scoring system approved by the Animal and VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00740 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 731 Animal and Plant Health Inspection Service, USDA § 113.69 Plant Health Inspection Service, the Master Seed is unsatisfactory. (6) An Outline of Production change must be made before authority for use of a new lot of Master Seed is granted by the Animal and Plant Health In spection Service. (c) Test requirements for release. Each serial and subserial shall meet the ap plicable general requirements pre scribed in §§ 113.8 and 113.64 and the re quirements in this paragraph. Any se rial or subserial found unsatisfactory by a prescribed test shall not be re leased. (1) Safety test. Samples of completed product from each serial or first sub serial shall be tested for safety in calves as provided in §§ 113.41(a) and 113.41(b) except, that the equivalent of two doses of vaccine shall be used and administered in the manner rec ommended on the label. (2) Bacterial count requirements. Final container samples of completed prod uct shall be tested for bacterial count using the method used in paragraph (b)(2) of this section. Two replicate ti trations shall be conducted on each se rial and subserial. Each sample shall be rehydrated with accompanying sterile diluent to the volume indicated on the label. To be eligible for release, each serial and subserial shall have a bac terial count sufficiently greater than that of the vaccine used in the immunogenicity test to assure that, when tested at any time within the ex piration period, each serial and sub serial shall have a bacterial count at least two times greater than that used in the immunogenicity test. [55 FR 35559, Aug. 31, 1990, as amended at 72 FR 72564, Dec. 21, 2007] § 113.69 Pasteurella Multocida Vac cine, Bovine. Pasteurella Multocida Vaccine, Bo vine, shall be prepared as a desiccated live culture bacterial vaccine of an avirulent or modified strain of Pasteurella multocida, of bovine origin. Only Master Seed which has been es tablished as pure, safe, and immunogenic shall be used for vaccine production. All serials of vaccine shall be prepared from the first through the fifth passage from the Master Seed. (a) The Master Seed shall meet the applicable general requirements pre scribed in § 113.64 and the requirements in this section. (b) Each lot of Master Seed used for vaccine production shall be tested for immunogenicity. The immunogenicity of a selected bacterial count from the lot of Master Seed shall be established as follows: (1) Fifteen Pasteurella multocida sus ceptible calves shall be used as test animals (10 vaccinates and 5 controls) for each route of administration rec ommended on the label. (2) An arithmetic mean count of the colony forming units from vaccine pro duced from the highest passage of the Master Seed shall be established before the immunogenicity test is conducted. The 10 calves to be used as vaccinates shall be injected as recommended on the label with a predetermined quan tity of vaccine bacteria. The five con trol calves shall be held separately from the vaccinates. To confirm the dosage calculation, arithmetic mean count shall be established by con ducting five replicate titrations on a sample of the bacterial vaccine used. Only plates containing between 30 and 300 colonies shall be considered a valid test. (3) The vaccinates and controls shall be examined and their average body temperature determined prior to chal lenge. Fourteen to twenty-one days post vaccination, the vaccinates and controls shall each be challenged by the respiratory route with a (virulent) pneumonia producing Pasteurella multocida culture and observed for 4 to 10 days. The challenge culture and in structions for preparation for use shall be furnished or approved by the Animal and Plant Health Inspection Service. (4) A satisfactory challenge shall be evidenced in the controls by progres sion of clinical signs consistent with respiratory system infection following challenge, including but not limited to acute illness with higher body tem perature and respiration rate, lacrimation, mucoid nasal exudate, ex piratory dyspnea, tachypnea, pul monary rales, and cough, possibly ter minating in death; moribundity, de pression with anorexia; diarrhea with VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00741 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 732 9 CFR Ch. I (1–1–18 Edition) § 113.70 substantial weight loss; or any com bination of these symptoms. (5) Lung lesion response to challenge will be assessed in all calves. Lung le sions will be assessed at necropsy in calves that succumb to challenge. Sur viving calves will be euthanized on day 4 to 10 following challenge and lung le sions assessed at necropsy. Lung lesion scores will be used in the assessment of the response to challenge exposure. If a significant difference in lung lesion scores cannot be demonstrated between vaccinates and controls using a scoring system approved by the Animal and Plant Health Inspection Service, the Master Seed is unsatisfactory. (6) An Outline of Production change must be made before authority for use of a new lot of Master Seed is granted by the Animal and Plant Health In spection Service. (c) Test requirements for release. Each serial and subserial shall meet the ap plicable general requirements pre scribed in §§ 113.8 and 113.64 and the re quirements in this paragraph. Any se rial or subserial found unsatisfactory by a prescribed test shall not be re leased. (1) Safety Test. Samples of completed product from each serial or first sub serial shall be tested for safety in calves as provided in §§ 113.41(a) and 113.41(b), except that the equivalent of two doses of vaccine shall be used and administered in the manner rec ommended on the label. (2) Bacterial count requirements. Final container samples of completed prod uct shall be tested for bacterial count using the method used in paragraph (b)(2) of this section. Two replicate ti trations shall be conducted on each se rial and subserial. Each sample shall be rehydrated with accompanying sterile diluent to the volume indicated on the label. To be eligible for release, each serial and subserial shall have a bac terial count sufficiently greater than that of the vaccine used in the immunogenicity test count per dose es tablished to assure that, when tested at any time within the expiration pe riod, each serial and subserial shall have a bacterial count at least two times greater than that used in the immunogenicity test. [55 FR 35560, Aug. 31, 1990, as amended at 72 FR 72564, Dec. 21, 2007] § 113.70 Pasteurella Multocida Vac cine, Avian Isolate. Pasteurella Multocida Vaccine, Avian Isolate, shall be prepared as a desiccated live culture of an avirulent or modified strain of Pasteurella multocida. Only Master Seed which has been established as pure, safe, and immunogenic shall be used for vaccine production. (a) The Master Seed shall meet the applicable general requirements pre scribed in § 113.64 and the requirements in this section. (b) Each lot of Master Seed used for vaccine production shall be tested for immunogenicity in each species and for each serotype for which the Master Seed is claimed to give protection. (1) Thirty Pasteurella multocida sus ceptible birds shall be used as test ani mals (20 vaccinates and 10 controls) for each bird species, route of administra tion, and serotype for which protection is claimed on the label. (2) An arithmetic mean count of col ony forming units from vaccine pro duced from the highest passage of Mas ter Seed shall be established before the immunogenicity test is conducted. The 20 birds to be used as vaccinates shall be inoculated, as recommended on the label with a predetermined quantity of vaccine bacteria. The 10 control birds shall be held separately from the vac cinates. To confirm the dosage calcula tion, an arithmetic mean count shall be established by conducting five rep licate titrations on a sample of the bacterial vaccine used. Only plates containing between 30 and 300 colonies shall be considered in a valid test. (3) Not less than 14 days after vac cination, each of 20 vaccinates and each of 10 unvaccinated controls shall be challenged intramuscularly or by other methods acceptable to the Ani mal and Plant Health Inspection Serv ice with a virulent Pasteurella multocida strain, for which protection is claimed, and observed daily for a 14 day post challenge period. (4) Eight or more of the unvaccinated controls must die for the test to be VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00742 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 733 Animal and Plant Health Inspection Service, USDA § 113.71 valid. If at least 16 of 20 of the vac cinates do not survive the 14-day postchallenge period, the Master Seed is unsatisfactory at the selected bac terial count. (c) Test requirements for release. Each serial and subserial shall meet the ap plicable requirements in §§ 113.8 and 113.64 and the requirements in this paragraph. Any serial or subserial found unsatisfactory by a prescribed test shall not be released. (1) Safety test. Samples of completed product from each serial or first sub serial shall be tested for safety. (i) Ten birds of a species for which the vaccine is recommended shall be given the equivalent of 10 doses each of the vaccine and observed for 10 days. If the vaccine is recommended for more than one species, only one species needs to be tested. (ii) If unfavorable reactions attrib utable to the vaccine occur during the observation period in two or more of the test birds, the serial is unsatisfac tory. (iii) If unfavorable reactions occur which are not attributable to the test vaccine, the test is a No Test and may be repeated. If the results of the next test are not satisfactory, or if the test is not repeated, the serial shall be con sidered unsatisfactory. (2) Bacterial count requirements. Final container samples of completed prod uct shall be tested for bacterial count using the method used in paragraph (b)(2) of this section. Two replicate ti trations shall be conducted on each se rial and subserial. Each sample shall be rehydrated with accompanying sterile diluent to the volume indicated on the label. To be eligible for release, each serial and subserial shall have a bac terial count sufficiently greater than that of the vaccine used in the immunogenicity test count per dose es tablished to assure that, when tested at any time within the expiration pe riod, each serial and subserial shall have a bacterial count at least two times greater than that used in the immunogenicity test. [55 FR 35560, Aug. 31, 1990, as amended at 59 FR 19633, Apr. 25, 1994; 64 FR 43044, Aug. 9, 1999; 72 FR 72564, Dec. 21, 2007] § 113.71 Chlamydia Psittaci Vaccine (Feline Pneumonitis), Live Chlamydia. Chlamydia Psittaci Vaccine (Feline Pneumonitis), Live Chlamydia, shall be prepared from chlamydia-bearing cell culture fluids or embryonated chicken eggs. Only Master Seed which has been established as pure, safe, and immunogenic shall be used for vaccine production. All serials of vaccine shall be prepared from the first through the fifth passage from the Master Seed. (a) The Master Seed shall meet the applicable requirements prescribed in § 113.300 and the requirements in this section. Master Seed propagated in chicken embryos shall be tested for pathogens by the chicken embryo test prescribed in § 113.37. If found unsatis factory by any prescribed test, the Master Seed shall not be used. (b) Each lot of Master Seed used for vaccine production shall be tested for immunogenicity. The immunogenicity of a selected dose from the lot of Mas ter Seed shall be established as follows: (1) Thirty feline pneumonitis suscep tible cats shall be used as test animals (20 vaccinates and 10 controls). Blood samples shall be drawn and individual serum samples tested. The cats shall be considered suitable for use if all serums are negative for pneumonitis antibody in a complement fixation test or other test of equal sensitivity. (2) A geometric mean titer of the dried vaccine produced from the high est passage of the Master Seed shall be established before the immunogenicity test is conducted. The 20 cats used as vaccinates shall be administered a pre determined quantity of vaccine by the method to be recommended on the label and the remaining 10 cats shall be held as controls. To confirm the dosage calculations, five replicate titrations shall be conducted on a sample of the vaccine dilution used. If two doses are used, five replicate confirming titra tions shall be conducted on each dose. (3) Fourteen or more days after the final dose of vaccine, the vaccinates and controls shall each be challenged intranasally with a minimum of 10,000 yolk sac LD50 of virulent feline pneu monitis furnished or approved by the Animal and Plant Health Inspection Service and observed each day for 28 VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00743 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 734 9 CFR Ch. I (1–1–18 Edition) § 113.100 days postchallenge. The rectal tem perature of each animal shall be taken and the presence or absence of clinical signs noted and recorded each day. (i) If less than 8 of 10 controls show clinical signs of feline pneumonitis in fection other than fever, the test is a No Test and may be repeated. (ii) If a significant difference in clin ical signs other than fever or chlamydia shedding cannot be dem onstrated between vaccinates and con trols using a scoring system approved by the Animal and Plant Health In spection Service, the Master Seed is unsatisfactory. (4) An Outline of Production change must be made before authority for use of a new lot of Master Seed is granted by the Animal and Plant Health In spection Service. (c) Test requirements for release: Ex cept for § 113.300(a)(3)(ii), each serial and subserial shall meet the require ments prescribed in § 113.300 and in this paragraph. Final container samples of completed product shall be tested. Any serial or subserial found unsatisfactory by a prescribed test shall not be re leased. (1) The test for pathogens prescribed in § 113.37 shall be conducted on each serial or one subserial of avian origin vaccine. (2) Chlamydia titer requirements. Final container samples of completed prod uct shall be tested for chlamydia titer using the titration method used in paragraph (b)(2) of this section. To be eligible for release, each serial and each subserial shall have a titer suffi ciently greater than the titer of vac cine used in the immunogenicity test prescribed in paragraph (b) of this sec tion to assure that when tested at any time within the expiration period, each serial and subserial shall have a titer 0.7 greater than that used in such immunogenicity test but not less than 2.5 ID50 per dose. [55 FR 35561, Aug. 31, 1990, as amended at 56 FR 66786, Dec. 26, 1991; 72 FR 72564, Dec. 21, 2007] INACTIVATED BACTERIAL PRODUCTS § 113.100 General requirements for in activated bacterial products. Unless otherwise prescribed in an ap plicable Standard Requirement or in the filed Outline of Production, an in activated bacterial product shall meet the applicable requirements in this sec tion. (a) Purity tests. (1) Final container samples of completed product from each serial and each subserial shall be tested for viable bacteria and fungi as provided in § 113.26. (2) Each lot of Master Seed Bacteria shall be tested for the presence of ex traneous viable bacteria and fungi in accordance with the test provided in § 113.27(d). (b) Safety tests. Bulk or final con tainer samples of completed product from each serial shall be tested for safety in young adult mice in accord ance with the test provided in § 113.33(b) unless: (1) The product contains material which is inherently lethal for mice. In such instances, the guinea pig safety test provided in § 113.38 shall be con ducted in place of the mouse safety test. (2) The product is recommended for poultry. In such instances, the product shall be safety tested in poultry as de fined in the specific Standard Require ment or Outline of Production for the product. (3) The product is recommended for fish, other aquatic species, or reptiles. In such instances, the product shall be safety tested in fish, other aquatic spe cies, or reptiles as required by specific Standard Requirement or Outline of Production for the product. (c) Identity test. Methods of identi fication of Master Seed Bacteria to the genus and species level by laboratory tests shall be sufficient to distinguish the bacteria from other similar bac teria according to criteria described in the most recent edition of ‘‘Bergey’s Manual of Systematic Bacteriology’’ or the American Society for Microbiology ‘‘Manual of Clinical Microbiology’’. If Master Seed Bacteria are referred to by serotype, serovar, subtype, pilus type, strain or other taxonomic subdivision VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00744 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 735 Animal and Plant Health Inspection Service, USDA § 113.101 1 The procedures for performing the ferric chloride test for residual free formaldehyde may be obtained from USDA, APHIS, Center for Veterinary Biologics-Laboratory, 1800 Dayton Road, P.O. Box 844, Ames, IA 50010. below the species level, adequate test ing must be used to identify the bac teria to that level. Tests which may be used to identify Master Seed Bacteria include, but are not limited to: (1) Cultural characteristics, (2) Staining reaction, (3) Biochemical reactivity, (4) Fluorescent antibody tests, (5) Serologic tests, (6) Toxin typing, (7) Somatic or flagellar antigen char acterization, and (8) Restriction endonuclease analysis. (d) Ingredient requirements. Ingredi ents used for the growth and prepara tion of Master Seed Bacteria and of final product shall meet the require ments provided in § 113.50. Ingredients of animal origin shall meet the appli cable requirements provided in § 113.53. (e) Only serials tested for viricidal activity in accordance with the test provided in § 113.35 and found satisfac tory by such test shall be packaged as diluent for desiccated fractions in com bination packages. (f) If formaldehyde is used as the in activating agent, and the serial has not been found satisfactory by the viricidal activity test, bulk or final container samples of completed product from each serial must be tested for residual free formaldehyde content using the ferric chloride test. 1 Firms currently using tests for residual free formalde hyde content other than the ferric chloride test have until July 14, 2004 to update their Outline of Production to be in compliance with this require ment. (1) The residual free formaldehyde content of biological products con taining clostridial antigens must not exceed 1.85 grams per liter (g/L). (2) The residual free formaldehyde content of bacterins, bacterin-toxoids, and toxoids, other than those con taining clostridial antigens, must not exceed 0.74 grams per liter (g/L). [39 FR 16862, May 10, 1974. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 60 FR 14355, Mar. 17, 1995; 68 FR 35283, June 13, 2003; 79 FR 31021, May 30, 2014] § 113.101 Leptospira Pomona Bacterin. Leptospira Pomona Bacterin shall be produced from a culture of Leptospira pomona which has been inactivated and is nontoxic. Each serial of biological product containing Leptospira pomona fraction shall meet the applicable re quirements in § 113.100 and shall be tested for purity, safety, and potency as prescribed in this section. A serial found unsatisfactory by any prescribed test shall not be released. (a) Purity test. Final container sam ples of completed product from each se rial and each subserial shall be tested for viable bacteria and fungi as pro vided in § 113.26. (b) Safety test. Bulk or final container samples of completed product from each serial shall be tested for safety as provided in § 113.38. (c) Potency test. Bulk or final con tainer samples of completed product shall be diluted with physiological sa line so that each 0.25 ml contains not more than 1/800th of the dose rec ommended on the label and shall be tested for potency, using the two-stage test provided in this paragraph. (1) Vaccinates. Inject each of at least 10 but not more than 12 young adult hamsters, each weighing 50 to 90 grams, with 0.25 ml of the diluted bacterin either subcutaneously or intramuscularly, in accordance with the label recommendations for use. (2) Controls. Retain at least 10 but not more than 12 additional hamsters from the same group as unvaccinated con trols. (3) Challenge. From 14 to 18 days postvaccination, challenge each of 10 vaccinates and each of 10 controls intraperitoneally with a suspension of virulent Leptospira pomona organisms, using a dose of 10–10,000 hamster LD50 as determined by titration. (4) Post-challenge period. Observe the vaccinates and controls for 14 days post-challenge and record all deaths. If eight or more controls die of lepto spirosis, the test is valid and the re sults shall be evaluated according to the following table: VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00745 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 736 9 CFR Ch. I (1–1–18 Edition) § 113.102 Stage Number of vac cinates Cumu lative number of vac cinates Cumulative total dead hamsters for satisfactory serial Cumulative total dead hamsters for unsatisfactory serial 1 .......... 10 ........ 10 ........ 2 or less ..... 5 or more. 2 .......... 10 ........ 20 ........ 5 or less ..... 6 or more. (5) If three or four vaccinates die in the first stage, the second stage shall be conducted in a manner identical to the first stage. (6) If the second stage is used, each serial shall be evaluated according to the second part of the table. On the basis of cumulative results, each serial shall either pass or fail. [39 FR 16862, May 10, 1974, as amended at 40 FR 20067, May 8, 1975; 45 FR 40100, June 13, 1980. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66785, Dec. 26, 1991] § 113.102 Leptospira Icterohaemorrhagiae Bacterin. Leptospira Icterohaemorrhagiae Bacterin shall be produced from a cul ture of Leptospira icterohaemorrhagiae which has been inactivated and is nontoxic. Each serial of biological product containing Leptospira icterohaemorrhagiae fraction shall meet the applicable requirements in § 113.100 and be tested for purity, safety, and po tency as prescribed in this section. A serial found unsatisfactory by any pre scribed test shall not be released. (a) Purity test. Final container sam ples of completed product from each se rial and each subserial shall be tested for viable bacteria and fungi as pro vided in § 113.26. (b) Safety test. Bulk or final container samples of completed product from each serial shall be tested for safety as provided in § 113.38. (c) Potency test. Bulk or final con tainer samples of completed product shall be diluted with physiological sa line so that each 0.25 ml contains not more than 1/80th of the dose rec ommended on the label and shall be tested for potency, using the two-stage test provided in this paragraph. (1) Vaccinates. Inject each of at least 10 but not more than 12 young adult hamsters, each weighing 50 to 90 grams, with 0.25 ml of the diluted bacterin either subcutaneously or intramuscularly, in accordance with the label recommendations for use. (2) Controls. Retain at least 10 but not more than 12 additional hamsters from the same group as unvaccinated con trols. (3) Challenge. From 14 to 18 days postvaccination, challenge each of 10 vaccinates and each of 10 controls intraperitoneally with a suspension of virulent Leptospira icterohaemorrhagiae organisms, using a dose of 10–10,000 hamster LD50 as determined by titra tion. (4) Post-challenge period. Observe the vaccinates and controls for 14 days post-challenge and record all deaths. If eight or more controls die from lepto spirosis, the test is valid and the re sults shall be evaluated according to the following table: Stage Number of vac cinates Cumu lative number of vac cinates Cumulative total dead hamsters for satisfactory serial Cumulative total dead hamsters for unsatisfactory serial 1 .......... 10 ........ 10 ........ 2 or less ..... 5 or more. 2 .......... 10 ........ 20 ........ 5 or less ..... 6 or more. (5) If three or four vaccinates die in the first stage, the second stage shall be used. The second stage shall be con ducted in a manner identical to the first stage. (6) If the second stage is used, each serial shall be evaluated according to the second part of the table. On the basis of cumulative results, each serial shall either pass or fail. [39 FR 16862, May 10, 1974, as amended at 45 FR 40100, June 13, 1980. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66785, Dec. 26, 1991] § 113.103 Leptospira Canicola Bacterin. Leptospira Canicola Bacterin shall be produced from a culture of Leptospira canicola which has been inactivated and is nontoxic. Each serial of biologi cal product containing Leptospira canic ola fraction shall meet the applicable requirements in § 113.100 and shall be tested for purity, safety, and potency as prescribed in this section. Serials found unsatisfactory by any prescribed test shall not be released. (a) Purity test. Final container sam ples of completed product from each se rial and each subserial shall be tested for viable bacteria and fungi as pro vided in § 113.26. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00746 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 737 Animal and Plant Health Inspection Service, USDA § 113.104 (b) Safety test. Bulk or final container samples of completed product from each serial shall be tested for safety as provided in § 113.38. (c) Potency test. Bulk or final con tainer samples of completed product shall be diluted with physiological sa line so that each 0.25 ml contains not more than 1/80th of the dose rec ommended on the label and shall be tested for potency, using the two-stage test provided in this paragraph. (1) Vaccinates. Inject each of at least 10 but not more than 12 young adult hamsters, each weighing 50 to 90 grams, with 0.25 ml of the diluted bacterin either subcutaneously or intramuscularly, in accordance with the label recommendations for use. (2) Controls. Retain at least 10 but not more than 12 additional hamsters from the same group as unvaccinated con trols. (3) Challenge. From 14 to 18 days postvaccination, challenge each of 10 vaccinates and each of 10 controls intraperitoneally with a suspension of virulent Leptospira canicola organisms, using a dose of 10–10,000 hamster LD50 as determined by titration. (4) Post-challenge period. Observe the vaccinates and controls for 14 days post-challenge and record all deaths. If eight or more controls die from lepto spirosis, test is valid and the results shall be evaluated according to the fol lowing table: Stage Number of vac cinates Cumu lative number of vac cinates Cumulative total dead hamsters for satisfactory serial Cumulative total dead hamsters for unsatisfactory serial 1 .......... 10 ........ 10 ........ 2 or less ..... 5 or more. 2 .......... 10 ........ 20 ........ 5 or less ..... 6 or more. (5) If three or four vaccinates die in the first stage, the second stage shall be used. The second stage shall be con ducted in a manner identical to the first stage. (6) If the second stage is used, each serial shall be evaluated according to the second part of the table. On the basis of cumulative results, each serial shall either pass or fail. [39 FR 16862, May 10, 1974, as amended at 45 FR 40100, June 13, 1980. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66785, Dec. 26, 1991] § 113.104 Leptospira Grippotyphosa Bacterin. Leptospira Grippotyphosa Bacterin shall be produced from a culture of Leptospira grippotyphosa which has been inactivated and is nontoxic. Each se rial of biological product containing Leptospira grippotyphosa fraction shall meet the applicable requirements in § 113.100 and shall be tested for purity, safety, and potency as prescribed in this section. A serial found unsatisfac tory by any prescribed test shall not be released. (a) Purity test. Final container sam ples of completed product from each se rial and each subserial shall be tested for viable bacteria and fungi as pro vided in § 113.26. (b) Safety test. Bulk or final container samples of completed product from each serial shall be tested for safety as provided in § 113.38. (c) Potency test. Bulk or final con tainer samples of completed product shall be diluted with physiological sa line so that each 0.25 ml contains not more than 1/800th of the dose rec ommended on the label and shall be tested for potency, using the two-stage test provided in this paragraph. (1) Vaccinates. Inject each of at least 10 but not more than 12 young adult hamsters, each weighing 50 to 90 grams, with 0.25 ml of the diluted bacterin either subcutaneously or intramuscularly, in accordance with the label recommendations for use. (2) Controls. Retain at least 10 but not more than 12 additional hamsters from the same group as unvaccinated con trols. (3) Challenge. From 14 to 18 days postvaccination, challenge each of 10 vaccinates and each of 10 controls intraperitoneally with a suspension of virulent Leptospira grippotyphosa orga nisms, using a dose of 10–10,000 hamster LD50 as determined by titration. (4) Post-challenge period. Observe the vaccinates and controls for 14 days post-challenge and record all deaths. If eight or more controls die of lepto spirosis, the test is valid and the re sults shall be evaluated according to the following table: VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00747 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 738 9 CFR Ch. I (1–1–18 Edition) § 113.105 CUMULATIVE TOTALS Stage Number of vaccinates Dead hamsters for acceptance Dead hamsters for rejection 1 ........... 10 ................. 2 or less .......... 5 or more. 2 ........... 20 ................. 5 or less .......... 6 or more. (5) If three or four vaccinates die in the first stage, the second stage shall be conducted in a manner identical to the first stage. (6) If the second stage is used, each serial shall be evaluated according to the second part of the table. On the basis of cumulative results, each serial shall either pass or fail. [40 FR 17003, Apr. 16, 1975, as amended at 40 FR 23989, June 4, 1975; 45 FR 40100, June 13, 1980. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66785, Dec. 26, 1991] § 113.105 Leptospira Hardjo Bacterin. Leptospira Hardjo Bacterin shall be produced from a culture of Leptospira hardjo which has been inactivated and is nontoxic. Each serial of biological product containing Leptospira hardjo fraction shall meet the applicable re quirements in § 113.100 and shall be tested for purity, safety, and potency as prescribed in this section. A serial found unsatisfactory by any prescribed test shall not be released. (a) Purity test. Final container sam ples of completed product from each se rial and each subserial shall be tested for viable bacteria and fungi as pro vided in § 113.26. (b) Safety test. Bulk or final container samples of completed product from each serial shall be tested for safety as provided in § 113.38. (c) Potency test. Bulk or final con tainer samples of completed product from each serial shall be tested for po tency using the test written into the filed Outline of Production. [40 FR 17003, Apr. 16, 1975, as amended at 40 FR 20067, May 8, 1975. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66785, Dec. 26, 1991] § 113.106 Clostridium Chauvoei Bacterin. Clostridium Chauvoei Bacterin shall be produced from a culture of Clos tridium chauvoei which has been inac tivated and is nontoxic. Each serial of biological product containing Clos tridium chauvoei fraction shall meet the applicable requirements in § 113.100 and shall be tested for purity, safety, and potency as prescribed in this section. Serials found unsatisfactory by any prescribed test shall not be released. (a) Purity test. Final container sam ples of completed product from each se rial and each subserial shall be tested for viable bacteria and fungi as pro vided in § 113.26. (b) Safety test. Bulk or final container samples of completed product from each serial shall be tested for safety as provided in § 113.38. (c) Potency test. Bulk or final con tainer samples of completed product from each serial shall be tested for po tency using the two-stage test provided in this paragraph. (1) Each of at least 8 but not more than 10 guinea pigs, each weighing 300 to 500 grams, shall be injected subcutaneously with a guinea pig dose. A second guinea pig dose shall be in jected 21 to 23 days after the first dose. Each guinea pig dose shall be one-fifth of the dose recommended on the label for a calf. (2) Clostridium chauvoei challenge ma terial, available upon request from Animal and Plant Health Inspection Service, shall be used for challenge 14 to 15 days following the last injection of the product. Each of eight vac cinates and each of five additional non vaccinated guinea pigs for controls shall be injected intramuscularly with approximately 100 LD50 of challenge material. This dose shall be determined by statistical analysis of results of ti trations of the challenge material. The vaccinates and controls shall be ob served for 3 days postchallenge and all deaths recorded. (3) For a valid test, at least 80 per cent of the controls shall die within the 3 day post-challenge observation period. If this requirement is met, the results of the potency test shall be evaluated according to the following table: Stage Number of vac cinates Cumu lative number of vac cinates Cumulative total number of deaths for a satisfac tory test Cumulative total number of deaths for an unsatisfac tory test 1 .......... 8 .......... 8 .......... 1 or less ..... 3 or more. 2 .......... 8 .......... 16 ........ 4 or less ..... 5 or more. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00748 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 739 Animal and Plant Health Inspection Service, USDA § 113.108 The second stage shall be required only when exactly two animals die in the first stage. The second stage shall be conducted in a manner identical to the first stage. [39 FR 16862, May 10, 1974, as amended at 45 FR 40100, June 13, 1980. Redesignated at 55 FR 35562, Aug. 31, 1990 and amended at 56 FR 66784, 66785, Dec. 26, 1991] § 113.107 Clostridium Haemolyticum Bacterin. Clostridium Haemolyticum Bacterin shall be produced from a culture of Clostridium haemolyticum which has been inactivated and is nontoxic. Each serial of biological product containing Clostridium haemolyticum fraction shall meet the applicable requirements in § 113.100 and shall be tested for purity, safety, and potency as prescribed in this section. A serial found unsatisfac tory by any prescribed test shall not be released. (a) Purity test. Final container sam ples of completed product from each se rial and each subserial shall be tested for viable bacteria and fungi as pro vided in § 113.26. (b) Safety test. Bulk or final container samples of completed product from each serial shall be tested for safety as provided in § 113.38. (c) Potency test. Bulk or final con tainer samples of completed product from each serial shall be tested for po tency using the two-stage test provided in this paragraph. (1) Each of at least 8 but not more than 10 guinea pigs, each weighing 300 to 500 grams, shall be injected subcutaneously with a guinea pig dose. A second guinea pig dose shall be in jected 21 to 23 days after the first dose. Each guinea pig dose shall be one-fifth of the dose recommended on the label for a calf. (2) Clostridium haemolyticum challenge material, available upon request from Animal and Plant Health Inspection Service, shall be used for challenge 14 to 15 days following the last injection of the product. Each of eight vac cinates and each of five additional non vaccinated guinea pigs for controls shall be injected intramuscularly with approximately 100 LD50 of challenge material. This dose shall be determined by statistical analysis of results of ti trations of the challenge material. The vaccinates and controls shall be ob served for 3 days postchallenge and all deaths recorded. (3) For a valid test, at least 80 per cent of the controls shall die within the 3 day post-challenge observation period. If this requirement is met, the results of the potency test shall be evaluated according to the following table: Stage Number of vac cinates Cumu lative number of vac cinates Cumulative total number of deaths for a satisfac tory test Cumulative total number of deaths for an unsatisfac tory test 1 .......... 8 .......... 8 .......... 1 or less ..... 3 or more. 2 .......... 8 .......... 16 ........ 4 or less ..... 5 or more. The second stage shall be required only when exactly two animals die in the first stage. The second stage shall be conducted in a manner identical to the first stage. [39 FR 16862, May 10, 1974, as amended at 40 FR 20067, May 8, 1975; 45 FR 40100, June 13, 1980. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66784, 66785, Dec. 26, 1991] § 113.108 Clostridium Novyi Bacterin Toxoid. Clostridium Novyi Bacterin-Toxoid shall be produced from a culture of Clostridium novyi which has been inac tivated and is nontoxic. Each serial of biological product containing Clos tridium novyi fraction shall meet the applicable requirements in § 113.100 and shall be tested for purity, safety, and potency as prescribed in this section. A serial found unsatisfactory by any pre scribed test shall not be released. (a) Purity test. Final container sam ples of completed product from each se rial and each subserial shall be tested for viable bacteria and fungi as pro vided in § 113.26. (b) Safety test. Bulk or final container samples of completed product from each serial shall be tested for safety as provided in § 113.38. (c) Potency test. Bulk or final con tainer samples of completed product from each serial shall be tested for po tency using the Alpha toxin-neutraliza tion test provided in this paragraph. (1) When used in this test, the fol lowing words and terms shall mean: VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00749 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 740 9 CFR Ch. I (1–1–18 Edition) § 113.108 (i) International antitoxin unit. (I.U.) That quantity of Alpha Antitoxin which reacts with Lo and L + doses of Standard Toxin according to their defi nitions. (ii) Lo dose. The largest quantity of toxin which can be mixed with one unit of Standard Antitoxin and not cause sickness or death in injected mice. (iii) L + dose. The smallest quantity of toxin which can be mixed with one unit of Standard Antitoxin and cause death in at least 80 percent of injected mice. (iv) Standard antitoxin. The Alpha Antitoxin preparation which has been standardized as to antitoxin unitage on the basis of the International Clos tridium novyi Alpha Antitoxin Standard and which is either supplied by or ac ceptable to the Animal and Plant Health Inspection Service. The anti toxin unit value shall be stated on the label. (v) Standard toxin. The Alpha toxin preparation which is supplied by or is acceptable to the Animal and Plant Health Inspection Service. (vi) Diluent. The solution used to make proper dilutions prescribed in this test. Such solutions shall be made by dissolving 1 gram of peptone and 0.25 gram of sodium chloride in each 100 ml of distilled water; adjusting the pH to 7.2; autoclaving at 121 °C for 25 min utes; and storing at 4 °C until used. (2) Each of at least eight rabbits of a strain acceptable to the Animal and Plant Health Inspection Service, each weighing 4–8 pounds, shall be injected subcutaneously with not more than half of the recommended cattle dose. Provided, That, if the product is rec ommended only for sheep, half of the recommended sheep dose shall be used. A second dose shall be given not less than 20 days nor more than 23 days after the first dose. (3) Fourteen to seventeen days after the second dose, all surviving rabbits shall be bled, and the serum tested for antitoxin content. (i) At least seven rabbits are required to make an acceptable serum pool. (ii) Equal quantities of serum from each rabbit shall be combined and test ed as a single pooled serum. (iii) If less than seven rabbits are available, the test is invalid and shall be repeated: Provided, That, if the test is not repeated, the serial shall be de clared unsatisfactory. (4) The antitoxin content of the rab bit serums shall be determined by the serum neutralization test as follows: (i) Make a dilution of Standard Anti toxin to contain 0.1 International Unit of antitoxin per ml. (ii) Make a dilution of Standard Toxin in which 0.1 Lo dose is contained in a volume of 1 ml or less. Make a sec ond dilution of Standard Toxin in which 0.1 L + dose is contained in a vol ume of 1 ml or less. (iii) Combine 0.1 International Unit of Standard Antitoxin with 0.1 Lo dose of diluted Standard Toxin and combine 0.1 International Unit of Standard Antitoxin with 0.1 L + dose of diluted Standard Toxin. Each mixture is ad justed to a final volume of 2.0 ml with diluent. (iv) Combine 0.1 Lo dose of diluted Standard Toxin with a 0.2 ml volume of undiluted serum. The mixture is ad justed to a final volume of 2.0 ml with diluent. (v) Neutralize all toxin-antitoxin mixtures at room temperature for 1 hour and hold in ice water until injec tions of mice can be made. (vi) Five Swiss white mice, each weighing 16–20 grams, shall be used for each toxin-antitoxin mixture. A dose of 0.2 ml shall be injected intravenously into each mouse. Conclude the test 72 hours post injection and record all deaths. (5) Test Interpretation shall be as fol lows: (i) If any mice inoculated with the mixture of 0.1 International Unit of Standard Antitoxin and 0.1 Lo doses of Standard Toxin die, the results of the serum neutralization test are a No Test and shall be repeated: Provided, That, if the test is not repeated, the serial shall be declared unsatisfactory. (ii) If less than 80 percent of the mice inoculated with the mixture of 0.1 International Unit of Standard Anti toxin and 0.1 L + doses of Standard Toxin die, the results of the serum neu tralization test are a No Test and shall be repeated: Provided, That, if the test is not repeated, the serial shall be de clared unsatisfactory. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00750 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 741 Animal and Plant Health Inspection Service, USDA § 113.109 (iii) If any mice inoculated with the mixture of 0.2 ml undiluted serum with 0.1 Lo dose of Standard Toxin die, the serum is considered to contain less than 0.50 International Units per ml. (iv) If the single pooled serum from seven or more rabbits contains less than 0.5 International Unit per ml, the serial is unsatisfactory. [39 FR 16862, May 10, 1974, as amended at 45 FR 40101, June 13, 1980. Redesignated at 55 FR 35562, Aug. 31, 1990; 56 FR 37825, Aug. 9, 1991, as amended at 56 FR 66784, 66785, Dec. 26, 1991] § 113.109 Clostridium Sordellii Bacterin-Toxoid. Clostridium Sordellii Bacterin-Tox oid shall be produced from a culture of Clostridium sordellii which has been in activated and is nontoxic. Each serial of biological product containing Clos tridium sordellii fraction shall meet the applicable requirements in § 113.100 and shall be tested for purity, safety, and potency as prescribed in this section. A serial found unsatisfactory by any pre scribed test shall not be released. (a) Purity test. Final container sam ples of completed product from each se rial and each subserial shall be tested for viable bacteria and fungi as pro vided in § 113.26. (b) Safety test. Bulk or final container samples of completed product from each serial shall be tested for safety as provided in § 113.38. (c) Potency test. Bulk or final con tainer samples of completed product from each serial shall be tested for po tency using the toxin-neutralization test provided in this paragraph. (1) When used in this test, the fol lowing words and terms shall mean: (i) International antitoxin unit. (I.U.) That quantity of antitoxin which re acts with Lo and L + doses of Standard Toxin according to their definitions. (ii) Lo dose. The largest quantity of toxin which can be mixed with one unit of Standard Antitoxin and not cause sickness or death in injected mice. (iii) L + dose. The smallest quantity of toxin which can be mixed with one unit of Standard Antitoxin and cause death in at least 80 percent of injected mice. (iv) Standard antitoxin. The antitoxin preparation which has been standard ized as to antitoxin unitage on the basis of the International Clostridium sordellii Antitoxin Standard and which is either supplied by or acceptable to the Animal and Plant Health Inspec tion Service. The antitoxin unit value shall be stated on the label. (v) Standard toxin. The toxin prepara tion which is supplied by or is accept able to the Animal and Plant Health Inspection Service. (vi) Diluent. The solution used to make proper dilutions prescribed in this test. Such solutions shall be made by dissolving 1 gram of peptone and 0.25 gram of sodium chloride in each 100 ml of distilled water; adjusting the pH to 7.2; autoclaving at 121 °C for 25 min utes; and storing at 4 °C until used. (2) Each of at least eight rabbits of a strain acceptable to the Animal and Plant Health Inspection Service, each weighing 4–8 pounds, shall be injected subcutaneously with not more than half of the recommended cattle dose: Provided, That, if the product is rec ommended only for sheep, half of the recommended sheep dose shall be used. A second dose shall be given not less than 20 days nor more than 23 days after the first dose. (3) Fourteen to seventeen days after the second dose, all surviving rabbits shall be bled, and the serum tested for antitoxin content. (i) At least seven rabbits are required to make an acceptable serum pool. (ii) Equal quantities of serum from each rabbit shall be combined and test ed as a single pooled serum. (iii) If less than seven rabbits are available, the test is a No Test and shall be repeated: Provided, That, if the test is not repeated, the serial shall be declared unsatisfactory. (4) The antitoxin content of the rab bit serums shall be determined by the serum neutralization test as follows: (i) Make a dilution of Standard Anti toxin to contain 1.0 international unit of antitoxin per ml. (ii) Make a dilution of Standard Toxin in which 1.0 Lo dose is contained in a volume of 1 ml or less. Make a sec ond dilution of Standard Toxin in which 1.0 L + dose is contained in a vol ume of 1 ml or less. (iii) Combine 1.0 International Unit Standard Antitoxin with 1.0 Lo dose of VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00751 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 742 9 CFR Ch. I (1–1–18 Edition) § 113.110 diluted Standard Toxin and combine 1.0 International Unit of Standard Anti toxin with 1.0 L + dose of diluted Standard Toxin. Each mixture is ad justed to a final volume of 2.0 ml with diluent. (iv) Combine 1.0 Lo dose of diluted Standard Toxin with a 1.0 ml volume of undiluted serum. This mixture is ad justed to a final volume of 2.0 ml with diluent. (v) Neutralize all toxin-antitoxin mixtures at room temperature for 1 hour and hold in ice water until injec tions of mice can be made. (vi) Five Swiss white mice, each weighing 16–20 grams, shall be used for each toxin-antitoxin mixture. A dose of 0.2 ml shall be injected intravenously into each mouse. Conclude the test 72 hours post injection and record all deaths. (5) Test Interpretation shall be as fol lows: (i) If any mice inoculated with the mixture of 1.0 International Unit of Standard Antitoxin and 1.0 Lo doses of Standard Toxin die, the results of the serum neutralization test are a No Test and shall be repeated: Provided, That, if the test is not repeated, the serial shall be declared unsatisfactory. (ii) If less than 80 percent of the mice inoculated with the mixture of 1.0 International Unit of Standard Anti toxin and 1.0 L + doses of Standard Toxin die, the results of the serum neu tralization test are a No Test and shall be repeated: Provided, That, if the test is not repeated, the serial shall be de clared unsatisfactory. (iii) If any mice inoculated with the mixture of 1.0 ml undiluted serum with 1.0 Lo dose of Standard Toxin die, the serum is considered to contain less than 1.0 International Units per ml. (iv) If the single pooled serum from seven or more rabbits contains less than 1.0 International Unit per ml, the serial is unsatisfactory. [39 FR 16862, May 10, 1974, as amended at 42 FR 61247, Dec. 2, 1977; 45 FR 40101, June 13, 1980. Redesignated at 55 FR 35562, Aug. 31, 1990; 56 FR 37826, Aug. 9, 1991; 56 FR 66784, 66785, Dec. 26, 1991; 79 FR 55969, Sept. 18, 2014] § 113.110 Clostridium Botulinum Type C Bacterin-Toxoid. Clostridium Botulinum Type C Bacterin-Toxoid shall be produced from a culture of Clostridium botulinum Type C which has been inactivated and is nontoxic. Each serial of biological product containing Clostridium botu linum Type C fraction shall meet the applicable requirements in § 113.100 and shall be tested for purity, safety, and potency as prescribed in this section. A serial found unsatisfactory by any pre scribed test shall not be released. (a) Purity test. Final container sam ples of completed product from each se rial and each subserial shall be tested for viable bacteria and fungi as pro vided in § 113.26. (b) Safety test. Bulk or final container samples of completed product from each serial shall be tested for safety as provided in § 113.33(b). (c) Potency test. Bulk or final con tainer samples of completed product from each serial shall be tested for po tency, using susceptible mink as test animals. At least five vaccinates and three unvaccinated controls of the same source and approximately the same age shall be used. (1) Each of the vaccinates shall be in jected subcutaneously with the dose recommended on the label for mink. Twenty-one to twenty-eight days post injection, the vaccinates and the con trols shall be challenged intraperitoneally with botulinum Type C toxin which has been titrated in mice to provide for a 10 4.0 mouse MLD dose. The titration technique shall include inoculation of the mice intraperitoneally. (2) The vaccinates and controls shall be observed for 7 days post-challenge and signs of botulism and deaths noted. For a valid test, the controls shall die of botulism. If the test is valid and 80 percent of the vaccinates do not re main free of botulism, the serial is un satisfactory. [39 FR 16862, May 10, 1974, as amended at 40 FR 759, Jan. 3, 1975. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66785, Dec. 26, 1991] VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00752 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 743 Animal and Plant Health Inspection Service, USDA § 113.111 § 113.111 Clostridium Perfringens Type C Toxoid and Bacterin-Toxoid. Clostridium Perfringens Type C Tox oid and Clostridium Perfringens Type C Bacterin-Toxoid shall be produced from a culture of Clostridium perfringens Type C which has been inactivated and is nontoxic. Each serial shall meet the applicable requirements in § 113.100 and shall be tested for purity, safety, and potency as prescribed in this section. Any serial found unsatisfactory by a prescribed test shall not be released. (a) Purity test. Final container sam ples of completed product from each se rial and each subserial shall be tested for viable bacteria and fungi as pro vided in § 113.26. (b) Safety test. Bulk or final container samples of completed product from each serial shall be tested for safety as provided in § 113.33(b). (c) Potency test. Bulk or final con tainer samples of completed product from each serial shall be tested for po tency using the Beta toxin-neutraliza tion test provided in this paragraph. (1) When used in this test, the fol lowing words and terms shall mean: (i) International antitoxin unit. (I.U.) That quantity of Beta Antitoxin which reacts with L 0 and L + doses of Standard Toxin according to their defi nitions. (ii) L 0 dose. The largest quantity of toxin which can be mixed with one unit of Standard Antitoxin and not cause sickness or death in injected mice. (iii) L + dose. The smallest quan tity of toxin which can be mixed with one unit of Standard Antitoxin and cause death in at least 80 percent of in jected mice. (iv) Standard antitoxin. The Beta Antitoxin preparation which has been standardized as to antitoxin unitage on the basis of the International Clos tridium perfringens Beta Antitoxin Standard and which is either supplied by or acceptable to Animal and Plant Health Inspection Service. The anti toxin unit value shall be stated on the label. (v) Standard toxin. The Beta toxin preparation which is supplied by or is acceptable to Animal and Plant Health Inspection Service. (vi) Diluent. The solution used to make proper dilutions prescribed in this test. Such solutions shall be made by dissolving 1 gram of peptone and 0.25 grams of sodium chloride in each 100 ml of distilled water; adjusting the pH to 7.2; autoclaving at 250 °F for 25 minutes; and storing at 4 °C until used. (2) Each of at least eight rabbits of a strain acceptable to APHIS, each weighing 4–8 pounds, shall be injected subcutaneously with not more than half of the largest recommended dose for any species indicated on the prod uct label. A second equivalent dose shall be given not less than 20 days nor more than 23 days after the first does. (3) Fourteen to seventeen days after the second dose, all surviving rabbits shall be bled and the serum tested for antitoxin content. (i) At least seven rabbits are required to make an acceptable serum pool. (ii) Equal quantities of serum from each rabbit shall be combined and test ed as a single pooled serum. (iii) If less than seven rabbits are available, the test is a No Test and shall be repeated: Provided, That, if the test is not repeated, the serial shall be declared unsatisfactory. (4) The antitoxin content of the rab bit serums shall be determined as fol lows: (i) Make a dilution of Standard Anti toxin to contain 10 International Units of antitoxin per ml. (ii) Make one dilution of Standard Toxin to contain 10 L 0 doses per ml and make a second dilution of Standard Toxin to contain 10 L ∂ doses per ml. (iii) Combine 10 International Units of Standard Antitoxin with 10 L 0 doses of diluted Standard Toxin and combine 10 International Units of Standard Antitoxin with 10 L ∂ doses of diluted Standard Toxin. (iv) Combine 1 ml of undiluted serum with 10 L0 doses of diluted Standard Toxin. (v) Neutralize all toxin-antitoxin mixtures at room temperature for 1 hour and hold in ice water until injec tions of mice can be made. (vi) Five Swiss white mice, each weighing 16–20 grams, shall be used for each toxin-antitoxin mixture. A dose of 0.2 ml shall be injected intravenously into each mouse. Conclude the test 24 hours post-injection and record all deaths. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00753 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 744 9 CFR Ch. I (1–1–18 Edition) § 113.112 (5) Test Interpretation shall be as fol lows: (i) If any mice inoculated with the mixture of 10 International Units of Standard Antitoxin and 10 L0 doses of Standard Toxin die, the results of the test are a No Test and shall be re peated: Provided, That, if the test is not repeated, the serial shall be declared unsatisfactory. (ii) If less than 80 percent of the mice inoculated with mixture of 10 Inter national Units of Standard Antitoxin and 10 L ∂ doses of Standard Toxin die, the results of the test are a No Test and shall be repeated: Provided, That, if the test is not repeated, the serial shall be declared unsatisfactory. (iii) If any mice inoculated with the mixture of serum with 10 L0 doses of Standard Toxin die, the serum is con sidered to contain less than 10 Inter national Units per ml. and the serial is unsatisfactory [39 FR 16862, May 10, 1974, as amended at 40 FR 759, Jan. 3, 1975; 40 FR 41088, Sept. 5, 1975. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66784, 66785, Dec. 26, 1991; 62 FR 31330, June 9, 1997; 79 FR 55969, Sept. 18, 2014] § 113.112 Clostridium Perfringens Type D Toxoid and Bacterin-Toxoid. Clostridium Perfringens Type D Tox oid and Clostridium Perfringens Type D Bacterin-Toxoid shall be produced from a culture of Clostridium perfringens Type D which has been inactivated and is nontoxic. Each serial shall meet the applicable requirements in § 113.100 and shall be tested for purity, safety, and potency as prescribed in this section. Any serial found unsatisfactory by a prescribed test shall not be released. (a) Purity test. Final container sam ples of completed product from each se rial and each subserial shall be tested for viable bacteria and fungi as pro vided in § 113.26. (b) Safety test. Bulk or final container samples of completed product from each serial shall be tested for safety as provided in § 113.33(b). (c) Potency test. Bulk or final con tainer samples of completed product from each serial shall be tested for po tency using the Epsilon toxin-neutral ization test provided in this paragraph. (1) When used in this test, the fol lowing words and terms shall mean: (i) International antitoxin unit. (I.U.) That quantity of Epsilon Antitoxin which reacts with L0 and L ∂ doses of Standard Toxin according to their defi nitions. (ii) L0 dose. The largest quantity of toxin which can be mixed with one tenth unit of Standard Antitoxin and not cause sickness or death in injected mice. (iii) L ∂ dose. The smallest quantity of toxin which can be mixed with one tenth unit of Standard Antitoxin and cause death in at least 80 percent of in jected mice. (iv) Standard antitoxin. The Epsilon Antitoxin preparation which has been standardized as to antitoxin unitage on the basis of the International Clos tridium perfringens Epsilon Antitoxin Standard and which is either supplied by or acceptable to Animal and Plant Health Inspection Service. The anti toxin unit value shall be stated on the label. (v) Standard toxin. The Epsilon toxin preparation which is supplied by or is acceptable to Animal and Plant Health Inspection Service. (vi) Diluent. The solution used to make proper dilutions prescribed in this test. Such solutions shall be made by dissolving 1 gram of peptone and 0.25 gram of sodium chloride in each 100 ml of distilled water; adjusting the pH to 7.2; autoclaving at 250 °F for 25 min utes; and storing at 4 °C until used. (2) Each of at least eight rabbits of a strain acceptable to APHIS, each weighing 4–8 pounds, shall be injected subcutaneously with not more than half of the largest recommended dose for any species indicated on the prod uct label. A second equivalent dose shall be given not less than 20 days nor more than 23 days after the first dose. (3) Fourteen to seventeen days after the second dose, all surviving rabbits shall be bled, and the serum tested for antitoxin content. (i) At least seven rabbits are required to make an acceptable serum pool. (ii) Equal quantities of serum from each rabbit shall be combined and test ed as a single pooled serum. (iii) If less than seven rabbits are available, the test is a No Test and VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00754 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 745 Animal and Plant Health Inspection Service, USDA § 113.113 shall be repeated: Provided, That, if the test is not repeated, the serial shall be declared unsatisfactory. (4) The antitoxin content of the rab bit serums shall be determined as fol lows: (i) Make a dilution of Standard Anti toxin to contain 1 International Unit of antitoxin per ml. (ii) Make one dilution of Standard Toxin to contain 10 Lo doses per ml and make a second dilution of Standard Toxin to contain 10 L ∂ doses per ml. (iii) Combine 1 International Unit of Standard Antitoxin with 10 Lo doses of diluted Standard Toxin and Combine 1 International Unit of Standard Anti toxin with 10 L ∂ doses of diluted Standard Toxin. (iv) Dilute 1 ml of serum with 1 ml of diluent (1:2) and combine 1 ml of this solution with 10 Lo doses of diluted Standard Toxin. (v) Neutralize all toxin-antitoxin mixtures at room temperature for 1 hour and hold in ice water until injec tions of mice can be made. (vi) Five Swiss white mice, each weighing 16–20 grams, shall be used for each toxin-antitoxin mixture. A dose of 0.2 ml shall be injected intravenously into each mouse. Conclude the test 24 hours post-injection and record all deaths. (5) Test Interpretation shall be as fol lows: (i) If any mice inoculated with the mixture of 1 International Unit of Standard Antitoxin and 10 Lo doses of Standard Toxin die, the results of the test are a No Test and shall be re peated: Provided, That, if the test is not repeated, the serial shall be declared unsatisfactory. (ii) If less than 80 percent of the mice inoculated with mixture of 1 Inter national Unit of Standard Antitoxin and 10 L ∂ doses of Standard Toxin die, the results of the test area No Test and shall be repeated: Provided, That, if the test is not repeated, the serial shall be declared unsatisfactory. (iii) If any mice inoculated with the mixture of serum with 10 Lo doses of Standard Toxin die, the serum is con sidered to contain less than 2 Inter national Units per ml, and the serial is unsatisfactory. [39 FR 16865, May 10, 1974; 39 FR 20783, June 14, 1974. Redesignated at 39 FR 25463, July 11, 1974, and amended at 40 FR 759, Jan. 3, 1975; 40 FR 41088, Sept. 5, 1975. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66784, 66785, Dec. 26, 1991; 62 FR 31331, June 9, 1997; 79 FR 55969, Sept. 18, 2014] § 113.113 Autogenous biologics. Autogenous biologics shall be pre pared from cultures of microorganisms which have been inactivated and are nontoxic. Such products shall be pre pared only for use by or under the di rection of a veterinarian under a vet erinarian-client-patient relationship, Provided, That, such products may be prepared for use under the direction of a person of appropriate expertise in specialized situations such as aqua culture, if approved by the Adminis trator. Each serial of an autogenous biologic shall meet the requirements in this section, and if found unsatisfactory by any prescribed test shall not be used. (a) Seed requirements. The microorga nisms used as seed to prepare autog enous biologics shall be microorga nisms which are isolated from sick or dead animals in the herd of origin and which there is reason to believe are the causative agent(s) of the current dis ease affecting such animals. (1) More than one microorganism iso lated from the same herd may be used as seed. (2) Under normal circumstances, microorganisms from one herd must not be used to prepare an autogenous biologic for another herd. The Adminis trator, however, may authorize prepa ration of an autogenous biologic for use in herds adjacent to the herd of ori gin, when adjacent herds are consid ered to be at risk. To request author ization to prepare a product for use in herds adjacent to the herd of origin, the establishment seeking authoriza tion must submit to the Administrator (in c/o the Director, Center for Veteri nary Biologics, Inspection and Compli ance, 1920 Dayton Avenue, P.O. Box 844, Ames, IA 50010) the following informa tion. (If any of the data are unavail able, the applicant for authorization VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00755 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 746 9 CFR Ch. I (1–1–18 Edition) § 113.113 should indicate that such data are un available and why.) (i) Name, address, and phone number of the owner of the herd of origin. (ii) Attending veterinarian’s name, address, and phone number. (iii) Animal species and number in herd of origin. (iv) Identification of microorga nism(s), at least to genus. (v) Diagnosis or clinical signs of the disease observed. (vi) Name and address of the person who isolated the microorganism(s) and the date of isolation. (vii) Number of doses of autogenous biologic requested and vaccination schedule. (viii) Each adjacent herd owner’s name, address, and phone number. (ix) Number of animals and species in each adjacent herd. (x) The attending veterinarian’s or approved specialist’s assessment of the involvement of the adjacent herd(s) with the disease observed. The applicant shall give notice to the State Veterinarian or other appro priate State Official in writing when an autogenous biologic is to be used in ad jacent herds. (3) The Administrator may authorize preparation of an autogenous biologic for use in herds which are not adjacent to the herd of origin, but which he or she considers to be at risk of infection with the same microorganism(s). Ex cept as provided below, the same infor mation which is required for prepara tion of such product for use in herds adjacent to the herd of origin must be submitted to the Administrator (in c/o the Director, Center for Veterinary Biologics, Inspection and Compliance, 1920 Dayton Avenue, P.O. Box 844, Ames, IA 50010) for authorization to prepare a product for use in herds not adjacent to the herd of origin. Because the recipient herd involved may not be known when autogenous biologics are to be used in other geographic areas, the following data may be used in place of the data required in paragraphs (a)(2)(viii) and (a)(2)(ix) of this section. (i) Names and addresses of practi tioners in the area in place of the name, address, and phone number of the adjacent herd owner. (ii) The geographic designations of the area involved. (iii) A summary of the epidemiology of the disease situation that links the designated geographic areas with the herd of origin. In addition, an applicant for authoriza tion under this paragraph (a)(3) shall provide written approval from the State Veterinarian or other appro priate State Official in the State in which the autogenous biologic is to be used in nonadjacent herds. (4) Under normal circumstances, microorganism(s) used for the produc tion of autogenous biologics may not be older than 15 months from the date of isolation, or 12 months from the date of harvest of the first serial of product produced from the microorganism(s), whichever comes first. The Adminis trator, however, may authorize produc tion of additional serials from micro organism(s) older than the above stat ed time periods, Provided, That, the person requesting such authorization submits the following supporting infor mation to the address listed in para graph (a)(3): (i) The attending veterinarian’s or approved specialist’s current assess ment of the continued involvement of a herd with the originally isolated microorganism(s), including a sum mary of the diagnostic work that has been done to support this assessment. (ii) Evidence of satisfactory protec tion from the previous use of the au togenous biologic produced from the microorganisms involved. (iii) Any other information the Ad ministrator may require in order to de termine the need to use the microorga nism to make additional serials. (b) Restrictions. Unless otherwise au thorized by the Administrator, each se rial of an autogenous biologic shall be subject to the following restrictions: (1) Microorganisms used to prepare autogenous biologics shall not be main tained in the licensed establishment beyond the time authorized for use in production. (2) The expiration date of the autog enous biologic shall not exceed 18 months from the date of harvest. (c) Testing requirements for autogenous biologics. (1) Final container samples of completed product from the first serial VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00756 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 747 Animal and Plant Health Inspection Service, USDA § 113.113 or subserial of an autogenous biologic produced from an isolate shall be test ed for purity as prescribed in § 113.26, and for safety as prescribed in § 113.33(b) or § 113.38 except that: (i) When the number of final con tainers in a serial or subserial is 50 or less, two final container samples from each serial and subserial shall be tested as prescribed in § 113.26(b): Provided, That, 1 ml aliquots from each sample may be inoculated into five cor responding individual test vessels of each of the test media required. (ii) Serials which are satisfactory after the third day of observation of purity test cultures and of safety test animals may be released for shipment to the customer and the tests contin ued throughout the required period; and (iii) Serials released on the basis of satisfactory results of third day obser vations shall be immediately recalled if evidence of contamination occurs in test cultures or if any of the test ani mals used to demonstrate product safe ty, sicken, or die during the observa tion period. (iv) Test summaries must be sub mitted to the Administrator (in c/o the Director, Center for Veterinary Bio logics, Inspection and Compliance, 1920 Dayton Avenue, P.O. Box 844, Ames, IA 50010) on a quarterly basis by the 21st day of January, April, July, and Octo ber or more often as required by the Administrator. (2) Each serial or subserial of autog enous bacterial product other than the first serial or subserial produced from an isolate shall meet the applicable general requirements prescribed in § 113.100 and the special requirements prescribed in this section. Each serial or subserial of autogenous viral prod uct other than the first serial or sub serial produced from an isolate shall meet the applicable general require ments prescribed in § 113.200 and the special requirements prescribed in this section. A serial or subserial found un satisfactory by any prescribed test shall not be released. (i) Purity test. Final container sam ples of completed product from each se rial and subserial shall be tested for viable bacteria and fungi as provided in § 113.26. When the number of final con tainers in a serial or subserial is 50 or less, two final container samples from each serial and subserial shall be tested as prescribed in § 113.26(b): Provided, That, 1 ml aliquots from each sample may be inoculated into five cor responding individual test vessels of each of the test media required. (ii) Safety test. Bulk of final container samples of completed product from each serial shall be tested for safety as provided in § 113.33 (b) or § 113.38. (iii) Identification. All microorga nisms used for the production of autog enous biologics shall be identified as follows: Bacteria, fungi, and myco plasma shall be identified at least to genus and species. Viruses shall be identified at least to family. After 15 months from the date of isolation, or 12 months from the harvest date of the first serial of autogenous product pro duced from a microorganism, which ever comes first, characterization and identification shall be completed to strain and/or serotype before such microorganism may be used for produc tion. (iv) Antigenicity, or immunogenicity, and potency. Persons seeking author ization to prepare additional serials of autogenous biologics from microorga nisms that are older than 24 months from the date of isolation, shall be re quired to conduct the following addi tional tests: (A) Completed product shall be tested for antigenicity or immunogenicity in the species for which the product is recommended or in another animal species whose immunological response has been shown in the scientific lit erature to correlate with the response of the species for which the product is recommended. Such tests shall be con ducted in accordance with a protocol developed by the licensee and approved by the Administrator and the results submitted to the Director, Center for Veterinary Biologics, Policy, Evalua tion, and Licensing, 1920 Dayton Ave nue, P.O. Box 844, Ames, IA 50010 for re view. Microorganisms not shown to be antigenic (that is, not shown to induce a significant serological response) or immunogenic by such approved tests shall not be used for the preparation of such product. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00757 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 748 9 CFR Ch. I (1–1–18 Edition) § 113.114 (B) Bulk or final container samples of completed product from each serial of such autogenous biologics con taining fractions for which standard re quirement potency test procedures have been established shall be tested for potency in accordance with applica ble standard requirement potency tests provided in 9 CFR part 113. If the cul ture of microorganisms used to produce such fractions is shown to be of a dif ferent strain or serotype than the rea gent or challenge microorganisms used in the standard requirement potency test, reagents or challenges of the same strain or serotype as the microorga nism used for production may be used. (C) If no standard requirement po tency test procedures have been estab lished for a fraction(s) in the autog enous biologic, such fraction(s) of each serial of product shall be tested for po tency using a developmental potency test described in the filed outline of production or shall at least be stand ardized to contain an antigenic mass for such fraction(s) that has been shown to be antigenic or immunogenic in accordance with paragraph (c)(2)(iv)(A) of this section. [57 FR 38756, Aug. 27, 1992, as amended at 59 FR 67616, Dec. 30, 1994; 64 FR 43044, Aug. 9, 1999; 67 FR 15714, Apr. 3, 2002; 75 FR 20773, Apr. 21, 2010] § 113.114 Tetanus Toxoid. Tetanus Toxoid shall be produced from a culture of Clostridium tetani which has been inactivated and is nontoxic. The toxoid may be either ab sorbed, precipitated, or purified and concentrated. Each serial of biological product containing tetanus toxoid frac tion shall meet the applicable require ments in § 113.100 and shall be tested for purity, safety, and potency as pre scribed in this section. A serial or sub serial found unsatisfactory by any pre scribed test shall not be released. (a) Purity test. Final container sam ples of completed product from each se rial and subserial shall be tested for viable bacteria and fungi as provided in § 113.26. (b) Safety test. Bulk or final container samples of completed product from each serial shall be tested for safety as provided in § 113.33(b). (c) Potency test. Bulk or final con tainer samples of completed product from each serial shall be tested for po tency. A group of 10 guinea pigs con sisting of an equal number of males and females weighing 450 to 550 grams shall each be injected subcutaneously with 0.4 of the largest dose rec ommended on the product labels. (1) Six weeks after injection, all sur viving guinea pigs shall be bled and equal portions of serum, but not less than 0.5 ml from each, shall be pooled. For a valid test, the pool shall contain the serum from at least eight animals. (2) The antitoxin titer of the pooled serum shall be determined in antitoxin units (A.U.) per ml using an enzyme linked immunosorbent assay method acceptable to the Animal and Plant Health Inspection Service. (3) If the antitoxin titer of the serum pool is at least 2.0 A.U. per ml, the se rial is satisfactory. If the antitoxin titer of the serum pool is less than 2.0 A.U. per ml, the serial may be retested by the following procedure: Provided, That, if the serial is not retested, it shall be declared unsatisfactory. (4) For serials in which the serum pool contains less than 2.0 A.U. per ml, the individual serum that constituted the pool may be tested by the enzyme linked immunosorbent assay. If at least 80 percent of the individual se rums have an antitoxin titer of at least 2.0 A.U. per ml, the serial is satisfac tory. If less than 80 percent of the indi vidual serums have an antitoxin titer of at least 2.0 A.U. per ml, the serial may be retested in 10 guinea pigs using the procedure described in (c)(1) and (2) above. The antitoxin titer of the pooled serum from the guinea pigs used in the retest shall be averaged with the anti toxin level of the pooled serum from the initial test. If the average of the two pools is at least 2.0 A.U. per ml, the serial is satisfactory. If the average of the two pools is less than 2.0 A.U. per ml, the serial is unsatisfactory and shall not be retested further. [39 FR 16862, May 10, 1974, as amended at 46 FR 23224, Apr. 24, 1981; 50 FR 24905, June 14, 1985. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 37827, Aug. 9, 1991; 56 FR 66785, Dec. 26, 1991] VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00758 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 749 Animal and Plant Health Inspection Service, USDA § 113.116 § 113.115 Staphylococcus Aureus Bacterin-Toxoid. Staphylococcus Aureus Bacterin Toxoid shall be prepared from toxoided broth cultures of selected toxogenic strains of Staphylococcus aureus which has been inactivated and is nontoxic. Each serial of biological product con taining Staphylococcus Aureus Bacterin-Toxoid shall meet the appli cable requirements in § 113.100 and shall be tested for purity, safety, and po tency as prescribed in this section. A serial found unsatisfactory by any pre scribed test shall not be released. (a) Purity test. Final container sam ples of completed product from each se rial shall be tested for viable bacteria and fungi as provided in § 113.26. (b) Safety test. Bulk or final container samples of completed product shall be tested for safety as provided in § 113.33(b). Also, the rabbits used in the potency test provided in paragraph (c) of this section shall constitute an addi tional safety test. If unfavorable reac tions attributable to the product occur in any of the rabbits during the obser vation period, the serial is unsatisfac tory. (c) Potency test. Rabbits, each weigh ing 2000–3000 grams, shall be used as test animals. Either a five rabbit indi vidual serum test or an eight rabbit pooled serum test shall be conducted. At the start of the test, individual se rums from the five rabbits or pooled se rums from the eight rabbits shall con tain less than 0.2 alpha antitoxin units per ml. (1) Each rabbit shall be given a series of not more than three intramuscular injections at 7 day intervals (1.0 ml, 2.0 ml, 3.0 ml) and observed from 7–14 days following the third injection. At the end of the observation period, a blood sample shall be taken from each rabbit. (2) The sample of serum from each rabbit, if the five rabbit individual test is conducted or a pooled sample of equal quantities of serum from the rab bits if the eight rabbit pooled serum test is conducted, shall be tested to de termine the staphylococcus alpha anti toxin units per ml as provided in para graphs (c)(3), (4), (5), (6), (7), and (8) of this section. (3) Inactivate rabbit serum 56 °C for 30 minutes. (4) Make serial twofold dilutions of the serum samples and conduct the test, using 1 ml of the serial dilutions. Appropriate controls should be in cluded for accurate interpretations. (5) Add 1 ml of the standardized toxin containing the established ‘‘Lh’’ dose. The ‘‘Lh’’ dose is the amount of toxin which when mixed with one unit of standard antitoxin produces a 50 per cent hemolysis of rabbit red blood cells. (6) Incubate toxin-antitoxin mixture at room temperature for 30 minutes and add 1 ml of a 1.5 percent suspension of washed freshly drawn rabbit red blood cells suspended in normal saline to each tube. Mix and incubate the combined product in a 37 °C water bath for 1 hour. Refrigerate at 5 °C over night. (7) Read the hemolysis produced and establish the 50 percent end point. The 50 percent end point of hemolysis should be established by determining the size of the button produced by the unlysed red blood cells. (8) Determine the units of antitoxin per 1 ml of serum. (9) If the individual samples from four of the five rabbits in the indi vidual serum test or the pooled sam ples from the eight rabbits in the pooled serum test do not contain three alpha antitoxin units per ml, the serial is unsatisfactory. [39 FR 16862, May 10, 1974. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66785, Dec. 26, 1991] § 113.116 Pasteurella Multocida Bacterin, Avian Isolate, Type 4. Pasteurella Multocida Bacterin, Avian Isolate, Type 4 shall be prepared from cultures of Pasteurella multocida, avian isolate, Type 4 (Little and Lyons classification), which have been inac tivated, and are nontoxic. Each serial of biological product containing Pasteurella Multocida Bacterin, Avian Isolate, Type 4, shall meet the applica ble requirements in § 113.100 and shall be tested for purity, safety, and po tency, as prescribed in this section. A serial found unsatisfactory by any pre scribed test shall not be released. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00759 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 750 9 CFR Ch. I (1–1–18 Edition) § 113.117 (a) Purity test. Final container sam ples of completed product shall be test ed for viable bacteria and fungi as pro vided in 9 CFR 113.26. (b) Safety test. Observation of the vac cinated turkeys during the prechallenge period of the potency test provided in paragraph (c) of this sec tion shall constitute the safety test. If unfavorable reactions that are attrib utable to the product occur, the serial is unsatisfactory. If unfavorable reac tions that are not attributable to the product occur in one turkey, test re sults shall be determined by observing the remaining 20 turkeys. The test is a No Test and may be repeated if unfa vorable reactions that are not attrib utable to the product occur in two or more turkeys, but the serial is unsatis factory if the test is not repeated. (c) Potency test. Bulk or final con tainer samples of completed product shall be tested for potency of the Type 4 strain, using the two-stage test pro vided in this paragraph. Turkeys at least 6 weeks old obtained from the same source and hatch shall be prop erly identified and used as provided in this paragraph. (1) Vaccinates. Each of not more than 21 turkeys shall be vaccinated with the dose and by the route recommended on the label. A second dose shall be given after 3 weeks and the turkeys observed for an additional 2-week prechallenge period. (2) Unvaccinated controls. Each of not more than 11 turkeys shall be held as controls. (3) Challenge. Not less than 14 days after the second dose, each of 20 vac cinates, and each of 10 unvaccinated controls shall be challenged intramuscularly with virulent Pasteurella multocida, Strain P–1662, Type 4 (Little and Lyons classification) and observed daily for a 14-day postchallenge period. Only dead birds shall be considered in evaluating the product. (4) Validity requirements. Eight or more unvaccinated controls must die for the test to be valid. If this require ment is met, the potency test results are evaluated according to stage one of the following table. The test is a No Test and may be repeated if the valid ity requirement is not met, but the se rial is unsatisfactory if the test is not repeated. Stage Number of vac cinates Cumu lative number of vac cinates Cummulative total number of dead vaccinates forlll Satisfactory serial Unsatisfac tory serial 1 ............ 20 20 6 or less ..... 9 or more. 2 ............ 20 40 15 or less ... 16 or more. (5) The serial shall pass or fail based on the stage one results of the potency test. However, the second stage may be conducted if seven or eight vaccinates die in stage one, but the serial is unsat isfactory if the second stage is not con ducted. (6) The second stage shall be con ducted in a manner identical to the first stage. The serial shall be evalu ated according to stage two of the table. On the basis of accumulated re sults from the data of both stage tests, a serial shall either pass or fail the sec ond stage. [47 FR 5795, Feb. 4, 1982; 47 FR 6817, Feb. 17, 1982, as amended at 52 FR 9117, Mar. 23, 1987. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66785, Dec. 26, 1991] § 113.117 Pasteurella Multocida Bacterin, Avian Isolate, Type 1. Pasteurella Multocida Bacterin, Avian Isolate, Type 1, shall be prepared from cultures of Pasteurella multocida, avian isolate, Type 1 (Little and Lyons classification), which have been inac tivated and are nontoxic. Each serial of biological product containing Pasteurella Multocida Bacterin, Avian Isolate, Type 1, shall meet the applica ble requirements in § 113.100 and shall be tested for purity, safety, and po tency as prescribed in this section. A serial found unsatisfactory by any pre scribed test shall not be released. (a) Purity test. Final container sam ples of completed product shall be test ed for viable bacteria and fungi as pro vided in § 113.26. (b) Safety test. Observation of the vac cinated chickens during the prechallenged period of the potency test provided in paragraph (c) of this section shall constitute the safety test. If unfavorable reactions that are at tributable to the product occur, the se rial is unsatisfactory. If unfavorable reactions that are not attributable to VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00760 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 751 Animal and Plant Health Inspection Service, USDA § 113.118 the product occur in one chicken, test results shall be determined by observ ing the remaining 20 chickens. The test is a No Test and may be repeated if un favorable reactions that are not attrib utable to the product occur in two or more chickens, but the serial is unsat isfactory if the test is not repeated. (c) Potency test. Bulk or final con tainer samples of completed product shall be tested for potency of the Type 1 strain, using the two-stage test pro vided in this paragraph. Chickens, at least 12 weeks of age, obtained from the same source and hatch, shall be properly identified and used as pro vided in this paragraph. (1) Vaccinates. Each of not more than 21 chickens shall be injected with the dose and by the route recommended on the label. A second dose shall be in jected after 3 weeks and the chickens observed for an additional 2 week prechallenge period. (2) Unvaccinated controls. Each of not more than 11 chickens shall be held as controls. (3) Challenge. Not less than 14 days after the second injection, each of 20 vaccinates, and each of 10 unvaccinated controls shall be challenged intramuscularly with a minimum of 250 colony-forming units of virulent Pasteurella multocida, Strain X–73, Type 1 (Little and Lyons classification) and observed daily for a 14-day postchallenge period. Only dead birds shall be considered in evaluating the product. (4) Validity requirements. Eight or more unvaccinated controls must die for the test to be valid. If these re quirement are met, the potency test re sults are evaluated according to stage one of the following table. The test is a No Test and may be repeated if the va lidity requirements are not met, but the serial is unsatisfactory if the test is not repeated. Stage Number of vac cinates Cumu lative number of vac cinates Cummulative total number of dead vaccinates forlll Satisfactory serial Unsatisfac tory serial 1 ............ 20 20 6 or less .... 9 or more. 2 ............ 20 40 15 or less .. 16 or more. (5) The serial shall pass or fail based on the stage one results of the potency test. However, the second stage may be conducted if seven or eight vaccinates die in stage one, but the serial is unsat isfactory if the second stage is not con ducted. (6) The second stage shall be con ducted in a manner identical to the first stage. The serial shall be evalu ated according to stage two of the table. On the basis of accumulated re sults from the data of both stage tests, a serial shall either pass or fail the sec ond stage. [39 FR 16866, May 10, 1974; 39 FR 20368, June 10, 1974, as amended at 40 FR 759, Jan. 3, 1975; 40 FR 23989, June 4, 1975; 47 FR 5195, Feb. 4, 1982; 52 FR 9118, Mar. 23, 1987. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66785, Dec. 26, 1991] § 113.118 Pasteurella Multocida Bacterin, Avian Isolate, Type 3. Pasteurella Multocida Bacterin, Avian Isolate, Type 3, shall be prepared from culture of Pasteurella multocida, avian isolate, Type 3 (Little and Lyons classification), which have been inac tivated and are nontoxic. Each serial of biological product containing Pasteurella Multocida Bacterin, Avian Isolate, Type 3, shall meet the applica ble requirements in § 113.100 and shall be tested for purity, safety, and po tency, as prescribed in this section. A serial found unsatisfactory by any pre scribed test shall not be released. (a) Purity test. Final container sam ples of completed product shall be test ed for viable bacteria and fungi as pro vided in § 113.26. (b) Safety test. Observation of the vac cinated turkeys during the prechallenge period of the potency test provided in paragraph (c) of this sec tion shall constitute the safety test. If unfavorable reactions that are attrib utable to the product occur, the serial is unsatisfactory. If unfavorable reac tions that are not attributable to the product occur in one turkey, test re sults shall be determined by observing the remaining 20 turkeys. The test is a No Test and may be repeated if unfa vorable reactions that are not attrib utable to the product occur in two or more turkeys, but the serial is unsatis factory if the test is not repeated. (c) Potency test. Bulk or final con tainer samples of completed product VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00761 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 752 9 CFR Ch. I (1–1–18 Edition) § 113.119 shall be tested for potency of the Type 3 strain, using the two-stage test pro vided in this paragraph. Turkeys, at least 6 weeks of age, obtained from the same source and hatch, shall be prop erly identified and used as provided in this paragraph. (1) Vaccinates. Each of not more than 21 turkeys shall be injected with the dose and by the route recommended on the label. A second dose shall be in jected after 3 weeks and the turkeys observed for an additional 2 week prechallenge period. (2) Unvaccinated controls. Each of not more than 11 turkeys shall be held as controls. (3) Challenge. Not less than 14 days after the second injection, each of 20 vaccinates, and each of 10 unvaccinated controls shall be challenged intramuscularly with a minimum of 150 colony-forming units of virulent Pasteurella multocida, Strain P–1059, Type 3 (Little and Lyons Classifica tion) and observed daily for a 14-day postchallenge period. Only dead birds shall be considered in evaluating the product. (4) Validity requirements. Eight or more unvaccinated controls must die for the test to be valid. If these re quirements are met, the potency test results are evaluated according to stage one of the following table. The test is a No Test and may be repeated if the validity requirements are not met, but the serial is unsatisfactory if the test is not repeated. Stage Number of vac cinates Cumu lative number of vac cinates Cummulative total number of dead vaccinates forlll Satisfactory serial Unsatisfac tory serial 1 ............ 20 20 6 or less ..... 9 or more. 2 ............ 20 40 15 or less ... 16 or more. (5) The serial shall pass or fail based on the stage one results of the potency test. However, the second stage may be conducted if seven or eight vaccinates die in stage one, but the serial is unsat isfactory if the second stage is not con ducted. (6) The second stage shall be con ducted in a manner identical to the first stage. The serial shall be evalu ated according to stage two of the table. On the basis of accumulated re sults from the data of both stage tests, a serial shall either pass or fail the sec ond stage. [39 FR 16862, May 10, 1974, as amended at 40 FR 759, Jan. 3, 1975; 47 FR 5196, Feb. 4, 1982; 52 FR 9118, Mar. 23, 1987. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66785, Dec. 26, 1991] § 113.119 Erysipelothrix Rhusiopathiae Bacterin. Erysipelothrix Rhusiopathiae Bacterin shall be produced from a cul ture of Erysipelothrix rhusiopathiae which has been inactivated and is nontoxic. Each serial of biological product containing Erysipelothrix rhusiopathiae shall meet the applicable requirements in § 113.100 and shall be tested for purity, safety, and potency as prescribed in this section. A serial found unsatisfactory by any prescribed test shall not be released. (a) Purity test. Final container sam ples of completed product from each se rial and each subserial shall be tested for viable bacteria and fungi as pro vided in § 113.26. (b) Safety test. Bulk or final container samples of completed product from each serial shall be tested for safety as provided in § 113.38. (c) Potency test. Bulk or final con tainer samples of completed product from each serial shall be tested for po tency using the mouse protection test provided in this paragraph. A mouse dose shall be 1⁄10 of the least dose rec ommended on the label for swine. Such swine dose shall not be less than 1 ml. (1) The ability of the bacterin being tested (Unknown) to protect mice shall be compared with a Standard Reference Bacterin (Standard) which is either supplied by or acceptable to Animal and Plant Health Inspection Service. (2) At least three threefold dilutions shall be made with the Standard and the same threefold dilutions shall be made for each Unknown. Dilutions shall be made with physiological saline solution. (3) For each dilution of the Standard and each dilution of an Unknown, a group of at least 20 mice, each weigh ing 16 to 22 grams, shall be used. Each mouse in each group shall be injected subcutaneously with one mouse dose of the appropriate dilution. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00762 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 753 Animal and Plant Health Inspection Service, USDA § 113.120 (4) Each of 20 injected mice from each group shall be challenged subcutaneously 14 to 21 days after being injected. A dose containing at least 100 mouse LD50 of a suitable cul ture of Erysipelothrix rhusiopathiae shall be used. All survivors in each group of mice shall be recorded 10 days postchallenge. (5) Test for valid assay: At least two dilutions of the Standard shall protect more than 0 percent and two dilutions shall protect less than 100 percent of the mice injected. The lowest dilution of the Standard shall protect more than 50 percent of the mice. The high est dilution of the Standard shall pro tect less than 50 percent of the mice. (6) The relative potency (RP) of the Unknown is determined by comparing the 50 percent endpoint dilution (high est bacterin dilution protecting 50 per cent of the mice) of the Unknown with that of the standard by the following formula: RP reciprocal reciprocal = of percent endpoint dilution of Unknown of percent endpoint dilution of Standard 50 50 (7) If the RP of the Unknown is less than 0.6, the serial being tested is un satisfactory. (8) If the 50 percent endpoint of an Unknown in a valid test cannot be cal culated because the lowest dilution does not exceed 50 percent protection, that serial may be retested in a man ner identical to the initial test: Pro vided, That, if the Unknown is not re tested or if the protection provided by the lowest dilution of the Standard ex ceeds the protection provided by the lowest dilution of the Unknown by six mice or more; or, if the total number of mice protected by the Standard ex ceeds the total number of mice pro tected by the Unknown by eight mice or more, the serial is unsatisfactory. (9) If the 50 percent endpoint of an Unknown in a valid test cannot be cal culated because the highest dilution exceeds 50 percent protection, the Un known is satisfactory without addi tional testing. (10) If the RP is less than 0.6, the se rial may be retested by conducting two independent replicate tests in a man ner identical to the initial test. The av erage of the RP values obtained in the retests shall be determined. If the aver age RP is less than 0.6, the serial is un satisfactory without further testing. If the average RP obtained in the retests is equal to or greater than 0.6, the fol lowing shall apply: (i) If the RP obtained in the original test is one-third or less than the aver age RP obtained in the retests, the ini tial RP may be considered a result of test system error and the serial is sat isfactory for potency. (ii) If the RP value obtained in the original test is more than one-third the average RP obtained in the retests, a new average shall be determined using the RP values obtained in all tests. If the new average is less than 0.6, the se rial is unsatisfactory. [39 FR 16862, May 10, 1974, as amended at 40 FR 759, Jan. 3, 1975; 40 FR 20067, May 8, 1975; 40 FR 51414, Nov. 5, 1975; 44 FR 71408, Dec. 11, 1979; 50 FR 23795, June 6, 1985; 51 FR 23731, July 1, 1986. Redesignated at 55 FR 35562, Aug. 31, 1990; 56 FR 66558, Dec. 24, 1991; 56 FR 66784, 66785, Dec. 26, 1991] § 113.120 Salmonella Typhimurium Bacterin. Salmonella Typhimurium Bacterin shall be prepared from a culture of Sal monella typhimurium which has been in activated and is nontoxic. Each serial of biological product containing Sal monella typhimurium fraction shall meet the applicable requirements in § 113.100 and shall be tested for purity, safety, and potency as prescribed in this section. A serial found unsatisfac tory by any prescribed test shall not be released. (a) Purity test. Final container sam ples of completed product shall be test ed for viable bacteria and fungi as pro vided in § 113.26. (b) Safety test. Bulk or final container samples of completed product from each serial shall be tested for safety as provided in § 113.33(b). (c) Potency test. Bulk or final con tainer samples of completed product from each serial shall be tested for po tency using the mouse test provided in this paragraph. A mouse dose shall be 1⁄20 of the least dose recommended on the label for other animals which shall not be less than 2 ml. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00763 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31 EC 1 4 N O 9 1 .0 1 6 < /M A T H > kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 754 9 CFR Ch. I (1–1–18 Edition) § 113.121 (1) The ability of the bacterin being tested (Unknown) to protect mice shall be compared with a Standard Reference Bacterin (Standard) which is either supplied by or acceptable to Animal and Plant Health Inspection Service. (2) At least three tenfold dilutions shall be made with the Standard and the same tenfold dilutions shall be made for each Unknown. The dilutions shall be made in Phosphate Buffered Saline. (3) For each dilution of the Standard and each dilution of an Unknown, a group of at least 20 mice, each weigh ing 16-22 grams, shall be used. Each mouse in a group shall be injected intraperitoneally with one mouse dose of the appropriate dilution. Each mouse shall be revaccinated on day 14, using the same schedule. (4) Each of 20 vaccinated mice per group shall be challenged intraperitoneally 7–10 days after the second vaccination with a 0.25 ml dose containing 100-10,000 mouse LD50 as de termined by titration, of a suitable cul ture of Salmonella typhimurium. All sur vivors in each group of mice shall be recorded 14 days postchallenge. (5) Test for valid assay: At least two dilutions of the Standard shall protect more than 0 percent and two dilutions shall protect less than 100 percent of the mice injected. The lowest dilution of the Standard shall protect more than 50 percent of the mice. The high est dilution of the Standard shall pro tect less than 50 percent of the mice. (6) The relative potency (RP) of the Unknown is determined by comparing the 50 percent endpoint dilution (high est bacterin dilution protecting 50 per cent of the mice) of the Unknown with that of the Standard by the following formula: RP reciprocal reciprocal = of percent endpoint dilution of Unknown of percent endpoint dilution of Standard 50 50 (7) If the RP of the Unknown is less than 0.30, the serial being tested is un satisfactory. (8) If the 50 percent endpoint of an Unknown cannot be calculated because the lowest dilution does not exceed 50 percent protection, that serial may be retested in a manner identical to the initial test; Provided, That, if the Un known is not retested or if the protec tion provided by the lowest dilution of the Unknown by six mice or more; or, if the total number of mice protected by the Standard exceeds the total num ber of mice protected by the Unknown by eight mice or more, the serial being tested is unsatisfactory. (9) If the 50 percent endpoint of an Unknown in a valid test cannot be cal culated because the highest dilution exceeds 50 percent protection, the Un known is satisfactory without addi tional testing. (10) If the RP is less than the min imum required in paragraph (c)(7) of this section, the serial may be retested by conducting two independent rep licate tests in a manner identical to the initial test. The average of the RP values obtained in the retests shall be determined. If the average RP is less than the required minimum, the serial is unsatisfactory. If the average RP ob tained in the retests is equal to or greater than the required minimum, the following shall apply: (i) If the RP obtained in the original test is one-third or less than the aver age RP obtained in the retests, the ini tial RP may be considered a result of test system error and the serial is sat isfactory. (ii) If the RP value obtained in the original test is more than one-third the average RP obtained in the retests, a new average shall be determined using the RP values obtained in all tests. If the new average is less than the min imum required in paragraph (c)(7) of this section, the serial is unsatisfac tory. [40 FR 17003, Apr. 16, 1975, as amended at 42 FR 59487, Nov. 18, 1977; 48 FR 31008, July 6, 1983. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66784, 66785, Dec. 26, 1991] § 113.121 Pasteurella Multocida Bacterin. Pasteurella Multocida Bacterin shall be prepared from a culture of Pasteurella multocida strains other than avian which have been inactivated and are nontoxic. Each serial of biological product containing Pasteurella VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00764 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31 EC 1 4 N O 9 1 .0 1 7 < /M A T H > kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 755 Animal and Plant Health Inspection Service, USDA § 113.121 multocida fraction shall meet the appli cable requirements in § 113.100 and shall be tested for purity, safety, and po tency as prescribed in this section. A serial found unsatisfactory by any pre scribed test shall not be released. (a) Purity test. Final container sam ples of completed product from each se rial and each subserial shall be tested for viable bacteria and fungi as pro vided in § 113.26. (b) Safety test. Bulk or final container samples of completed product from each serial shall be tested for safety as provided in § 113.33(b). The subcuta neous route is to be used. (c) Potency test. Bulk or final con tainer samples of completed product from each serial shall be tested for po tency using the mouse test provided in this paragraph. A mouse dose shall be 1⁄20 of the least dose recommended on the label for other animals which shall not be less than 2 ml. (1) The ability of the bacterin being tested (Unknown) to protect mice shall be compared with a Standard Reference Bacterin (Standard) which is either supplied by or acceptable to Animal and Plant Health Inspection Service. (2) At least three fivefold dilutions shall be made with the Standard and the same fivefold dilutions shall be made for each Unknown. The dilutions will be made in Phosphate Buffered Sa line. (3) For each dilution of the Standard and each dilution of each Unknown, a group of at least 20 mice, each weigh ing 16–22 grams, shall be used. Each mouse in a group shall be injected intraperitoneally with one mouse dose of the appropriate dilution. Each mouse shall be revaccinated on day 14, using the same schedule. (4) Each of 20 injected mice per group shall be challenged intraperitoneally 10–12 days after the second vaccination with a 0.2 ml dose containing 100–10,000 mouse LD50, as determined by titra tion, of a suitable culture of Pasteurella multocida. All survivors in each group of mice shall be recorded 10 days postchallenge. (5) Test for valid assay: At least two dilutions of the Standard shall protect more than 0 percent and two dilutions shall protect less than 100 percent of the mice injected. The lowest dilution of the Standard shall protect more than 50 percent of the mice. The high est dilution of the Standard shall pro tect less than 50 percent of the mice. (6) The relative potency (RP) of the Unknown is determined by comparing the 50 percent endpoint dilution (high est bacterin dilution protecting 50 per cent of the mice) of the Unknown with that of the Standard by the following formula: RP reciprocal reciprocal = of percent endpoint dilution of Unknown of percent endpoint dilution of Standard 50 50 (7) If the RP of the Unknown is less than 0.50, the serial being tested is un satisfactory. (8) If the 50 percent endpoint of an Unknown cannot be calculated because the lowest dilution does not exceed 50 percent protection, that serial may be retested in a manner identical to the initial test: Provided, That, if the Un known is not retested or if the protec tion provided by the lowest dilution of the Standard exceeds the protection provided by the lowest dilution of the Unknown by six mice or more; or, if the total number of mice protected by the Standard exceeds the total number of mice protected by the Unknown by eight mice or more, the serial being tested is unsatisfactory. (9) If the 50 percent endpoint of an Unknown in a valid test cannot be cal culated because the highest dilution exceeds 50 percent protection, the Un known is satisfactory without addi tional testing. (10) If the RP is less than the min imum required in paragraph (c)(7) of this section, the serial may be retested by conducting two independent rep licate tests in a manner identical to the initial test. The average of the RP values obtained in the retests shall be determined. If the average RP is less than the required minimum, the serial is unsatisfactory. If the average RP ob tained in the retests is equal to or greater than the required minimum, the following shall apply: (i) If the RP obtained in the original test is one-third or less than the aver age RP obtained in the retests, the ini tial RP may be considered a result of VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00765 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31 EC 1 4 N O 9 1 .0 1 8 < /M A T H > kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 756 9 CFR Ch. I (1–1–18 Edition) § 113.122 test system error and the serial is sat isfactory. (ii) If the RP value obtained in the original test is more than one-third the average RP obtained in the retests, a new average shall be determined using the RP values obtained in all tests. If the new average is less than the min imum required in paragraph (c)(7) of this section, the serial is unsatisfac tory. [40 FR 17004, Apr. 16, 1975, as amended at 42 FR 59487, Nov. 18, 1977; 48 FR 31008, July 6, 1983. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66784, 66785, Dec. 26, 1991] § 113.122 Salmonella Choleraesuis Bacterin. Salmonella Choleraesuis Bacterin shall be prepared from a culture of Sal monella choleraesuis which has been in activated and is nontoxic. Each serial of biological product containing Sal monella choleraesuis fraction shall meet the applicable requirements in 9 CFR 113.100 and shall be tested for purity, safety, and potency as prescribed in this section. A serial found unsatisfac tory by any prescribed test shall not be released. (a) Purity test. Final container sam ples of completed product shall be test ed for viable bacteria and fungi as pro vided in 9 CFR 113.26. (b) Safety test. Bulk or final container samples of completed product from each serial shall be tested for safety as provided in 9 CFR 113.33(b). The subcutaneous route shall be used when the product is in combination with Pasteurella Multocida Bacterin. (c) Potency test. Bulk or final con tainer samples of completed product from each serial shall be tested for po tency using the mouse test provided in this paragraph. A mouse dose shall be 1⁄20 of the least dose recommended on the label for other animals which shall not be less than 2 ml. (1) The ability of the bacterin being tested (Unknown) to protect mice shall be compared with a Standard Reference Bacterin (Standard) which is either supplied by or acceptable to Veterinary Services. (2) At least three fivefold dilutions shall be made with the Standard and the same fivefold dilution shall be made for each Unknown. The dilutions shall be made in Phosphate-Buffered Saline. (3) For each dilution of the Standard and each dilution of an Unknown, a group of at least 20 mice, each weigh ing 16 to 22 grams, shall be used. Each mouse in a group shall be injected intraperitoneally with one mouse dose of the appropriate dilution. Each mouse shall be revaccinated on day 14, using the same schedule. (4) Each of 20 vaccinated mice per group shall be challenged intraperitoneally 7 to 10 days after the second vaccination with a 0.25 ml dose containing 10–1,000 mouse LD50 as de termined by titration of a suitable cul ture of Salmonella choleraesuis. All sur vivors in each group of mice shall be recorded 14 days postchallenge. (5) Test for valid assay: At least two dilutions of the Standard shall protect more than 0 percent and two dilutions shall protect less than 100 percent of the mice injected. The lowest dilution of the Standard shall protect more than 50 percent of the mice. The high est dilution of the Standard shall pro tect less than 50 percent of the mice. (6) The relative potency (RP) of the Unknown is determined by comparing the 50 percent endpoint dilution (high est bacterin dilution protecting 50 per cent of the mice) of the Unknown with that of the Standard by the following formula: RP reciprocal reciprocal = of percent endpoint dilution of Unknown of percent endpoint dilution of Standard 50 50 (7) If the RP of the Unknown is less than 0.50, the serial being tested is un satisfactory. (8) If the 50 percent endpoint of an Unknown cannot be calculated because the lowest dilution does not exceed 50 percent protection, that serial may be retested in a manner identical to the initial test; Provided, That, if the Un known is not retested or if the protec tion provided by the lowest dilution of the Standard exceeds the protection provided by the lowest dilution of the Unknown by six mice or more; or, if the total number of mice protected by the Standard exceeds the total number VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00766 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31 EC 1 4 N O 9 1 .0 1 9 < /M A T H > kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 757 Animal and Plant Health Inspection Service, USDA § 113.123 of mice protected by the Unknown by eight mice or more, the serial being tested is unsatisfactory. (9) If the 50 percent endpoint of an Unknown in a valid test cannot be cal culated because the highest dilution exceeds 50 percent protection, the Un known is satisfactory without addi tional testing. (10) If the RP is less than the min imum required in paragraph (c)(7) of this section, the serial may be retested by conducting two independent rep licate tests in a manner identical to the initial test. The average of the RP values obtained in the retests shall be determined. If the average RP is less than the required minimum, the serial is unsatisfactory. If the average RP ob tained in the retests is equal to or greater than the required minimum, the following shall apply: (i) If the RP obtained in the original test is one-third or less than the aver age RP obtained in the retests, the ini tial RP may be considered a result of test system error and the serial is sat isfactory. (ii) If the RP value obtained in the original test is more than one-third the average RP obtained in the retests, a new average shall be determined using the RP values obtained in all tests. If the new average is less than the min imum required in paragraph (c)(7) of this section, the serial is unsatisfac tory. [43 FR 25077, June 9, 1978, as amended at 48 FR 31008, July 6, 1983. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66785, Dec. 26, 1991] § 113.123 Salmonella Dublin Bacterin. Salmonella Dublin Bacterin shall be prepared from a culture of Salmonella dublin which has been inactivated and is nontoxic. Each serial of biological product containing Salmonella dublin fraction shall meet the applicable re quirements in 9 CFR 113.100 and shall be tested for purity, safety, and po tency as prescribed in this section. A serial found unsatisfactory by any pre scribed test shall not be released. (a) Purity test. Final container sam ples of completed product shall be test ed for viable bacteria and fungi as pro vided in 9 CFR 113.26. (b) Safety test. Bulk or final container samples of completed product from each serial shall be tested for safety as provided in 9 CFR 113.33(b). (c) Potency test. Bulk or final con tainer samples of completed product from each serial shall be tested for po tency using the mouse test provided in this paragraph. A mouse dose shall be 1⁄20 of the least dose recommended on the label for other animals which shall not be less than 2 ml. (1) The ability of the bacterin being tested (Unknown) to protect mice shall be compared with a Standard Reference Bacterin (Standard) which is either supplied by or acceptable to Veterinary Services. (2) At least three tenfold dilutions shall be made with the Standard and the same tenfold dilutions shall be made for each Unknown. The dilutions shall be made in Phosphate-Buffered Saline. (3) For each dilution of the Standard and each dilution of an Unknown, a group of at least 20 mice, each weigh ing 16 to 22 grams, shall be used. Each mouse in a group shall be injected intraperitoneally with one mouse dose of the appropriate dilution. Each mouse shall be revaccinated on day 14, using the same schedule. (4) Each of 20 vaccinated mice per group shall be challenged intraperitoneally 7 to 10 days after the second vaccination with a 0.25 ml dose containing 1,000–100,000 mouse LD50 as determined by titration of a suitable culture of Salmonella dublin. All sur vivors in each group of mice shall be recorded 14 days postchallenge. (5) Test for valid assay: At least two dilutions of the Standard shall protect more than 0 percent and two dilutions shall protect less than 100 percent of the mice injected. The lowest dilution of the Standard shall protect more than 50 percent of the mice. The high est dilution of the Standard shall pro tect less than 50 percent of the mice. (6) The relative potency (RP) of the Unknown is determined by comparing the 50 percent endpoint dilution (high est bacterin dilution protecting 50 per cent of the mice) of the Unknown with that of the Standard by the following formula: VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00767 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 758 9 CFR Ch. I (1–1–18 Edition) § 113.200 RP reciprocal reciprocal = of percent endpoint dilution of Unknown of percent endpoint dilution of Standard 50 50 (7) If the RP of the Unknown is less than 0.30, the serial being tested is un satisfactory. (8) If the 50 percent endpoint of an Unknown cannot be calculated because the lowest dilution does not exceed 50 percent protection, that serial may be retested in a manner identical to the initial test; Provided, That, if the Un known is not retested or if the protec tion provided by the lowest dilution of the Standard exceeds the protection provided by the lowest dilution of the Unknown by six mice or more; or, if the total number of mice protected by the Standard exceeds the total number of mice protected by the Unknown by eight mice or more, the serial being tested is unsatisfactory. (9) If the 50 percent endpoint of an Unknown in a valid test cannot be cal culated because the highest dilution exceeds 50 percent protection, the Un known is satisfactory without addi tional testing. (10) If the RP is less than the min imum required in paragraph (c)(7) of this section, the serial may be retested by conducting two independent rep licate tests in a manner identical to the initial test. The average of the RP values obtained in the retests shall be determined. If the average RP is less than the required minimum, the serial is unsatisfactory. If the average RP ob tained in the retests is equal to or greater than the required minimum, the following shall apply: (i) If the RP obtained in the original test is one-third or less than the aver age RP obtained in the retests, the ini tial RP may be considered a result of test system error and the serial is sat isfactory. (ii) If the RP value obtained in the original test is more than one-third the average RP obtained in the retests, a new average shall be determined using the RP values obtained in all tests. If the new average is less than the min imum required in paragraph (c)(7) of this section, the serial is unsatisfac tory. [43 FR 25077, June 9, 1978, as amended at 48 FR 31009, July 6, 1983. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66785, Dec. 26, 1991] KILLED VIRUS VACCINES § 113.200 General requirements for killed virus vaccines. When prescribed in an applicable Standard Requirement or in the filed Outline of Production, a killed virus vaccine shall meet the applicable re quirements in this section. (a) Killing agent. The vaccine virus shall be killed (inactivated) by an ap propriate agent. The procedure in volved may be referred to as inactiva tion. Suitable tests to assure complete inactivation shall be written into the filed Outline of Production. (b) Cell culture requirements. If cell cultures are used in the preparation of the vaccine, primary cells shall meet the requirements in § 113.51 and cell lines shall meet the requirements in § 113.52. (c) Purity tests—(1) Bacteria and fungi. Final container samples of completed product from each serial shall be tested as prescribed in § 113.26. (2) Avian origin vaccine. Bulk pooled material or final container samples from each serial shall also be tested for: (i) Salmonella contamination as pre scribed in § 113.30; and (ii) Lymphoid leukosis virus contami nation as prescribed in § 113.31; and (iii) Hemagglutinating viruses as pre scribed in § 113.34. (3) Mycoplasma. If the licensee cannot demonstrate that the agent used to kill the vaccine virus would also kill myco plasma, each serial of the vaccine shall be tested for mycoplasma as prescribed in § 113.28, prior to adding the killing agent. Material found to contain myco plasma is unsatisfactory for use. (4) Extraneous viruses. Each lot of Master Seed Virus used to prepare killed virus vaccine recommended for animals other than poultry shall meet the requirements for extraneous vi ruses as prescribed in § 113.55. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00768 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31 EC 1 4 N O 9 1 .0 2 0 < /M A T H > kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 759 Animal and Plant Health Inspection Service, USDA § 113.205 2 The procedures for performing the ferric chloride test for residual free formaldehyde may be obtained from USDA, APHIS, Center for Veterinary Biologics-Laboratory, 1800 Dayton Road, P.O. Box 844, Ames, IA 50010. (d) Safety tests. Final container sam ples of completed product from each se rial shall be tested for safety in guinea pigs as prescribed in § 113.38 and for safety in mice as prescribed in § 113.33: Provided, That, vaccines recommended for use only in poultry are exempt from this requirement. (e) Viricidal activity test. Only serials tested for viricidal activity in accord ance with the test provided in § 113.35 and found satisfactory by such test shall be packaged as diluent for des iccated fractions in combination pack ages. (f) Formaldehyde content. If formalde hyde is used as the killing agent, the residual free formaldehyde content must not exceed 0.74 grams per liter (g/ L) as determined using the ferric chlo ride test. 2 Firms currently using tests for residual free formaldehyde content other than the ferric chloride test have until July 14, 2004 to update their Out line of Production to be in compliance with this requirement. [39 FR 27428, July 29, 1974, as amended at 40 FR 23989, June 4, 1975; 43 FR 49528, Oct. 24, 1978. Redesignated at 55 FR 35562, Aug. 31, 1990; 68 FR 35283, June 13, 2003; 79 FR 31021, May 30, 2014] §§ 113.201–113.203 [Reserved] § 113.204 Mink Enteritis Vaccine, Killed Virus. Mink Enteritis Vaccine, Killed Virus, shall be prepared from virus-bearing cell culture fluids or tissues obtained from mink that have developed mink enteritis following inoculation with virulent mink enteritis virus. Each se rial shall meet the applicable require ments prescribed in § 113.200 and special requirements prescribed in this sec tion. Any serial found unsatisfactory by a prescribed test shall not be re leased. (a) Safety test. Vaccinates used in the potency test in paragraph (b) of this section shall be observed each day prior to challenge. If unfavorable reac tions attributable to the vaccine occur, the serial is unsatisfactory. If unfavor able reactions not attributable to the vaccine occur, the test shall be de clared a No Test and may be repeated: Provided, That, if the test is not re peated, the serial is unsatisfactory. (b) Potency test. Bulk or final con tainer samples of completed product shall be tested for potency using 10 mink enteritis susceptible mink (five vaccinates and five controls) as fol lows: (1) Vaccination. Each of the five vac cinates shall be injected with one dose of vaccine as recommended on the label and observed each day for 14 days. (2) Challenge. At least 2 weeks after the last inoculation, the five vac cinates and the five controls shall be challenged with virulent mink enter itis virus and observed each day for 12 days. Fecal material shall be collected on one day between days 4–8 (inclusive) postchallenge from each test animal that remains free of enteric signs and tested for the presence of mink enter itis virus by cell culture with fluores cent antibody examination. (3) Interpretation. A serial is satisfac tory if at least 80 percent of the vac cinates remain free of enteric signs and do not shed virus in the feces, while at least 80 percent of the controls develop clinical signs of mink enteritis or shed virus in the feces. If at least 80 percent of the vaccinates remain free of enteric signs and do not shed virus in the feces, while less than 80 percent of the con trols develop clinical signs of mink en teritis or shed virus in the feces, the test is considered a No Test and may be repeated: Provided, That, if at least 80 percent of the vaccinates do not re main well and free of detectable virus in the feces, the serial is unsatisfac tory. [39 FR 27428, July 29, 1974. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66786, Dec. 26, 1991; 60 FR 14361, Mar. 17, 1995] § 113.205 Newcastle Disease Vaccine, Killed Virus. Newcastle Disease Vaccine (Killed Virus) shall be prepared from virus bearing tissues or fluids obtained from embryonated chicken eggs or cell cul tures. With the exception of § 113.200(c)(2)(iii), each serial shall meet the applicable general requirements VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00769 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 760 9 CFR Ch. I (1–1–18 Edition) § 113.206 prescribed in § 113.200 and special re quirements prescribed in this section. A serial found unsatisfactory by a pre scribed test shall not be released. (a) Safety test. The prechallenge part of the potency test in paragraph (b) of this section shall constitute a safety test. If unfavorable reactions attrib utable to the product occur in any of the vaccinates, the serial is unsatisfac tory. If unfavorable reactions which are not attributable to the product occur, the test shall be declared a No Test and may be repeated: Provided, That, if the test is not repeated, the se rial shall be declared unsatisfactory. (b) Potency test. A vaccination-chal lenge test shall be conducted using sus ceptible chickens 2 to 6 weeks of age at time of vaccination, properly identified and obtained from the same source and hatch. (1) Ten or more chickens shall be vac cinated as recommended on the label and kept isolated under observation for at least 14 days. (2) After at least 14 days post-vac cination, the vaccinates and at least 10 unvaccinated chickens that have been kept isolated as controls shall be chal lenged with a virulent strain of New castle disease virus supplied by or ap proved by Veterinary Services and the vaccinates observed each day for 14 days. (3) If at least 90 percent of the con trols do not show typical signs of New castle disease or die, the test is a No Test and may be repeated. If at least 90 percent of the vaccinates do not re main normal, the serial is unsatisfac tory. [39 FR 27428, July 29, 1974. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66786, Dec. 26, 1991] § 113.206 Wart Vaccine, Killed Virus. Wart Vaccine, Killed Virus, shall be prepared from virus-bearing epidermal tumors (warts) obtained from a bovine. Each serial shall meet the require ments prescribed in this section and any serial found unsatisfactory by a prescribed test shall not be released. (a) Purity. Final container samples of completed product shall meet the re quirements for purity as prescribed in § 113.200 (c)(1) and (3). (b) Safety. Bulk or final container samples of completed product shall meet the requirements for safety as prescribed in §§ 113.33(b) and 113.38. (c) Formaldehyde content. Bulk or final container samples of completed product shall meet the requirements for formaldehyde content as prescribed in § 113.200(f). (d) Potency and efficacy. The efficacy of wart vaccine has been demonstrated to the satisfaction of Veterinary Serv ices as being a valuable biological product. The inherent nature of the product precludes the possible develop ment of serial to serial potency tests and none is required: Provided, That, (1) The vaccine shall be a tissue ex tract representing at least 10 percent weight to volume suspension of wart tissue; and (2) The vaccine shall be limited to use in the prevention of warts in cat tle. Labeling recommendations shall be in accordance with § 112.7(h). [40 FR 14084, Mar. 28, 1975, as amended at 40 FR 23989, June 4, 1975; 40 FR 30803, July 23, 1975. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66786, Dec. 26, 1991; 81 FR 59436, Aug. 30, 2016] § 113.207 Encephalomyelitis Vaccine, Eastern, Western, and Venezuelan, Killed Virus. Encephalomyelitis Vaccine, Eastern, Western, and Venezuelan, Killed Virus, shall be prepared from virus-bearing cell culture fluids. Each serial or sub serial shall meet the requirements pre scribed in this section and the general requirements prescribed in § 113.200, ex cept those in § 113.200(d). Any serial or subserial found unsatisfactory by a prescribed test shall not be released. (a) Safety test. Bulk samples of com pleted product from each serial shall be tested for encephalomyelitis virus in activation. (1) Each of at least ten 6 to 12 hour old chickens shall be injected subcutaneously with 0.5 ml of the prod uct and the chickens observed each day for 10 days. (2) If unfavorable reactions attrib utable to the product occur in the chickens during the observation period, the serial is unsatisfactory. If unfavor able reactions not attributable to the product occur, the test is a No Test and VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00770 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 761 Animal and Plant Health Inspection Service, USDA § 113.208 may be repeated: Provided, That, if the test is not repeated, the serial is unsat isfactory. (b) Potency test. Bulk or final con tainer samples of completed product from each serial shall be tested for po tency in accordance with the two-stage test provided in this paragraph. For each fraction contained in the prod uct—Eastern type, Western type, or Venezuelan type—the serological inter pretations required in this test shall be made independently. A serial or sub serial found unsatisfactory for any of the fractions shall not be released. (1) For this test, a guinea pig dose shall be one-half the amount rec ommended on the label for a horse and shall be administered as recommended for a horse. Each of 10 healthy guinea pigs (vaccinates) shall be injected with two guinea pig doses with an interval of 14 to 21 days between doses. Two ad ditional guinea pigs from the same source shall be held as controls. (2) Fourteen to 21 days after the sec ond injection, serum samples from each vaccinate and each control shall be tested by a plaque reduction, serum neutralization test using Vero 76 cells. (3) If the control serum samples show a titer of 1:4 or greater for any frac tion, the test is a No Test for that frac tion and may be repeated: Provided, That, if four or more of the vaccinate serum samples show a titer of less than 1:40 for the Eastern type fraction, less than 1:40 for the Western type fraction, or less than 1:4 for the Venezuelan type fraction, the serial or subserial is un satisfactory without further testing. (4) If two or three of the vaccinate serum samples show a titer of less than 1:40 for the Eastern type fraction, less than 1:40 for the Western type fraction, or less than 1:4 for the Venezuelan type fraction, the second stage of the test may be used for the relevant frac tion(s): Provided, That, if a fraction is found acceptable by the first stage of the test, the second stage need not be conducted for that fraction. (5) If the second stage is used and four or more of the vaccinate serum samples show a titer of less than 1:40 for the Eastern type fraction or the Western type fraction, or less than 1:4 for the Venezuelan type fraction, the serial or subserial is unsatisfactory. (6) The results shall be evaluated ac cording to the following table: CUMULATIVE TOTALS Stage Vaccinates Failures for acceptance Failures for rejection 1 ................... 10 ................ 1 or less ...... 4 or more. 2 ................... 20 ................ 3 or less ...... Do. [39 FR 44714, Dec. 27, 1974, as amended at 40 FR 14084, Mar. 28, 1975; 42 FR 45284, Sept. 9, 1977. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66786, Dec. 26, 1991; 61 FR 67930, Dec. 26, 1996] § 113.208 Avian Encephalomyelitis Vaccine, Killed Virus. Avian Encephalomyelitis Vaccine (Killed Virus) shall be prepared from virus-bearing tissues or fluids obtained from embryonated chicken eggs. Each serial shall meet the general require ments prescribed in § 113.200 and the re quirements prescribed in this section. Any serial found unsatisfactory by a prescribed test shall not be released. (a) Safety tests. (1) The prechallenge part of the potency test prescribed in paragraph (b) of this section shall con stitute a safety test. If any of the vac cinates develop clinical signs of disease or die due to causes attributable to the product, the serial is unsatisfactory. (2) An inactivation test for viable avian encephalomyelitis (AE) virus shall be conducted on each serial. The test shall be conducted using suscep tible chicken embryos: Provided, That, if a non-embryo adapted virus is used for vaccine production, the test shall be conducted in susceptible chickens. (i) Chicken Embryo Test. Each of 15 or more AE susceptible 5 or 6 day old em bryos shall be injected in the yolk sac with 0.2 ml of the vaccine. For a valid test, at least 80 percent of the embryos shall survive for 48 hours post-inocula tion (PI). Eleven to 13 days PI, all em bryos surviving the 48 hour PI period shall be examined for gross lesions of AE; all these embryos shall be normal or the serial is unsatisfactory. Concur rently, five additional embryos from the same source shall be injected with live AE virus of the production strain to serve as positive controls. At least 4 of the 5 embryos shall show evidence of AE virus infection during the 11 to 13 VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00771 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 762 9 CFR Ch. I (1–1–18 Edition) § 113.209 day PI period or the test shall be con sidered a No Test and repeated: Pro vided, That, if the test is not repeated, the serial shall be declared unsatisfac tory. (ii) Chicken test. Each of 10 or more AE susceptible 7 day old chickens shall be injected intracerebrally with 0.1 ml vaccine each. The chickens shall be ob served each day for 28 days. If any chickens show clinical signs of AE, the serial is unsatisfactory. Concurrently, 5 additional chickens from the same source shall be injected intracerebrally with live AE virus of the production strain to serve as positive controls. At least 4 of the 5 controls shall show evi dence of AE virus infection during the observation period or the test shall be a No Test and may be repeated: Pro vided, That, if the test is not repeated, the serial shall be unsatisfactory. (b) Potency test. Bulk or final con tainer samples of completed product from each serial or one subserial shall be tested. Ten or more AE-susceptible chickens (vaccinates), 4 weeks or older, properly identified and obtained from the same source and hatch, shall be in jected as recommended on the label. At least 10 additional AE-susceptible chickens, properly identified and ob tained from the same source and hatch shall be kept in isolation as controls. (1) At least 28 days post-injection, the vaccinates and the controls shall be challenged intramuscularly with a virulent AE virus and the chickens ob served each day for 21 days. (2) If at least 80 percent of the con trols do not show clinical signs of or die from AE infection, the test is a No Test and may be repeated. (3) If at least 80 percent of the vac cinates do not remain normal, the se rial is unsatisfactory. [39 FR 12958, Dec. 27, 1974, as amended at 40 FR 41088, Sept. 5, 1975. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66786, Dec. 26, 1991] § 113.209 Rabies Vaccine, Killed Virus. Rabies Vaccine (Killed Virus) shall be prepared from virus-bearing cell cul tures or nerve tissues obtained from animals that have developed rabies in fection following injection with rabies virus. Only Master Seed Virus which has been established as pure, safe, and immunogenic shall be used for pre paring the production seed virus for vaccine production. All serials of vac cine shall be prepared from the first through the fifth passage from the Master Seed Virus. (a) The Master Seed Virus shall meet the applicable requirements prescribed in § 113.200 and the requirements pre scribed in this section. (1) Each lot of Master Seed Virus propagated in tissue or cells of avian origin shall also be tested for extra neous pathogens by procedures pre scribed in § 113.37. (2) Each lot of Master Seed Virus propagated in primary cell cultures of mouse or hamster origin or brain tis sues of mouse origin shall be tested for lymphocytic choriomeningitis (LCM) virus by the procedure prescribed in § 113.42. If LCM virus is detected, the Master Seed Virus is unsatisfactory. (b) The immunogenicity of vaccine prepared with virus at the highest pas sage from the Master Seed shall be es tablished in each species for which the vaccine is recommended. Tests shall be conducted in accordance with a pro tocol filed with Animal and Plant Health Inspection Service before initi ation of the tests. The vaccine shall be prepared using methods prescribed in the Outline of Production. If Rabies Vaccine is to be in combination with other fractions, the product to be test ed shall include all fractions to be test ed. (1) The preinactivation virus titer must be established as soon as possible after harvest by at least five separate virus titrations. A mean relative po tency value of the vaccine to be used in the host animal potency test must be established by at least five replicate potency tests conducted in accordance with the standard NIH test for potency in chapter 37 of ‘‘Laboratory Tech niques in Rabies,’’ Fourth Edition (1996), edited by F.X. Meslin, M.M. Kaplan, and H. Koprowski, World Health Organization, Geneva, Switzer land (ISBN 92 4 154479 1). The provisions of chapter 37 of ‘‘Laboratory Tech niques in Rabies,’’ Fourth Edition (1996), are the minimum standards for achieving compliance with this section and are incorporated by reference. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00772 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 763 Animal and Plant Health Inspection Service, USDA § 113.209 These provisions state that the chal lenge virus standard to be used as the challenge in the NIH test and the ref erence vaccine for the test are avail able from the national control author ity. In the United States, that author ity is the Animal and Plant Health In spection Service’s Center for Veteri nary Biologics Laboratory, located at 1920 Dayton Avenue, P.O. Box 844, Ames, IA 50010; phone (515) 337-6100; fax (515) 337-6120. This incorporation by ref erence was approved by the Director of the Federal Register in accordance with 5 U.S.C. 552(a) and 1 CFR part 51. Copies may be obtained from the World Health Organization Publications Cen ter USA, 49 Sheridan Avenue, Albany, NY 12210. Copies may be inspected at the Animal and Plant Health Inspec tion Service, Center for Veterinary Biologics, Policy, Evaluation, and Li censing, 1920 Dayton Avenue, P.O. Box 844, Ames, IA 50010, or at the National Archives and Records Administration (NARA). For information on the avail ability of this material at NARA, call 202–741–6030, or go to: http:// www.archives.gov/federallregister/ codeloflfederallregulations/ ibrllocations.html. (2) The dose of vaccine to be used in the immunogenicity test shall be no more than the amount which, on the basis of The NIH Test For Potency, has been diluted to the proposed minimum acceptable potency value. 1 (3) Test animals shall be uniform and have no neutralizing antibodies to ra bies as determined by serum-neutral ization (SN) tests. (i) Twenty-five or more animals shall be used as vaccinates. Each shall be ad ministered a dose of vaccine at the pro posed minimum potency level and by the method specified in the Outline of Production. (ii) Ten or more additional animals shall be held as controls. (iii) On or about 30, 90, 180, 270, and 365 days postvaccination, all test ani mals shall be bled and individual serum samples tested for neutralizing anti bodies to rabies virus. (iv) All surviving test animals shall be challenged intramuscularly with virulent rabies virus furnished or ap proved by Animal and Plant Health In spection Service 1 year after vaccina tions, except as provided in (b)(4) of this section. The challenged animals shall be observed each day for 90 days as prescribed in § 113.5(b). The brain of each test animal that dies following challenges shall be examined for rabies by the fluorescent antibody test or other method acceptable to Animal and Plant Health Inspection Service. (v) Requirements for acceptance in challenge tests shall be death due to rabies in at least 80 percent of the con trols while at least 22 of 25 or 26 of 30 or a statistically equivalent number of the vaccinates remain well for a period of 90 days. (4) An alternative to challenging all surviving test animals in accordance with paragraph (b)(3)(iv) of this section may be used when the test animals are of species other than carnivores. Vac cinates shall be challenged at 1 year postvaccination. These shall include five vaccinates with the lowest SN titers at the 270th-day bleeding, five vaccinates with the lowest SN titers at the 365th-day bleeding, and all vac cinates with SN titers below 1:10 by the mouse SN test or below 1:16 by the rapid-fluorescent-focus-inhibition test at any bleeding. At least five SN-nega tive controls of each species shall be challenged at the same time as the vaccinates. All SN titers shall be titrated to an endpoint. All of the chal lenged vaccinates must remain well for a period of 90 days, and at least 80 per cent of the controls must die of rabies for a satisfactory test without further challenge. If one or more of the vac cinates die from rabies, all the remain ing vaccines, regardless of titer, along with the five controls shall be chal lenged. The cumulative results from the two challenges shall be evaluated for acceptance as specified in para graph (b)(3)(v) of this section. (5) An Outline of Production change shall be made before authority for use a new lot of Master Seed Virus shall be granted by Animal and Plant Health Inspection Service. (c) If more than 1 year duration of immunity is to be claimed, a duration of immunity test for the additional time shall be conducted and inter preted as prescribed in paragraph (b) of this section for the 1 year test. The VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00773 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 764 9 CFR Ch. I (1–1–18 Edition) § 113.210 test animals shall be monitored sero logically at least every 180 days. The time of challenge may be adjusted ac cordingly. (d) Test requirements for release: Each serial and each subserial shall meet the general requirements pre scribed in § 113.200 and special require ments in this paragraph. (1) Purity test. Primary cell cultures of hamster origin or brain tissues of mouse origin used in vaccine produc tion shall be tested for LCM virus as prescribed in § 113.42. Hamster origin cells shall be disrupted and undiluted cell fluids from each lot shall be tested. Where mouse brains are used in produc tion, at least five mice which have not been injected with rabies virus shall be sacrificed and a 10 percent suspension of brain material shall be prepared and tested. (2) Safety tests. Bulk samples from each serial shall be tested for virus in activation and safety as follows: (i) At the end of the inactivation pe riod, each of 20 12 to 16 gram mice shall be injected intracerebrally with 0.03 ml and two rabbits shall be injected into each cerebral hemisphere with 0.25 ml and observed each day for 21 days. The brains of animals dying between the fourth and 21st day post-injection shall be checked for rabies virus. Material from each brain recovered shall be in jected into each of five mice and the mice observed each day for 14 days. The fluorescent antibody test or serum neu tralization test shall be used to con firm the presence or absence or live ra bies virus. If live rabies virus is con firmed, the serial is unsatisfactory un less reprocessed in accordance with § 114.18. (ii) A test for safety in three young seronegative animals of the most sus ceptible species for which the vaccine is recommended shall be conducted. Each shall in injected intramuscularly with one recommended dose of vaccine. If unfavorable reactions attributable to the product occur during a 28 day ob servation period, the serial is unsatis factory. (3) Potency test. Bulk or final con tainer samples of completed product from each serial must be tested for po tency by tests conducted in accordance with the standard NIH test for potency in Chapter 37 of ‘‘Laboratory Tech niques in Rabies,’’ Fourth Edition (1996), which is incorporated by ref erence at paragraph (b)(1) of this sec tion. The relative potency of each se rial must be at least equal to that used in an approved host animal immunogenicity test. [39 FR 44715, Dec. 27, 1974, as amended at 42 FR 6794, Feb. 4, 1977; 43 FR 49528, Oct. 24, 1978; 50 FR 20090, May 14, 1985. Redesignated at 55 FR 35562, Aug. 31, 1990; 56 FR 66784, 66786, Dec. 26, 1991; 61 FR 31823, June 21, 1996; 64 FR 45420, Aug. 20, 1999; 69 FR 18803, Apr. 9, 2004; 75 FR 20773, Apr. 21, 2010] § 113.210 Feline Calicivirus Vaccine, Killed Virus. Feline Calicivirus Vaccine, Killed Virus, shall be prepared from virus bearing cell culture fluids. Only Master Seed which has been established as pure, safe, and immunogenic shall be used for preparing seeds for vaccine production. All serials of vaccine shall be prepared from the first through the fifth passage from the Master Seed. (a) The Master Seed shall meet the applicable general requirements pre scribed in § 113.200. (b) The Master Seed shall be tested for chlamydial agents as prescribed in § 113.43. (c) The immunogenicity of vaccine prepared from the Master Seed in ac cordance with the Outline of Produc tion shall be established by a method acceptable to Animal and Plant Health Inspection Service. Vaccine used for this test shall be at the highest passage from the Master Seed and prepared at the minimum preinactivation titer specified in the Outline of Production. (d) Test requirements for release. Each serial and subserial shall meet the ap plicable general requirements pre scribed in § 113.200 and the special re quirements provided in this paragraph. Any serial or subserial found unsatis factory by a prescribed test shall not be released. (1) Safety. Vaccinates used in the po tency test in paragraph (d)(2) of this section shall be observed each day dur ing the prechallenge period. If unfavor able reactions occur, including oral le sions, which are attributable to the vaccine, the serial is unsatisfactory. If unfavorable reactions occur which are VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00774 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 765 Animal and Plant Health Inspection Service, USDA § 113.212 not attributable to the vaccine, the test is inconclusive and may be re peated. If the test is not repeated, the serial is unsatisfactory. (2) Potency. Bulk or final container samples of completed product shall be treated for potency as follows: (i) Eight feline calicivirus susceptible cats (five vaccinates and three con trols) shall be used as test animals. Throat and nasal swabs shall be col lected from each cat and individually tested on susceptible cell cultures for the presence of feline calicivirus. Blood samples shall be drawn and individual serum samples tested for neutralizing antibody. The cats shall be considered suitable for use if all swabs are nega tive for virus isolation and all serums are negative for calicivirus antibody at the 1:2 final dilution in a 50 percent plaque reduction test or other test of equal sensitivity. (ii) The five cats used as vaccinates shall be administered one dose of vac cine by the method recommended on the label. If two doses are rec ommended, the second dose shall be given after the interval recommended on the label. (iii) Fourteen or more days after the final dose of vaccine, the vaccinates and controls shall each be challenged intranasally with virulent feline calicivirus furnished or approved by Animal and Plant Health Inspection Service and observed each day for 14 days postchallenge. The rectal tem perature of each animal shall be taken and the presence or absence of clinical signs, particularly lesions on the oral mucosa, noted and recorded each day. (iv) If three of three controls do not show clinical signs of feline calicivirus infection other than fever, the test is inconclusive and may be repeated. (v) If a significant difference in clin ical signs cannot be demonstrated be tween vaccinates and controls using a scoring system approved by Animal and Plant Health Inspection Service and prescribed in the Outline of Pro duction, the serial is unsatisfactory. [50 FR 433, Jan. 4, 1985. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66784, 66786, Dec. 26, 1991] § 113.211 [Reserved] § 113.212 Bursal Disease Vaccine, Killed Virus. Bursal Disease Vaccine, Killed Virus, shall be prepared from virus-bearing cell culture fluids or embryonated chicken eggs. Only Master Seed which has been established as pure, safe, and immunogenic shall be used for pre paring seeds for vaccine production. All serials shall be prepared from the first through the fifth passage from the Master Seed. (a) The Master Seed shall meet the applicable requirements prescribed in § 113.200. (b) Each lot of Master Seed shall be tested for pathogens by the chicken embryo inoculation test prescribed in § 113.37, except that, if the test is a No Test because of a vaccine virus over ride, the chicken inoculation test pre scribed in § 113.36 may be conducted and the virus judged accordingly. (c) The immunogenicity of vaccine prepared in accordance with the Out line of Production shall be established by a method acceptable to Animal and Plant Health Inspection Service. Vac cine used for this test shall be at the highest passage from the Master Seed and prepared at the minimum preinactivation titer specified in the Outline of Production. The test shall establish that the vaccine, when used as recommended on the label, is capa ble of inducing an immune response in dams of sufficient magnitude to pro vide significant protection to offspring. (d) Test requirements for release. Each serial and subserial shall meet the ap plicable general requirements pre scribed in § 113.200 and the special re quirements in this paragraph. Any se rial or subserial found unsatisfactory by a prescribed test shall not be re leased. (1) Safety. Vaccinates used in the po tency test in paragraph (d)(2) of this section shall be observed each day dur ing the prechallenge period. If unfavor able reactions attributable to the vac cine occur, the serial is unsatisfactory. If unfavorable reactions which are not attributable to the vaccine occur, the test is a No Test and may be repeated. If the test is not repeated, the serial is unsatisfactory. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00775 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 766 9 CFR Ch. I (1–1–18 Edition) §§ 113.213–113.214 (2) Potency. Bulk or final container samples of completed product from each serial shall be tested for potency using the two-stage potency test pro vided in this paragraph. (i) Vaccinates. Inject each of 21 sus ceptible chickens 14 to 28 days of age, properly identified and obtained from the same source and hatch, with one dose of vaccine by the route rec ommended on the label and observe for at least 21 days. (ii) Controls. Retain at least 10 addi tional chickens from the same source and hatch as unvaccinated controls. (iii) Challenge. Twenty-one to 28 days postvaccination, challenge 20 vac cinates and 10 controls by eyedrop with a virulent infectious bursal disease virus furnished or approved by Animal and Plant Health Inspection Service. (iv) Postchallenge period. Four days postchallenge, necropsy all chickens and examine each for gross lesions of bursal disease. For purposes of this test, gross lesions shall include peribursal edema and/or edema and/or macroscopic hemorrhage in the bursal tissue. Vaccinated chickens showing gross lesions shall be counted as fail ures. If at least 80 percent of the con trols do not have gross lesions of bursal disease in a stage of the test, that stage is considered inconclusive and may be repeated. In a valid test, the re sults shall be evaluated according to the following table: Stage Number of vac cinates Cumu lative number of vac cinates Cumulative total number of failures for— Satisfactory serial Unsatisfac tory serial 1 ................................................................................................................. 20 20 3 or less ...... 6 or more. 2 ................................................................................................................. 20 40 8 or less ...... 9 or more. (v) If four or five vaccinates show le sions of bursal disease in the first stage, the second stage may be con ducted in a manner identical to the first stage. If the second stage is not conducted, the serial is unsatisfactory. (vi) If the second stage is used, each serial shall be evaluated according to the second part of the table on the basis of cumulative results. [50 FR 434, Jan. 4, 1985. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66784, 66786, Dec. 26, 1991; 79 FR 55969, Sept. 18, 2014] §§ 113.213–113.214 [Reserved] § 113.215 Bovine Virus Diarrhea Vac cine, Killed Virus. Bovine Virus Diarrhea Vaccine, Killed Virus, shall be prepared from virus-bearing cell culture fluids. Only Master Seed virus which has been es tablished as pure, safe, and immunogenic shall be used for pre paring seed cultures for vaccine pro duction. All serials of vaccine shall be prepared from the first through the fifth passage from the Master Seed. (a) The Master Seed shall meet the applicable general requirements pre scribed in § 113.200 and the require ments of this section. (b) The immunogenicity of vaccine prepared from the Master Seed in ac cordance with the Outline of Produc tion shall be established by a method acceptable to the Animal and Plant Health Inspection Service. Vaccine used for this test shall be at the high est passage from the Master Seed and at the minimum preinactivation titer provided in the Outline of Production. (c) Test requirements for release. Each serial and subserial shall meet the ap plicable general requirements pre scribed in § 113.200 and the special re quirements provided in this paragraph. Any serial or subserial found unsatis factory by a prescribed test shall not be released. (1) Safety. Vaccinates used in the po tency test in paragraph (c)(2) of this section shall be observed each day dur ing the prechallenge period. If unfavor able reactions occur, including res piratory signs, which are attributable to the vaccine, the serial is unsatisfac tory. If unfavorable reactions occur which are not attributable to the vac cine, the test is a No Test and may be VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00776 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 767 Animal and Plant Health Inspection Service, USDA § 113.216 repeated one time. If results of the sec ond test are not satisfactory, or if the test is not repeated, the serial is unsat isfactory. (2) Potency. Bulk or final container samples of completed product shall be tested for potency using the method described in this paragraph. (i) Eight bovine virus diarrhea sus ceptible calves (five vaccinates and three controls) shall be used as test animals. Individual serum samples shall be collected, inactivated, and in dividually tested for neutralizing anti body. (ii) A constant virus decreasing serum neutralization test in cell cul ture using 50–300 TCID50 of virus shall be used. Calves shall be considered sus ceptible if there is no neutralization at 1:2 final serum dilution. Other tests of equal sensitivity approved by the Ani mal and Plant Health Inspection Serv ice may be used. (iii) The five calves used as vac cinates shall be administered one dose of vaccine as recommended on the label. If two doses are recommended, the second dose shall be given accord ing to the interval recommended on the label. (iv) Fourteen days or more after the last vaccination, blood samples shall be drawn and the individual serum samples inactivated and tested for bo vine virus diarrhea virus neutralizing antibody by the same method used to determine susceptibility. (v) Test interpretation. If the controls have not remained seronegative at 1:2, the test is a No Test (NT) and may be repeated. If at least four of the five vaccinates in a valid test have not de veloped 50 percent endpoint titers of 1:8 or greater, the serial is unsatisfactory, except as provided in paragraph (c)(2)(vi) of this section. (vi) Virus Challenge Test. If the results of a valid serum neutralization test are unsatisfactory, the vaccinates and con trols may be challenged with virulent bovine virus diarrhea virus furnished or approved by the Animal and Plant Health Inspection Service. The animals shall be observed for 14 days post-chal lenge. If two of the three control calves do not show a temperature rise to 104.5 °F and develop respiratory or clinical signs of bovine virus diarrhea, the test is a No Test and may be repeated one time. If two or more vaccinates show a temperature of 104.0 °F for 2 or more days and develop respiratory or clinical or other signs, the serial is unsatisfac tory. (vii) The prevaccination and postvaccination sera from a satisfac tory potency test shall be submitted to the National Veterinary Services Lab oratories for confirmatory testing. [55 FR 35562, Aug. 31, 1990] § 113.216 Bovine Rhinotracheitis Vac cine, Killed Virus. Infectious Bovine Rhinotracheitis Vaccine, Killed Virus, shall be prepared from virus-bearing cell culture fluids. Only Master Seed virus which has been established as pure, safe, and immunogenic shall be used for pre paring seed cultures for vaccine pro duction. All serials of vaccine shall be prepared from the first through the fifth passage from the Master Seed. (a) The Master Seed shall meet the applicable general requirements pre scribed in § 113.200 and the require ments of this section. (b) The immunogenicity of vaccine prepared in accordance with the Out line of Production shall be established by a method acceptable to the Animal and Plant Health Inspection Service. Vaccine used for this test shall be at the highest passage from the Master Seed and at the minimum preinactivation titer provided in the Outline of Production. (c) Test requirements for release. Each serial and subserial shall meet the re quirements prescribed in § 113.200 and the special requirements provided in this paragraph. Any serial or subserial found unsatisfactory by a prescribed test shall not be released. (1) Safety. Vaccinates used in the po tency test in paragraph (c)(2) of this section shall be observed each day dur ing the prechallenge period. If unfavor able reactions occur, which are attrib utable to the vaccine, the serial is un satisfactory. If unfavorable reactions occur which are not attributable to the vaccine, the test is a No Test and may be repeated one time. If the results of the second test are not satisfactory, or if the test is not repeated, the serial is unsatisfactory. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00777 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 768 9 CFR Ch. I (1–1–18 Edition) § 113.300 (2) Potency. Bulk or final container samples of completed product shall be tested for potency using the method described in this paragraph. (i) Eight infectious bovine rhinotracheitis susceptible calves (five vaccinates, three controls) shall be used as test animals. Individual serum samples shall be collected, inactivated, and individually tested for neutralizing antibody. (ii) A constant virus decreasing serum neutralization test in cell cul ture using 50–300 TCID50 of virus shall be used. Calves shall be considered sus ceptible if there is no neutralization at 1:2 final serum dilution. Other tests of equal sensitivity acceptable to the Ani mal and Plant Health Inspection Serv ice may be used. (iii) The five calves used as vac cinates shall be administered one dose of vaccine as recommended on the label. If two doses are recommended, the second dose shall be given accord ing to the interval recommended on the label. (iv) Fourteen or more days after the last vaccination, blood samples shall be drawn and the individual serum samples inactivated and tested for in fectious bovine rhinotracheitis virus neutralizing antibody by the same method used to determine suscepti bility. (v) Test interpretation. If the three controls have not remained seronegative at 1:2, the test is a No Test (NT) and may be repeated. If at least four of the five vaccinates in a valid test have not developed 50 per cent endpoint titers of 1:8, the serial is unsatisfactory, except as provided in paragraph (c)(2)(vi) of this section. (vi) Virus Challenge Test. If the results of a valid serum neutralization test are unsatisfactory, the vaccinates and con trols may be challenged with virulent infectious bovine rhinotracheitis virus furnished or approved by the Animal and Plant Health Inspection Service. The animals shall be observed each day for 14 days post-challenge. If two of the three control calves do not show a tem perature rise to 104.5 °F and develop respiratory or other clinical signs of infectious bovine rhinotracheitis, the test is a No Test (NT) and may be re peated one time. If more than one of the vaccinates shows a temperature of 104.0 °F for 2 or more days or if more than one of the vaccinates develops respiratory or clinical or other signs, the serial is unsatisfactory. (vii) The prevaccination and postvaccination sera from a satisfac tory potency test shall be submitted to the National Veterinary Services Lab oratories for testing by the Animal and Plant Health Inspection Service. [55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66786, Dec. 26, 1991] LIVE VIRUS VACCINES § 113.300 General requirements for live virus vaccines. When prescribed in an applicable Standard Requirement or in the filed Outline of Production, a live virus vac cine shall meet the applicable require ments in this section. (a) Purity tests—(1) Bacteria and fungi. Final container samples of completed product and comparable samples of each lot of Master Seed Virus shall be tested for bacteria and fungi in accord ance with the test provided in § 113.27. (2) Mycoplasma. Final container sam ples of completed product and com parable samples of each lot of Master Seed Virus shall be tested for myco plasma in accordance with the test pro vided in § 113.28. (3) Avian Origin Vaccine. Samples of each lot of Master Seed Virus and bulk pooled material or final container sam ples from each serial shall also be test ed for: (i) Salmonella contamination as pre scribed in § 113.30; and (ii) Lymphoid leukosis virus con tamination as prescribed in § 113.31; and (iii) Hemagglutinating viruses as pre scribed in § 113.34. (4) Extraneous viruses. Each lot of Master Seed Virus used to prepare live virus vaccine recommended for animals other than poultry shall meet the re quirements for extraneous viruses as prescribed in § 113.55 (b) Safety tests. Samples of each lot of Master Seed Virus and final container samples of completed product from each serial or first subserial of live virus vaccine recommended for animals other than poultry shall be tested for safety in at least one species for which VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00778 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 769 Animal and Plant Health Inspection Service, USDA § 113.301 the vaccine is intended using methods prescribed in §§ 113.39, 113.40, 113.41, 113.44, and 113.45 or in a filed Outline of Production. The mouse safety test pre scribed in § 113.33(a) shall also be con ducted unless the virus or agent in the vaccine is inherently lethal for mice. (c) Virus identity test. At least one of the virus identity tests provided in this paragraph or a suitable identity test prescribed in the filed Outline of Pro duction shall be conducted on the Mas ter Seed Virus and final container sam ples from each serial or first subserial of biological product. (1) Fluorescent antibody test. The fluo rescent antibody test shall be con ducted using virus inoculated cells and uninoculated control cells. Cells shall be stained with fluorochrome con jugated specific antiserum. Fluores cence typical of the virus concerned shall be demonstrated in the inocu lated cells. The control cells shall re main free of such fluorescence. (2) Serum neutralization test. The serum neutralization test shall be con ducted using the constant serum-de creasing virus method with specific antiserum. For positive identification, at least 100 ID50 of vaccine virus shall be neutralized by the antiserum. (d) Cell Culture Requirements. If cell cultures are used in the preparation of Master Seed Virus or of the vaccine, primary cells shall meet the require ments prescribed in § 113.51, cell lines shall meet the requirements prescribed in § 113.52, and ingredients of animal or igin shall meet the applicable require ments in § 113.53. (e) Moisture content. (1) The max imum moisture content in desiccated vaccines must be stated in the filed Outline of Production. (2) Final container samples of com pleted product from each serial or sub serial must be tested for moisture con tent in accordance with the test pre scribed in § 113.29. [39 FR 27430, July 29, 1974, as amended at 43 FR 49528, Oct. 24, 1978; 50 FR 1042, Jan. 9, 1985; 54 FR 19352, May 5, 1989. Redesignated at 55 FR 35562, Aug. 31, 1990; 60 FR 24549, May 9, 1995; 68 FR 57608, Oct. 6, 2003] § 113.301 Ovine Ecthyma Vaccine. Ovine Ecthyma Vaccine shall be pre pared from tissue culture fluids or virus-bearing tissues obtained from sheep that have developed ovine ecthyma following inoculation with virulent ovine ecthyma virus. Ovine Ecthyma Vaccine is exempt from the requirements prescribed in §§ 113.27 and 113.300(a), (b), and (c). Each serial shall meet the moisture requirements in § 113.300(e) and the special requirements prescribed in this section. Any serial found unsatisfactory by a prescribed test shall not be released. (a) Safety tests. (1) Bulk or final con tainer samples of completed product from each serial shall be tested for safety as prescribed in § 113.38. (2) The prechallenge period of the po tency test shall constitute a safety test. If unfavorable reactions attrib utable to the vaccine occur in either of the vaccinates during the observation period, the serial is unsatisfactory. (b) Potency test. Final container sam ples of completed product from each se rial and each subserial shall be tested for potency using susceptible lambs. The vaccine shall be prepared as rec ommended for use on the label. (1) Each of two lambs (vaccinates) shall be vaccinated by application of the vaccine to a scarified area on the medial surface of the thigh and ob served each day for 14 days. (2) The immunity of the two vac cinates and one or more unvaccinated lambs (controls) shall be challenged in the same manner as for vaccination, using the opposite thigh. (3) If typical signs of ovine ecthyma, such as hyperemia, vesicles, and pustules do not develop on the controls during the first 2 weeks following chal lenge and persist for approximately 30 days, the test is a No Test and may be repeated. (4) If the vaccinates do not show a typical immune reaction, the serial is unsatisfactory: Provided, That, an ini tial active reaction with hyperemia which resolves progressively and dis appears within 2 weeks, may be charac terized as a typical immune reaction. [39 FR 27430, July 29, 1974. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66786, Dec. 26, 1991] VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00779 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 770 9 CFR Ch. I (1–1–18 Edition) § 113.302 § 113.302 Distemper Vaccine—Mink. Distemper Vaccine—Mink shall be prepared from virus-bearing cell cul ture fluids. Only Master Seed Virus which has been established as pure, safe, and immunogenic shall be used for preparing the production seed virus for vaccine production. All serials of vaccine shall be prepared from the first through the fifth passage from the Master Seed Virus. (a) The Master Seed Virus shall meet the applicable requirements prescribed in § 113.300 and the requirements pre scribed in this section. (b) The lot of Master Seed Virus shall be tested for extraneous viruses as fol lows: (1) To detect virulent canine dis temper virus, each of two distemper susceptible mink or ferrets shall be in oculated with 1 ml of the Master Seed Virus and observed each day for 21 days. If undesirable reactions occur in either test animal, the lot of Master Seed Virus is unsatisfactory. (2) Master Seed Virus propagated in chicken embryos shall be tested for pathogens by the chicken embryo test prescribed in § 113.37 except lesions typ ical of distemper virus may be dis regarded. If found unsatisfactory, the Master Seed Virus shall not be used. (c) Each lot of Master Seed Virus used for vaccine production shall be tested for immunogenicity. The se lected virus dose from the lot of Master Seed Virus shall be established as fol lows: (1) At least 25 distemper susceptible mink shall be used as test animals. Blood samples shall be drawn from these animals and individual serum samples tested. The mink shall be con sidered susceptible if the results are negative at a 1:2 final serum dilution in a varying serum-constant virus neu tralization test with less than 500 ID50 of canine distemper virus. Other means of insuring susceptibility may be used if prior approval from Animal and Plant Health Inspection Service is re ceived. (2) A geometric mean titer of the dried vaccine produced from the high est passage of the Master Seed Virus shall be established before the immunogenicity test is conducted. At least 20 mink shall be vaccinated with a predetermined quantity of vaccine virus and at least 5 additional mink shall be held as unvaccinated controls. To confirm the dosage calculations, five replicate virus titrations shall be conducted on a sample of the vaccine virus dilution used. (3) At least twenty-one days post-in jection, the immunity of each of the vaccinates and the controls shall be challenged with the same size dose of virulent distemper virus and observed each day for 21 days. (i) If at least 80 percent of the con trols do not die or show severe signs of distemper, the test is a No Test and may be repeated. (ii) If at least 19 of 20, 27 of 30, or 36 of 40 of the vaccinates do not survive without showing clinical signs of dis temper during the observation period, the Master Seed Virus is unsatisfac tory. (4) An Outline of Production change shall be made before authority for use of a new lot of Master Seed Virus shall be authorized by Animal and Plant Health Inspection Service. (d) Test requirements for release: Each serial and subserial shall meet the general requirements prescribed in § 113.300 and the requirements in this paragraph. Final container samples of completed product shall be tested. Any serial or subserial found unsatisfactory by a prescribed test shall not be re leased. (1) Mink safety test. Each of 2 mink shall be vaccinated with the equivalent of 10 doses of vaccine rehydrated with sterile diluent and administered in the manner recommended on the label. The mink shall be observed each day for 21 days. If unfavorable reactions attrib utable to the product occur in either of the mink during the observation pe riod, the serial or subserial is unsatis factory. If unfavorable reactions which are not attributable to the product occur, the test shall be declared a No Test and may be repeated: Provided, That if the test is not repeated, the se rial or subserial shall be declared un satisfactory. (2) Potency Test. An in vitro potency test shall be conducted. To be eligible for release, each serial and subserial shall have a virus titer sufficiently greater than the titer of vaccine virus VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00780 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 771 Animal and Plant Health Inspection Service, USDA § 113.303 used in the immunogenicity test pre scribed in paragraph (c) of this section to assure that, when tested at any time within the expiration period, each se rial and subserial shall have a virus titer 10 0.7 greater than that used in such immunogenicity test when tested by the method used in paragraph (c)(2) of this section. [40 FR 53000, Nov. 14, 1975, as amended at 48 FR 33471, July 22, 1983. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66784, 66786, Dec. 26, 1991; 72 FR 72564, Dec. 21, 2007] § 113.303 Bluetongue Vaccine. Bluetongue Vaccine shall be prepared from virus-bearing cell culture fluids. Only Master Seed which has been es tablished as pure, safe, and immunogenic shall be used for pre paring the seeds for vaccine produc tion. All serials of vaccine shall be pre pared from the first through the tenth passage from the Master Seed. (a) The Master Seed shall meet the applicable general requirements pre scribed in § 113.300 and the require ments in this section. (b) Each lot of Master Seed shall be tested for transmissibility and rever sion to virulence in sheep using a method acceptable to Animal and Plant Health Inspection Service. If re version to virulence is demonstrated, the Master Seed is unsatisfactory. (c) Each lot of Master Seed used for vaccine production shall be tested for immunogenicity. The selected virus dose from the lot of Master Seed shall be established as follows: (1) Twenty-five lambs, susceptible to the bluetongue virus serotype con tained in the vaccine, shall be used as test animals (20 vaccinates and 5 con trols). Blood samples shall be drawn from these animals and individual se rums tested. A lamb shall be consid ered susceptible if there is no neutral ization at a 1:2 final serum dilution in a constant virus varying serum neu tralization test with 60 to 300 TCID50 of bluetongue virus or another method ac ceptable to Animal and Plant Health Inspection Service. (2) A geometric mean titer of the vac cine produced from the highest passage from the Master Seed shall be estab lished before the immunogenicity test is conducted. The 20 lambs to be used as vaccinates shall be administered a predetermined quantity of vaccine virus by the method recommended on the label. To confirm the virus dosage administered, five replicate virus titra tions shall be conducted on a sample of the vaccine used. (3) At least once during the period of 14 to 18 days postvaccination, indi vidual serum samples shall be collected from each of the vaccinates and tested for virus neutralizing antibody using the 60 to 300 TCID50 of bluetongue virus. (4) Twenty-one to twenty-eight days postvaccination the vaccinates and the controls shall each be challenged with virulent bluetongue virus and observed for 14 days. The rectal temperature of each animal shall be taken and re corded for 17 consecutive days begin ning 3 days prechallenge. The presence or absence of lesions or other clinical signs of bluetongue noted and recorded on each of 14 consecutive days postchallenge. (i) If at least four of the five controls do not show clinical signs of bluetongue and a temperature rise of 3 °F or higher over the prechallenge mean temperature, the test shall be considered a No Test and may be re peated. (ii) If at least 19 of the 20 vaccinates tested as prescribed in paragraph (c)(3) of this section do not have bluetongue neutralizing antibody titers of 1:4 final serum dilution or higher, or if more than one of the vaccinates shows a temperature rise of 3 °F or higher than its prechallenge mean temperature for 2 or more days, or if more than one of the vaccinates exhibits clinical signs of bluetongue, the Master Seed is unsatis factory. (5) An Outline of Production change shall be made before authority for use of a new lot of Master Seed shall be granted by Animal and Plant Health Inspection Service. (d) Test requirements for release. Each serial and subserial shall meet the ap plicable general requirements pre scribed in § 113.300 and the require ments in this paragraph. Final con tainer samples of completed product shall be tested. Any serial or subserial VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00781 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 772 9 CFR Ch. I (1–1–18 Edition) § 113.304 found unsatisfactory by a prescribed test shall not be released. (1) Safety test. The mouse safety test prescribed in § 113.33(a) and the lamb safety test prescribed in § 113.45 shall be conducted. (2) Virus titer requirements. Final con tainer samples of completed product shall be tested for virus titer using the titration method used in paragraph (c)(2) of this section. To be eligible for release, each serial and subserial shall have a virus titer sufficiently greater than the titer of vaccine virus used in the immunogenicity test prescribed in paragraph (c) of this section to assure that when tested at any time within the expiration period, each serial and subserial shall have a virus titer of 10 0.7 greater than that used in such immunogenicity test. [50 FR 23796, June 6, 1985. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66784, 66786, Dec. 26, 1991; 72 FR 72564, Dec. 21, 2007] § 113.304 Feline Panleukopenia Vac cine. Feline Panleukopenia Vaccine shall be prepared from virus-bearing cell cul ture fluids. Only Master Seed Virus which has been established as pure, safe, and immunogenic shall be used for preparing the production seed virus for vaccine production. All serials of vaccine shall be prepared from the first through the fifth passage from the Master Seed Virus. (a) The Master Seed Virus shall meet the applicable general requirements prescribed in § 113.300 and the require ments prescribed in this section. (b) The lot of Master Seed Virus shall be tested for other agents as follows: (1) To detect virulent feline panleukopenia virus or virulent mink enteritis virus, each of two feline panleukopenia susceptible cats, as de termined by the criteria prescribed in paragraph (c)(1) of this section, shall be injected subcutaneously with the equivalent of one cat dose each and the cats observed each day for 21 days. If either or both cats show signs of dis ease or reduced white blood cell counts below 50 percent of the normal level es tablished by an average of three or more counts taken prior to injection, the Master Seed Virus is unsatisfac tory. (2) To detect chlamydial agents, the Master Seed Virus shall be tested as prescribed in § 113.43. (c) Each lot of Master Seed Virus used for vaccine production shall be tested for immunogenicity. The se lected virus dose from the lot of Master Seed Virus shall be established as fol lows: (1) Twenty-five feline panleukopenia susceptible cats shall be used as test animals (20 vaccinates and 5 controls). Blood samples drawn from each cat shall be individually tested for neutral izing antibody against feline panleukopenia virus to determine sus ceptibility. (i) A constant virus-carrying serum neutralization test in tissue culture using 100 to 300 TCID50 of virus shall be used. (ii) Cats shall be considered suscep tible if there is no neutralization at a 1:2 final serum dilution. (2) A geometric mean titer of the dried vaccine produced from the high est passage of the Master Seed Virus shall be established before the immunogenicity test is conducted. The 20 cats used as vaccinates shall be in jected with a predetermined quantity of vaccine virus and the remaining five cats held as uninjected controls. To confirm the dosage calculations, five replicate virus titrations shall be con ducted on a sample of the vaccine virus dilution used. (3) Fourteen days post-injection, the vaccinates and the controls shall be challenged with virulent feline panleukopenia virus furnished by Ani mal and Plant Health Inspection Serv ice and the cats observed each day for 14 days. (i) If at least 80 percent of the con trols do not show clinical signs of fe line panleukopenia during the observa tion period, the test is a No Test and may be repeated. Clinical signs of fe line panleukopenia shall include a pro nounced leukopenia wherein the white cell count drops to 4,000 or less per cubic mm, or the white cell count drops to less than 25 percent of the nor mal level established by an average of three or more counts taken prior to challenge. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00782 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 773 Animal and Plant Health Inspection Service, USDA § 113.305 (ii) If at least 19 of the 20 vaccinates do not survive the observation period without showing clinical signs of feline panleukopenia as described in para graph (c)(3)(i) of this section, the Mas ter Seed Virus is unsatisfactory. (4) An Outline of Production change shall be made before authority for use of a new lot of Master Seed Virus shall be granted by Animal and Plant Health Inspection Service. (d) Test requirements for release. Each serial and subserial shall meet the re quirements prescribed in § 113.300 and in this paragraph. Final container sam ples of completed product shall be test ed. Any serial or subserial found unsat isfactory by a prescribed test shall not be released. (1) Safety test. The mouse safety test prescribed in § 113.33(a) and the cat safety test prescribed in § 113.39 shall be conducted. (i) Each of two healthy cats shall be injected with 10 cat doses by the meth od recommended on the label and the cats observed each day for 14 days. (ii) If unfavorable reactions attrib utable to the biological product occur during the observation period, the se rial is unsatisfactory. If unfavorable reactions occur which are not attrib utable to the product, the test shall be declared a No Test and repeated: Pro vided, That, if not repeated, the serial shall be unsatisfactory. (2) Virus titer requirements. Final con tainer samples of completed product shall be tested for virus titer using the titration method used in paragraph (c)(2) of this section. To be eligible for release, each serial and each subserial shall have a virus titer sufficiently greater than the titer of vaccine virus used in the immunogenicity test pre scribed in paragraph (c) of this section to assure that when tested at any time within the expiration period, each se rial and subserial shall have a virus titer of 10 0.7 greater than that used in such immunogenicity test but not less than 10 2.5 TCID50 per dose. [39 FR 44716, Dec. 27, 1974, as amended at 40 FR 53378, Nov. 18, 1975; 43 FR 25078, June 9, 1978; 43 FR 41186, Sept. 15, 1978; 44 FR 58900, Oct. 12, 1979; 48 FR 33471, July 22, 1983. Redes ignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66784, 66786, Dec. 26, 1991; 72 FR 72564, Dec. 21, 2007] § 113.305 Canine Hepatitis and Canine Adenovirus Type 2 Vaccine. Canine Hepatitis Vaccine and Canine Adenovirus Type 2 Vaccine shall be prepared from virus-bearing cell cul ture fluids. Only Master Seed Virus which has been established as pure, safe, and immunogenic shall be used in preparing the production seed virus for vaccine production. All serials shall be prepared from the first through the fifth passage from the Master Seed Virus. (a) The Master Seed Virus shall meet the applicable requirements prescribed in § 113.300 except that the dog safety test prescribed in § 113.40(a) shall be conducted by the intravenous route. (b) Each lot of Master Seed Virus used for vaccine production shall be tested for immunogenicity by one or both of the following methods: (1) Immunogenicity for canine hepatitis. Twenty-five canine hepatitis suscep tible dogs shall be used as test animals (20 vaccinates and 5 controls). Blood samples shall be drawn from these ani mals and individual serum samples tested. The dogs shall be considered susceptible if the results are negative at a 1:2 final serum dilution in a vary ing serum-constant virus neutraliza tion test using 50 to 300 TCID50 of ca nine adenovirus. (i) A geometric mean titer of the dried vaccine produced from the high est passage of the Master Seed Virus shall be established before the immunogenicity test is conducted. The 20 dogs to be used as vaccinates shall be injected with a predetermined quan tity of vaccine virus and the remaining five dogs held as uninjected controls. To confirm the dosage calculations, five replicate virus titrations shall be conducted on a sample of the vaccine virus dilution used. (ii) Not less than 14 days postinjection, the vaccinates and the controls shall each be challenged intra venously with virulent infectious ca nine hepatitis virus furnished or ap proved by the Animal and Plant Health Inspection Service and observed each day for 14 days. (A) If at least four of the five con trols do not show severe clinical signs of canine hepatitis, the test is a No Test and may be repeated. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00783 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 774 9 CFR Ch. I (1–1–18 Edition) § 113.306 (B) If at least 19 of the 20 vaccinates do not survive without showing clinical signs of infectious canine hepatitis dur ing the observation period, the Master Seed Virus is unsatisfactory. (2) Immunogenicity for canine adenovirus Type 2. Thirty canine adenovirus type 2 susceptible dogs shall be used as test animals (20 vac cinates and 10 controls). Blood samples shall be drawn from these animals and individual serum samples tested. The dogs shall be considered susceptible if the results are negative at a 1:2 final serum dilution in a varying serum-con stant virus neutralization test using 50 to 300 TCID50 of canine adenovirus. (i) A geometric mean titer of the dried vaccine produced from the high est passage of the Master Seed Virus shall be established before the immunogenicity test is conducted. The 20 dogs to be used as vaccinates shall be injected with a predetermined quan tity of vaccine virus and the remaining 10 dogs held as uninjected controls. To confirm the dosage calculations, five replicate virus titrations shall be con ducted on a sample of the vaccine virus dilution used. (ii) Not less than 14 days postinjection, the vaccinates and the controls shall be challenged by expo sure to a nebulized aerosol of virulent canine adenovirus type 2 furnished or approved by the Animal and Plant Health Inspection Service and observed each day for 14 days postchallenge. The rectal temperature of each animal shall be taken and the presence of res piratory or other clinical signs of ca nine adenovirus type 2 noted and re corded each day. (A) If at least 6 of 10 controls do not show clinical signs of canine adenovirus type 2 infection other than fever, the test is a No Test and may be repeated. (B) If a significant difference in clin ical signs in a valid test cannot be demonstrated between vaccinates and controls using a scoring system ap proved by the Animal and Plant Health Inspection Service, the Master Seed Virus is unsatisfactory. (iii) An Outline of Production change shall be made before authorization for use of a new lot of Master Seed Virus shall be granted by the Animal and Plant Health Inspection Service. (c) Test requirements for release. Each serial and subserial shall meet the re quirements prescribed in § 113.300 and in this paragraph. Final container sam ples of completed product shall be test ed. Any serial or subserial found unsat isfactory by a prescribed test shall not be released. (1) Virus titer requirements. Final con tainer samples of completed product shall be tested for virus titer using the titration method used in paragraph (b)(1)(i) and/or (b)(2)(i) of this section. To be eligible for release, each serial and each subserial shall have a virus titer sufficiently greater than the titer of vaccine virus used in the immunogenicity test(s) prescribed in paragraph (b) of this section to assure that when tested at any time within the expiration period, each serial and subserial shall have a virus titer of 10 0.7 greater than that used in such immunogenicity test(s) but not less than 10 2.5 TCID50 dose. If both immunogenicity tests in paragraph (b) of this section are conducted and a dif ferent amount of virus is used in each test, the virus titer requirements shall be based on the higher of the two amounts. (2) [Reserved] [60 FR 14361, Mar. 17, 1995, as amended at 72 FR 72564, Dec. 21, 2007] § 113.306 Canine Distemper Vaccine. Canine Distemper Vaccine shall be prepared from virus-bearing cell cul ture fluids or embryonated chicken eggs. Only Master Seed Virus which has been established as pure, safe, and immunogenic shall be used for pre paring the production seed virus for vaccine production. All serials of vac cine shall be prepared from the first through the fifth passage from the Master Seed Virus. (a) Master Seed Virus. The Master Seed Virus shall meet the applicable requirements prescribed in § 113.300 and the requirements prescribed in this sec tion. (1) To detect ferret virulent canine distemper virus, each of five canine distemper susceptible ferrets shall be injected with a sample of the Master Seed Virus equivalent to the amount of VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00784 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 775 Animal and Plant Health Inspection Service, USDA § 113.308 virus to be used in one dog dose and ob served each day for 21 days. If undesir able reactions are observed during the observation period, the lot of Master Seed is unsatisfactory. (2) Master Seed Virus propagated in tissues or cells of avian origin shall be tested for pathogens by the chicken embryo test prescribed in § 113.37. If found unsatisfactory, the Master Seed Virus shall not be used. (b) Each lot of Master Seed Virus used for vaccine production shall be tested for immunogenicity. The se lected virus dose from the lot of Master Seed Virus shall be established as fol lows: (1) Twenty-five canine distemper sus ceptible dogs shall be used as test ani mals (20 vaccinates and 5 controls). Blood samples shall be drawn from these animals and individual serum samples tested. The dogs shall be con sidered susceptible if the results are negative at a 1:2 final serum dilution in a varying serum-constant virus neu tralization test using 50 to 300 TCID50 of canine distemper virus. (2) A geometric mean titer of the dried vaccine produced from the high est passage of the Master Seed Virus shall be established before the immunogenicity test is conducted. The 20 dogs used as vaccinates shall be in jected with a predetermined quantity of vaccine virus and the remaining five dogs held as uninjected controls. To confirm the dosage calculations, five replicate virus titrations shall be con ducted on a sample of the vaccine virus dilution used. (3) At least 14 days post-injection, the vaccinates and the controls shall each be challenged intracerebrally with virulent canine distemper virus furnished or approved by the Animal and Plant Health Inspection Service and observed each day for 21 days. (i) If at least four of the five controls do not die and the survivor, if any, does not show clinical signs of canine dis temper the test is a No Test and may be repeated. (ii) If at least 19 of the 20 vaccinates do not survive without showing clinical signs of canine distemper during the observation period, the Master Seed Virus is unsatisfactory. (4) An Outline of Production change shall be made before authorization for use of a new lot of Master Seed Virus shall be granted by the Animal and Plant Health Inspection Service. (c) Test requirements for release. Ex cept for § 113.300(a)(3)(ii), each serial and subserial shall meet the require ments prescribed in § 113.300 and in this paragraph. Final container samples of completed product shall be tested. Any serial or subserial found unsatisfactory by a prescribed test shall not be re leased. (1) The test for pathogens prescribed in § 113.37 shall be conducted on each serial or one subserial of avian origin vaccine. (2) Virus titer requirements. Final con tainer samples of completed product shall be tested for virus titer using the titration method used in paragraph (b)(2) of this section. To be eligible for release, each serial and subserial shall have a virus titer sufficiently greater than the titer of vaccine virus used in the immunogenicity test prescribed in paragraph (b) of this section to assure that when tested at any time within the expiration period, each serial and subserial shall have a virus titer of 10 0.7 greater than that used in such immunogenicity test but not less than 10 2.5 TCID50 per dose. [60 FR 14362, Mar. 17, 1995, as amended at 72 FR 72564, Dec. 21, 2007] § 113.308 Encephalomyelitis Vaccine, Venezuelan. Encephalomyelitis Vaccine, Ven ezuelan, shall be prepared from virus bearing cell culture fluids. Only Master Seed which has been established as pure, safe, and immunogenic shall be used for preparing seeds for vaccine production. All serials of vaccine shall be prepared from the first through the fifth passage from the Master Seed. (a) The Master Seed shall meet the applicable general requirements pre scribed in § 113.300 except (b), and the requirements prescribed in this sec tion. (b) Each lot of Master Seed shall be tested for immunogenicity. The se lected virus dose from the lot of Master Seed shall be established as follows: VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00785 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 776 9 CFR Ch. I (1–1–18 Edition) § 113.309 (1) Tests conducted by the Depart ment have established that horses hav ing Venezuelan equine encephalomyelitis antibody titers of 1:20 by the hemagglutination-inhibi tion (HI) method or 1:40 by the serum neutralization (SN) method were im mune to challenge with virulent virus. The immunogenicity test is based on the demonstration of a serological re sponse of at least that magnitude fol lowing vaccination of serologically negative horses. (2) At least 22 horses (20 vaccinates and 2 controls), susceptible to Ven ezuelan equine encephalomyelitis, shall be used as test animals. Blood samples shall be taken from each horse and the serums individually tested for neutralizing antibody. Horses shall be considered susceptible if there is no neutralization at a 1:2 final serum dilu tion in a constant virus-varying serum neutralization test using 60 to 300 TCID50 of Venezuelan equine encephalomyelitis virus. (3) A geometric mean titer of the vac cine produced from the highest passage of the Master Seed shall be established using a method acceptable to Veteri nary Services before the immunogenicity test is conducted. The 20 horses used as vaccinates shall be in jected with a predetermined quantity of vaccine virus by the method to be recommended on the label. To confirm the dosage administered, five replicate virus titrations shall be conducted on a sample of the vaccine virus dilution used. (4) Twenty-one to twenty-eight days postvaccination, blood samples shall be drawn from all test animals. For a valid test, the controls shall remain seronegative at 1:2 final serum dilu tion. In a valid test, if at least 19 of 20 vaccinates do not have antibody titers of at least 1:20 in a hemagglutination inhibition test or at least 1:40 in a serum neutralization test, the Master Seed is unsatisfactory. (5) An Outline of Production change shall be made before authority for use of a new lot of Master Seed shall be granted by Animal and Plant Health Inspection Service. (c) Test requirements for release. Each serial and subserial shall meet the ap plicable general requirements pre scribed in § 113.300 and special require ments in this paragraph. Any serial or subserial found unsatisfactory by a prescribed test shall not be released. (1) Safety test. The mouse safety test prescribed in § 113.33(b) shall be con ducted. (2) Virus titer requirements. Final con tainer samples of completed product shall be tested for virus titer using the method in paragraph (b)(3) of this sec tion. To be eligible for release, each se rial and subserial shall have a virus titer sufficiently greater than the titer of the vaccine used in the immunogenicity test prescribed in paragraph (b) of this section to assure that, when tested at any time within the expiration period, each serial and subserial shall have a virus titer of 10 0.7 greater than that used in the immunogenicity test, but not less than 10 2.5 TCID50 per dose. [50 FR 23797, June 6, 1985. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66784, 66786, Dec. 26, 1991; 72 FR 72564, Dec. 21, 2007] § 113.309 Bovine Parainfluenza 3 Vac cine. Bovine Parainfluenza3 Vaccine shall be produced from virus-bearing cell culture fluids. Only Master Seed Virus which has been established as pure, safe, and immunogenic shall be used for preparing the production seed virus for vaccine production. All serials of vaccine shall be prepared from the first through the tenth passage from the Master Seed Virus. (a) The Master Seed Virus shall meet the applicable general requirements prescribed in § 113.300. (b) Each lot of Master Seed Virus shall meet the special requirements prescribed in this section. (c) Each lot of Master Seed Virus used for vaccine production shall be tested for immunogenicity. The se lected virus dose from the lot of Master Seed Virus shall be established as fol lows: (1) Twenty-five bovine parainfluenza, susceptible calves shall be used as test animals (20 vaccinates and five con trols). Blood samples shall be drawn from these animals and individual se rums tested. Also, nasal specimens VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00786 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 777 Animal and Plant Health Inspection Service, USDA § 113.309 shall be collected for virus isolation at tempts. The calves shall be considered susceptible if: (i) The results are negative at a 1:2 final serum dilution in a varying serum constant virus neutralization test with less than 500 TCID50 of bovine parainfluenza3 virus; and (ii) Shall be negative to bovine parainfluenza3 virus isolation attempts from the nasal specimens on the day of injection. (2) A geometric mean titer of the dried vaccine produced from the high est passage of the Master Seed Virus shall be established before the immunogenicity test is conducted. The 20 calves to be used as vaccinates shall be injected with a predetermined quan tity of vaccine virus and the remaining five calves held as uninjected controls. To confirm the dosage calculation, five replicate virus titrations shall be con ducted on a sample of the vaccine virus dilution used. (3) The vaccinates and controls shall be examined for clinical signs of res piratory disease and the body tempera ture taken and recorded on each of the first 14 consecutive days post-injection. The vaccinates shall be bled on day 6 ±2 days post-injection. (4) Three to four weeks post-vaccina tion, all calves shall be bled for serum antibodies and nasal specimens shall be collected for PI3 virus isolation. On the same day, all vaccinates and controls shall be given acceptable challenge PI3 virus titrating at least 10 7.0 TCID50 per ml and the animals observed for 14 days. Two ml of the challenge virus shall be instilled in each nostril or shall be inhaled as an aerosol suspen sion. Upon request, challenge virus and instructions shall be furnished by Ani mal and Plant Health Inspection Serv ice. (5) Each animal shall be examined for clinical signs of respiratory disease and the body temperature recorded on each of the 14 consecutive days of the post challenge observation period. Each day for at least the first 10 days post-chal lenge, nasal specimens for virus isola tion attempts shall be taken. All ani mals shall be bled on day 6 ±2 days post-challenge, and all animals shall be bled at least once 14 to 28 days post challenge for serum antibody studies. (6) Satisfactory Test Criteria: (i) All virus isolations attempts shall be by culture and at least one subcul ture in PI3 susceptible cells for a total of at least 14 days. (ii) Two to four weeks post-vaccina tion, at least 19 of the 20 vaccinates shall have PI3 neutralizing antibody titers of at least 1:4 and all five con trols shall be negative at 1:2 dilution. None of the post-vaccination serums collected from the vaccinates on day 6 ±2 days shall reveal serum neutraliza tion antibody titers of 1:32 or greater based upon final dilution. (iii) Satisfactory resistance to chal lenge by vaccinates shall be deter mined by a significant difference be tween virus isolation rates from vac cinates and controls. The virus neu tralization titers of post-challenge se rums and respiratory symptoms and temperatures from all animals shall be considered in the evaluation of the test validity. (7) Designated animal alternates for test animals showing anamnestic anti body responses (titers 1:32 or greater) on day 6 serums may be included in the study under the following provisions: (i) No more than five alternates shall be allowed for the vaccinates and no more than two for the controls. (ii) Alternates shall be subject to all requirements outlined for the animals for which they are alternates. (iii) Antibody values from alternate animals may be used only to replace values from up to and including five vaccinates which develop antibody of 1:32 or greater by day 6 ±2 days post vaccination or up to and including two controls which develop antibody titers of 1:32 or greater by day 6 ±2 days post challenge. (8) A sequential test procedure may be used in lieu of the 20 calf require ment. A beta value of .05 and a toler ance level of .78 shall be required. (9) An Outline of Production change shall be made before authority for use of a new lot of Master Seed Virus shall be granted by Animal and Plant Health Inspection Service. (d) Test requirements for release: Each serial and subserial shall meet the applicable general requirements VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00787 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 778 9 CFR Ch. I (1–1–18 Edition) § 113.310 prescribed in § 113.300 and the require ments in this paragraph. Final con tainer samples of completed product shall be tested except as prescribed in paragraph (d)(1) of this section. Any se rial or subserial found unsatisfactory by a prescribed test shall not be re leased. (1) Purity test. The test for Brucella contamination prescribed in § 113.32 shall be conducted on each batch of pri mary cells intended for production use. (2) Safety test. The mouse safety test prescribed in § 113.33(a) and the calf safety test prescribed in § 113.41 shall be conducted. (3) Virus titer requirements. Final con tainer samples of completed product shall be tested for virus titer using the titration method used in paragraph (c)(2) of this section. To be eligible for release, each serial and each subserial shall have a virus titer per dose suffi ciently greater than the titer of vac cine virus used in the immunogenicity test prescribed in paragraph (c) of this section to assure that when tested at any time within the expiration period, each serial and subserial shall have a virus titer of 10 0.7 greater than that used in the immunogenicity test but not less than 10 2.5 TCID50 per dose. [39 FR 44719, Dec. 27, 1974, as amended at 40 FR 41089, Sept. 5, 1975; 43 FR 49529, Oct. 24, 1978; 48 FR 33472, July 22, 1983. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66784, 66786, Dec. 26, 1991; 60 FR 14357, Mar. 17, 1995; 72 FR 72564, Dec. 21, 2007] § 113.310 Bovine Rhinotracheitis Vac cine. Bovine Rhinotracheitis Vaccine shall be prepared from virus-bearing cell cul ture fluids. Only Master Seed Virus which has been established as pure, safe, and immunogenic shall be used for preparing the production seed virus for vaccine production. All serials of vaccine shall be prepared from the first through the tenth passage from the Master Seed Virus. (a) The Master Seed Virus shall meet the applicable general requirements prescribed in § 113.300. (b) Each lot of Master Seed Virus shall meet the special requirements prescribed in this section. (c) Each lot of Master Seed Virus used for vaccine production shall be tested for immunogenicity. The se lected virus dose from the lot of Master Seed Virus shall be established as fol lows: (1) Twenty-five infectious bovine rhinotracheitis susceptible calves shall be used as test animals (20 vaccinates and five controls). Blood samples shall be drawn from these animals and indi vidual serums tested. The calves shall be considered susceptible if the results are negative at a 1:2 final serum dilu tion by the virus plaque reduction method. (2) A geometric mean titer of the dried vaccine produced from the high est passage of the Master Seed Virus shall be established before the immunogenicity test is conducted. The 20 calves to be used as vaccinates shall be injected with a predetermined quan tity of vaccine virus and the remaining five calves held as uninjected controls. To confirm the dosage calculations, five replicate virus titrations shall be conducted on a sample of the vaccine virus dilution used. (3) At least once during a period of 14 to 28 days post-vaccination, individual serum samples shall be collected for virus-neutralization tests from each of the vaccinates. The test virus shall be 100 to 500 TCID50 bovine rhinotracheitis virus. Results shall be used in making a determination as prescribed in para graph (c)(6) of this section. (4) The vaccinates and the controls shall each be challenged with virulent infectious bovine rhinotracheitis virus and observed for 14 days. The rectal temperature of each animal shall be taken and the presence or absence of respiratory or other clinical signs of bovine rhinotracheitis noted and re corded on each of the 14 consecutive days. (5) If at least four of the five controls do not show clinical signs of infectious bovine rhinotracheitis and a marked temperature rise to 104.5 °F. or higher post-challenge, the test shall be consid ered a No Test and may be repeated. (6) If less than 19 of the post-injection serum samples tested as prescribed in paragraph (c)(3) of this section show neutralization in all tubes of the 1:2 final serum dilution, or if more than one of the vaccinates show a tempera ture of 103.5 °F. or higher for 2 or more VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00788 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 779 Animal and Plant Health Inspection Service, USDA § 113.311 days, or if more than one of the vac cinates exhibit respiratory or other clinical signs of infectious bovine rhinotracheitis, or both, the Master Seed Virus is unsatisfactory. (7) A sequential test procedure may be used in lieu of the 20 calf require ment. A beta value of .05 and a toler ance level of .78 shall be required. (8) An outline of Production change shall be made before authority for use of a new lot of Master Seed Virus shall be granted by Animal and Plant Health Inspection Service. (d) Test requirements for release: Each serial and subserial shall meet the applicable general requirements prescribed in § 113.300 and the require ments in this paragraph. Final con tainer samples of completed product shall be tested except as prescribed in paragraph (d)(1) of this section. Any se rial or subserial found unsatisfactory by a prescribed test shall not be re leased. (1) Purity test. The test for Brucella contamination prescribed in § 113.32 shall be conducted on each batch of pri mary cells intended for production use. (2) Safety test. The mouse safety test prescribed in § 113.33(a) and the calf safety test prescribed in § 113.41 shall be conducted. (3) Virus titer requirements. Final con tainer samples of completed product shall be tested for virus titer using the titration method used in paragraph (c)(2) of this section. To be eligible for release, each serial and each subserial shall have a virus titer per dose suffi ciently greater than the titer of vac cine virus used in the immunogenicity test prescribed in paragraph (c) of this section to assure that when tested at any time within the expiration period, each serial and subserial shall have a virus titer of 10 0.7 greater than that used in the immunogenicity test but not less than 10 2.5 TCID50 per dose. [39 FR 44720, Dec. 27, 1974, as amended at 40 FR 20067, May 8, 1975; 40 FR 23989, June 4, 1975; 40 FR 41089, Sept. 5, 1975; 43 FR 49529, Oct. 24, 1978; 48 FR 33472, July 22, 1983. Redes ignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66784, 66786, Dec. 26, 1991; 72 FR 72564, Dec. 21, 2007] § 113.311 Bovine Virus Diarrhea Vac cine. Bovine Virus Diarrhea Vaccine shall be prepared from virus-bearing cell cul ture fluids. Only Master Seed Virus which has been established as pure, safe, and immunogenic shall be used for preparing the production seed virus for vaccine production. All serials of vaccine shall be prepared from the first through the tenth passage from the Master Seed Virus. (a) The Master Seed Virus shall meet the applicable general requirements prescribed in § 113.300. (b) Each lot of Master Seed Virus shall meet the special requirements prescribed in this section. (c) Each lot of Master Seed Virus used for vaccine production shall be tested for immunogenicity. The se lected virus dose from the lot of Master Seed Virus shall be established as fol lows: (1) Twenty-five bovine virus diarrhea susceptible calves shall be used as test animals (20 vaccinates and five con trols). Blood samples shall be drawn from these animals and individuals serum samples tested. The calves shall be considered susceptible to bovine virus diarrhea virus infection if the re sults are negative at a 1:2 final serum dilution in a varying serum-constant virus neutralization test with less than 500 TCID50 of bovine virus diarrhea virus. (2) A geometric mean titer of the dried vaccine produced from the high est passage of the Master Seed Virus shall be established before the immunogenicity test is conducted. The 20 calves to be used as vaccinates shall be injected with a predetermined quan tity of vaccine virus and the remaining five calves held as uninjected controls. To confirm the dosage calculations, five replicate virus titrations shall be conducted on a sample of the vaccine virus dilution used. (3) At least once during a period 14 to 28 days post-vaccination, individual serum samples shall be collected for virus-neutralization tests from each of the vaccinates. The test virus shall be less than 500 TCID50 of bovine virus di arrhea virus. The white cell count for all vaccinates and controls shall be es tablished at least 3 days just before VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00789 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 780 9 CFR Ch. I (1–1–18 Edition) § 113.312 challenge. Results shall be used in making a determination as prescribed in paragraph (c)(5) of this section. (4) The vaccinates and the controls shall each be challenged with virulent bovine virus diarrhea virus and ob served for 14 consecutive days. The white cell count shall be determined daily on each animal from the second through the eighth day post-challenge. If leukopenia does not develop in at least four of the five controls as com pared with the vaccinates, the test shall be considered a No Test and may be repeated. (5) If less than 19 of the post-injection serum samples, tested as prescribed in paragraph (c)(3) of this section, show neutralization in all tubes of the 1:8 di lution; or if more than one of the vac cinates exhibits respiratory or other clinical signs of bovine virus diarrhea post-challenge; or both, the Master Seed Virus is unsatisfactory. (6) A sequential test procedure may be used in lieu of the 20 calf require ment. A beta value of .05 and a toler ance level of .78 shall be required. (7) An Outline of Production change shall be made before authority for use of a new lot of Master Seed Virus shall be granted by Animal and Plant Health Inspection Service. (d) Test requirements for release: Each serial and subserial shall meet the applicable general requirements prescribed in § 113.300 and the require ments in this paragraph. Final con tainer samples of completed product shall be tested except as prescribed in paragraph (d)(1) of this section. Any se rial or subserial found unsatisfactory by a prescribed test shall not be re leased. (1) Purity test. The test for Brucella contamination prescribed in § 113.32 shall be conducted on each batch of pri mary cells intended for production use. (2) Safety test. The mouse safety test prescribed in § 113.33(a) and the calf safety test prescribed in § 113.41 shall be conducted. (3) Virus titer requirements. Final con tainer samples of completed product shall be tested for virus titer using the titration method used in paragraph (c)(2) of this section. To be eligible for release, each serial and each subserial shall have a virus titer per dose suffi ciently greater than the titer of vac cine virus used in the immunogenicity test prescribed in paragraph (c) of this section to assure that when tested at any time within the expiration period, each serial and subserial shall have virus titer of 10 0.7 greater than that used in the immunogenicity test but not less than 10 2.5 TCID 50 per dose. [39 FR 44721, Dec. 27, 1974, as amended at 40 FR 20067, May 8, 1975; 40 FR 41089, Sept. 5, 1975; 43 FR 49529, Oct. 24, 1978; 48 FR 33472, July 22, 1983. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66784, 66786, Dec. 26, 1991; 72 FR 72564, Dec. 21, 2007] § 113.312 Rabies Vaccine, Live Virus. Rabies Vaccine shall be prepared from virus-bearing cell cultures or embryonated chicken eggs. Only Mas ter Seed Virus which has been estab lished as pure, safe and immunogenic shall be used for preparing the produc tion seed virus for vaccine production. All serials of vaccine shall be prepared from the first through the fifth passage from the Master Seed Virus. (a) The Master Seed Virus shall meet the applicable general requirements prescribed in § 113.300. (1) Each lot of Master Seed Virus shall meet the special requirements prescribed in this section. (2) Each lot of Master Seed Virus propagated in tissues or cells of avian origin shall be tested for pathogens by procedures prescribed in § 113.37. (3) Each lot of Master Seed Virus propagated in primary cell cultures of mouse or hamster origin or brain tis sues of mouse origin shall be tested for lymphocytic choriomeningitis (LCM) virus by the procedure prescribed in § 113.42. If LCM virus is detected, the Master Seed Virus is unsatisfactory. (4) The Master Seed Virus shall be studied in each species of carnivore or domesticated wild animal for which the vaccine is specifically rec ommended to attempt to determine the fate of the vaccine virus. Results shall be considered in evaluating safety of vaccine virus. (i) Obtain at least 10 unvaccinated animals, negative at 1:2 final serum di lution, of each species in which tests will be conducted. Divide each species into two groups of five animals. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00790 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 781 Animal and Plant Health Inspection Service, USDA § 113.312 (ii) For each species of animal, inject one group of five animals intramuscularly. Infiltrate a major nerve and the surrounding tissue in each of the five animals in the other group. Use 1.0 ml of high titer virus for each method of administration. (iii) Observe all animals for signs of rabies until scheduled time to sac rifice. If animals show definite symp toms, sacrifice and check regional lymph nodes, brain, salivary glands, and kidney for rabies virus by injection of suckling mice (not more than 7 days of age). Tissues may be held frozen at ¥70 °C. until suckling mice are avail able. Inject each mouse in one litter intracerebrally with 0.02 ml of a ground tissue suspension from each organ. Ob serve mice each day for 21 days. If any mice die, determine if the deaths were due to rabies virus in the brain by a fluorescent antibody test. (iv) Sacrifice animals that do not show signs of rabies according to the following schedule and check regional lymph nodes, brain, salivary glands, and kidney in suckling mice. Route of injection Days after injection Number of animals Intramuscularly ............. 15, 20, 25, 30, 35 1 each day. Intraneurally ................. 3, 6, 9, 15, 30 1 each day. (5) Each lot of Master Seed Virus shall be tested for safety in at least 10 unvaccinated serologically negative animals of each domestic species for which the vaccine is recommended. (i) Each group of 10 animals shall be divided into 2 groups of 5 animals. For each species, inject one group intramuscularly with 10 doses of high titer virus. (ii) Infiltrate a major nerve of each of the animals in the other group of 5 with 10 doses of the same high titer virus. For all species except dogs and cats, multiple injections along the cer vical spine in the proximity to the nerve trunks emerging from the spinal cord may be used: Provided, That a 1 dose volume shall be injected into each of four or more sites bilaterally. (iii) Observe all animals each day for 90 days. (iv) If any animals show clinical signs of rabies, sacrifice the animal and check appropriate brain tissue for ra bies virus by the fluorescent antibody test and by mouse injection. (v) If rabies is confirmed, the lot of Master Seed Virus is unsatisfactory. (b) The immunogenicity of vaccine prepared with virus at the highest pas sage of the Master Seed shall be estab lished in each species for which the vaccine is recommended. Tests shall be conducted in accordance with a pro tocol filed with Animal and Plant Health Inspection Service before initi ation of the tests. The vaccine shall be prepared using methods prescribed in the Outline of Production. If Rabies Vaccine is to be in combination with other fractions, the product tested shall include all fractions to be rec ommended. (1) A geometric mean virus titer of the dried vaccine produced from the highest passage of the Master Seed Virus shall be established before the immunogenicity test is conducted. To confirm the dosage calculations, five replicate virus titrations shall be con ducted on a sample of the vaccine virus dilution used. (2) The dose of vaccine to be used in the immunogenicity test shall be no more than the amount of rehydrated vaccine which, on the basis of previous titrations, has been diluted to the pro posed minimum acceptable virus titer. (3) Test animals shall be uniform and have no neutralizing antibodies to ra bies as determined by serum-neutral ization (SN) tests. (i) Twenty-five or more animals shall be used as vaccinates. Each shall be in jected intramuscularly at one site in the thigh with a dose of vaccine at the proposed minimum virus titer as speci fied in the filed Outline of Production. (ii) Ten or more additional animals shall be held as controls. (iii) On or about days 30, 90, 180, 270, and 365 postvaccination, all animals shall be bled and individual serums tested for neutralizing antibodies to ra bies virus. (iv) All surviving test animals of each species shall be challenged intramuscularly with virulent rabies virus furnished or approved by Animal and Plant Health Inspection Service 1 year after vaccination, except as pro vided in paragraphs (b)(4), (b)(5), and (b)(6) of this section. The challenged VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00791 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 782 9 CFR Ch. I (1–1–18 Edition) § 113.312 animals shall be observed each day for 90 days as prescribed in § 113.5(b). The brain of each test animal that dies fol lowing challenge shall be examined for rabies by the fluorescent antibody test or other method acceptable to Animal and Plant Health Inspection Service. (v) Requirements for acceptance in challenge tests shall be death due to rabies in at least 80 percent of controls while at least 22 of 25 or 26 of 30 or a statistically equivalent number of the vaccinates remain well for a period of 90 days. (4) An alternative to challenging all surviving test animals in accordance with paragraph (b)(3)(iv) of this section may be used when the test animals are of species other than carnivores. Vac cinates shall be challenged at 1 year postvaccination. These shall include five vaccinates with the lowest SN titers at the 270th-day bleeding, five vaccinates with the lowest SN titers at the 365th-day bleeding, and all vac cinates with SN titers below 1:10 by the mouse SN test or below 1:16 by the rapid-fluorescent-focus-inhibition test at any bleeding. At least five SN-nega tive controls of each species shall be challenged at the same time as the vaccinates. All SN titers shall be iterated to an endpoint. All of the chal lenged vaccinates must remain well for a period of 90 days, and at least 80 per cent of the controls must die of rabies for a satisfactory test without further challenge. If one or more of the vac cinates die from rabies, all the remain ing vaccinates, regardless of titer, along with the five controls shall be challenged. The cumulative results from the two challenges shall be evalu ated for acceptance as specified in paragraph (b)(3)(v) of this section. (5) An outline of Production change shall be made before authority for use of a new lot of Master Virus shall be granted by Animal and Plant Health Inspection Service. (c) If more than 1 year duration of immunity is to be claimed, a duration of immunity test for the additional time shall be conducted and inter preted as prescribed in paragraph (b) of this section for the 1 year test. The test animals shall be monitored sero logically at least every 180 days. The time of challenge may be adjusted ac cordingly. (d) Test requirements for release: Each serial and each subserial shall meet the general requirements pre scribed in § 113.300 and special require ments in this paragraph. (1) Purity and safety tests. Final con tainer samples of completed product from each serial or one subserial shall be tested. (i) The test for pathogens, prescribed in § 113.37 shall be conducted on each serial or one subserial of avian origin. If necessary, neutralize the rabies virus with specific rabies antiserum. (ii) A test for safety in three young seronegative animals of the most sus ceptible species for which the vaccine is recommended shall be conducted. Each shall be injected intramuscularly with 10 recommended doses of vaccine. If unfavorable reactions attributable to the product occur during a 28 day ob servation period, the serial is unsatis factory. (iii) If primary cell cultures of ham ster origin or of mouse origin are used vaccine production, they shall be test ed for LCM virus as prescribed in § 113.42. The cells shall be disrupted and undiluted cell fluids from each lot shall be tested. (2) Virus titrations. Final container samples of completed product shall be tested for virus titer using the titra tion method used in paragraph (b)(1) of this section. To be eligible for release, each serial and each subserial shall have a virus titer sufficiently higher than the titer of the vaccine virus used in paragraph (b) of this section to as sure that, when tested at any time within the expiration period, each se rial and subserial shall have a virus titer equal to or greater than that used in the immunogenicity test. (3) Young adult mice, each weighing 14 to 16 grams, shall be used as test ani mals when the virus in vaccine pre pared with a low egg passage Flury Strain or high cell passage Street Ala bama Dufferin Strain (HCP SAD) of ra bies virus is titrated. At least 10 mice for each dilution shall be used. (i) At least 10 mice shall be used for each dilution. Each shall be injected intracerebrally with 0.03 ml. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00792 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 783 Animal and Plant Health Inspection Service, USDA § 113.313 (ii) The injected young adult mice shall be observed each day for 14 days except when testing vaccines made with HCP SAD strain of rabies virus, in which case, the mice shall be observed each day for 21 days. Deaths and paral ysis occurring subsequent to the fourth day post-injection shall be noted and the LD50 titer calculated by the Reed and Muench Method. (iii) Virus titer requirements for re lease and at expiration date shall be determined for each vaccine on the basis of data available: Provided, That, the lowest titer permitted at expira tion date when determined by this test shall be 10 3.0 LD50 per 0.03 ml. (4) Suckling mice, 6 days of age or younger, shall be used as test animals when virus in vaccine prepared with a high egg passage Flury Strain of rabies virus is titrated. (i) Six to twelve mice shall be used for each dilution. Each shall be in jected intracerebrally with 0.02 ml. (ii) The injected suckling mice shall be observed each day for 21 days. Deaths and paralysis occurring subse quent to the fourth day post-injection shall be noted and the LD50 titer cal culated by the Reed and Muench Meth od; and (iii) Virus titer requirements for re lease and at expiration date shall be determined for each vaccine on the basis of data available: Provided, That, the lowest titer permitted at expira tion date when determined by this test shall be 10 3.0 LD50 per 0.02 ml. [39 FR 44721, Dec. 27, 1974, as amended at 40 FR 20067, May 8, 1975; 42 FR 6795, Feb. 4, 1977; 43 FR 49529, Oct. 24, 1978; 50 FR 20090, May 14, 1985; 50 FR 23797, June 6, 1985. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66784, 66786, Dec. 26, 1991; 61 FR 31823, June 21, 1996; 72 FR 72564, Dec. 21, 2007] § 113.313 Measles Vaccine. Measles Vaccine shall be prepared from virus-bearing cell culture fluids. Only Master Seed Virus which has been established as pure, safe, and immunogenic shall be used for pre paring the production seed virus for vaccine production. All serials of vac cine shall be prepared from the first through the fifth passage from the Master Seed Virus. (a) The Master Seed Virus shall meet the applicable general requirements prescribed in § 113.300. Each lot of Mas ter Seed Virus shall meet the special requirements prescribed in this sec tion. (b) To detect virulent canine dis temper virus, each of two canine dis temper susceptible ferrets shall be in jected with a sample of the Master Seed Virus equivalent to the amount of virus to be used in one dog dose and ob served each day for 21 days. If undesir able reactions occur in either ferret, the lot of Master Seed Virus is unsatis factory. (c) Each lot of Master Seed Virus used for vaccine production shall be tested for immunogenicity. The se lected virus dose from the lot of Master Seed Virus shall be established as fol lows: (1) Twenty-five dogs, less than 12 weeks of age and free of measles anti body, shall be used as test animals (20 vaccinates and five controls). Blood samples shall be drawn from these ani mals and individual serum samples tested. The dogs shall be considered susceptible if the results are negative at a 1:2 final serum dilution in a vary ing serum-constant virus neutraliza tion test with less than 500 ID50 of mea sles virus. (2) A geometric mean titer of the dried vaccine produced from the high est passage of the Master Seed Virus shall be established before the immunogenicity test is conducted. Twenty dogs shall be vaccinated with a predetermined quantity of vaccine virus and the remaining five dogs held as unvaccinated controls. To confirm the dosage calculations, five replicate virus titrations shall be conducted on a sample of the vaccine virus dilution used. (3) On the day of challenge, serum samples shall be obtained from each vaccinate and individually tested for antibody against canine distemper virus. For a valid test, each vaccinate shall be negative at a 1:4 final serum dilution in varying serum-constant virus neutralization test using less than 500 ID50 of canine distemper virus. (4) At least 21 days postinoculation, the immunity of the vaccinates and VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00793 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 784 9 CFR Ch. I (1–1–18 Edition) § 113.314 controls shall be challenged by expo sure to a uniform dose of aerosolized virulent canine distemper virus. All test dogs shall be observed daily for 21 days postchallenge. (i) If at least 4 of the 5 controls do not die or show signs of distemper, in cluding a temperature of 104.0 °F. or higher and at least 15 percent weight loss, the test is a No Test and may be repeated. (ii) If at least 19 of the 20 vaccinates do not survive without showing a tem perature of 104.0 °F. or higher and a weight loss exceeding 15 percent after day 8 postchallenge, the Master Seed Virus is unsatisfactory. (5) When approved in advance by Ani mal and Plant Health Inspection Serv ice, a sequential test procedure may be used in lieu of the 20 dog requirement. A beta value of 0.05 and a tolerance level of 0.78 shall be required. (6) An Outline of Production change shall be made before authority for use of a new lot of Master Seed Virus shall be granted by Animal and Plant Health Inspection Service. (d) Test requirements for release: Each serial and subserial shall meet the general requirements prescribed in § 113.300 and the requirements in this paragraph. Final container samples of completed product shall be tested. Any serial or subserial found unsatisfactory by a prescribed test shall not be re leased. (1) Safety tests. The dog safety test prescribed in § 113.40 and the mouse safety test prescribed in § 113.33(a) shall be conducted. (2) Virus titer requirements. Final con tainer samples of completed product shall be tested for virus titer using the titration method used in paragraph (c)(2) of this section. To be eligible for release, each serial and each subserial shall have a virus titer sufficiently greater than the titer of the vaccine virus used in the immunogenicity test prescribed in paragraph (c) of this sec tion to assure that when tested at any time within the expiration period, each serial and subserial shall have a virus titer of 10 0.7 greater than that used in the immunogenicity test but not less than 10 2.5 ID50 per dose. [40 FR 53001, Nov. 14, 1975, as amended at 43 FR 49529, Oct. 24, 1978; 48 FR 33472, July 22, 1983. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66784, 66786, Dec. 26, 1991; 72 FR 72564, Dec. 21, 2007] § 113.314 Feline Calicivirus Vaccine. Feline Calicivirus Vaccine shall be prepared from virus-bearing cell cul ture fluids. Only Master Seed Virus which has been established as pure, safe, and immunogenic shall be used for preparing the production seed virus for vaccine production. All serials of vaccine shall be prepared from the first through the fifth passage from the Master Seed Virus. (a) The Master Seed Virus shall meet the applicable general requirements prescribed in § 113.300. (b) The Master Seed Virus shall be tested for chlamydial agents as pre scribed in § 113.43. (c) Each lot of Master Seed Virus used for vaccine production shall be tested for immunogenicity. The se lected virus dose from the lot of Master Seed Virus shall be established as fol lows: (1) Thirty feline calicivirus suscep tible cats shall be used as test animals (20 vaccinates and 10 controls). Throat swabs shall be collected from each cat and individually tested on susceptible cell cultures for the presence of feline calicivirus. Blood samples shall be drawn and individual serum samples tested. The cats shall be considered suitable for use if all swabs are nega tive for virus isolation and if all se rums are negative for calicivirus anti body at the 1:2 final dilution in a 50 percent plaque reduction test or other SN test of equal sensitivity. (2) A geometric mean titer of the dried vaccine produced from the high est passage of the Master Seed Virus shall be established before the immunogenicity test is conducted. The 20 cats used as vaccinates shall be ad ministered a predetermined quantity of vaccine virus by the method to be rec ommended on the label and the re maining 10 cats shall be held as con trols. To confirm the dosage calcula tions, five replicate virus titrations shall be conducted on a sample of the VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00794 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 785 Animal and Plant Health Inspection Service, USDA § 113.315 vaccine virus dilution used. If two doses are used, five replicate con firming titrations shall be conducted on each dose. (3) Twenty-one or more days after the final dose of vaccine, the vac cinates and controls shall each be chal lenged intranasally with a minimum of 100,000 TCID50 or plaque forming units of virulent feline calicivirus furnished or approved by Animal and Plant Health Inspection Service and observed each day for 14 days postchallenge. The rectal temperature of each animal shall be taken and the presence or ab sence of clinical signs, particularly le sions on the oral mucosa, noted and re corded each day. (i) If less than 8 of 10 controls show clinical signs of feline calicivirus infec tion other than fever, the test is a No Test and may be repeated. (ii) If a significant difference in clin ical signs cannot be demonstrated be tween vaccinates and controls using a scoring system approved by Animal and Plant Health Inspection Service and prescribed in the Outline of Pro duction, the Master Seed Virus is un satisfactory. (4) An Outline of Production change shall be made before authority for use of a new lot of Master Seed Virus shall be granted by Animal and Plant Health Inspection Service. (d) Test requirements for release. Each serial and subserial shall meet the re quirements prescribed in § 113.300 and in this paragraph. Final container sam ples of completed product shall be test ed. Any serial or subserial found unsat isfactory by a prescribed test shall not be released. (1) Safety test. The mouse safety test prescribed in § 113.33(a) and the cat safety test prescribed in § 113.39(b) shall be conducted. (2) Virus titer requirements. Final con tainer samples of completed product shall be tested for virus titer using the titration method used in paragraph (c)(2) of this section. To be eligible for release, each serial and each subserial shall have a virus titer sufficiently greater than the titer of vaccine virus used in the immunogenicity test pre scribed in paragraph (c) of this section to assure that when tested at any time within the expiration period, each se rial and subserial shall have a virus titer of 10 0.7 greater than that used in the immunogenicity test but not less than 10 2.5 TCID50 or plaque forming units per dose. [44 FR 58899, Oct. 12, 1979; 44 FR 63083, Nov. 2, 1979, as amended at 48 FR 33472, July 22, 1983. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66784, 66786, Dec. 26, 1991; 72 FR 72564, Dec. 21, 2007] § 113.315 Feline Rhinotracheitis Vac cine. Feline Rhinotracheitis Vaccine shall be prepared from virus-bearing cell cul ture fluids. Only Master Seed Virus which has been established as pure, safe, and immunogenic shall be used for preparing the production seed virus for vaccine production. All serials of vaccine shall be prepared from the first through the fifth passage from the Master Seed Virus. (a) The Master Seed Virus shall meet the applicable general requirements prescribed in § 113.300. (b) The Master Seed Virus shall be tested for chlamydial agents as pre scribed in § 113.43. (c) Each lot of Master Seed Virus used for vaccine production shall be tested for immunogenicity. The se lected virus dose from the lot of Master Seed Virus shall be established as fol lows: (1) Thirty feline rhinotracheitis sus ceptible cats shall be used as test ani mals (20 vaccinates and 10 controls). Throat swabs shall be collected from each cat and individually tested on sus ceptible cell cultures for the presence of feline rhinotracheitis virus. Blood samples shall be drawn and individual serum samples tested. The cats shall be considered suitable for use if all swabs are negative for virus isolation and if all serums are negative for feline rhinotracheitis virus antibody at the 1:2 final dilution in a 50 percent plaque reduction test or other SN test of equal sensitivity. (2) A geometric mean titer of the dried vaccine produced from the high est passage of the Master Seed Virus shall be established before the immunogenicity test is conducted. The 20 cats used as vaccinates shall be ad ministered a predetermined quantity of VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00795 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 786 9 CFR Ch. I (1–1–18 Edition) § 113.316 vaccine virus by the method to be rec ommended on the label and the re maining 10 cats shall be held as con trols. To confirm the dosage calcula tions, five replicate virus titrations shall be conducted on a sample of the vaccine virus dilution used. If two doses are used, five replicate con firming titrations shall be conducted on each dose. (3) Twenty-one or more days after the final dose of vaccine, the vac cinates and controls shall each be chal lenged intranasally with a minimum of 100,000 TCID50 or plaque forming units of virulent feline rhinotracheitis virus furnished or approved by Animal and Plant Health Inspection Service and observed each day for 14 days post challenge. The rectal temperature of each animal shall be taken and the presence of respiratory or other clin ical signs of feline rhinotracheitis noted and recorded each day. (i) If less than 8 of 10 controls show clinical signs of feline rhinotracheitis infection other than fever, the test is a No Test and may be repeated. (ii) If a significant difference in clin ical signs cannot be demonstrated be tween vaccinates and controls using a scoring system approved by Veterinary Services and prescribed in the Outline of Production, the Master Seed Virus is unsatisfactory. (4) An Outline of Production change shall be made before authority for use of a new lot of Master Seed Virus shall be granted by Animal and Plant Health Inspection Service. (d) Test requirements for release. Each serial and subserial shall meet the re quirements prescribed in § 113.300 and in this paragraph. Final container sam ples of completed product shall be test ed. Any serial or subserial found unsat isfactory by a prescribed test shall not be released. (1) Safety test. The mouse safety test prescribed in § 113.33(a) and the cat safety test prescribed in § 113.39(b) shall be conducted. (2) Virus titer requirements. Final con tainer samples of completed product shall be tested for virus titer using the titration method used in paragraph (c)(2) of this section. To be eligible for release, each serial and each subserial shall have a virus titer sufficiently greater than the titer of vaccine virus used in the immunogenicity test pre scribed in paragraph (c) of this section to assure that when tested at any time within the expiration period, each se rial and subserial shall have a virus titer of 10 0.7 greater than that used in the immunogenicity test but not less than 10 2.5 TCID50 or plaque forming units per dose. [44 FR 58899, Oct. 12, 1979, as amended at 48 FR 33472, July 22, 1983. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66784, 66786, Dec. 26, 1991; 72 FR 72564, Dec. 21, 2007] § 113.316 Canine Parainfluenza Vac cine. Canine Parainfluenza Vaccine shall be prepared from virus-bearing cell cul ture fluids. Only Master Seed which has been established as pure, safe, and immunogenic shall be used for pre paring seeds for vaccine production. All serials of vaccine shall be prepared from the first through the fifth passage from the Master Seed. (a) The Master Seed shall meet the applicable general requirements pre scribed in § 113.300 and the require ments in this section. (b) Each lot of Master Seed shall be tested for immunogenicity. The se lected virus dose shall be established as follows: (1) Twenty-five canine parainfluenza susceptible dogs (20 vaccinates and 5 controls) shall be used as test animals. Nasal swabs shall be collected from each dog on the day the first dose of vaccine is administered and individ ually tested on susceptible cell cul tures for the presence of canine parainfluenza virus. Blood samples shall also be drawn and individual serum samples tested for neutralizing antibody. Dogs shall be considered sus ceptible if all swabs are negative for virus isolation and if all serums are negative for canine parainfluenza anti body at a 1:2 final dilution in a con stant virus-varying serum neutraliza tion test using 50 to 300 TCID50 of ca nine parainfluenza virus. (2) A geometric mean titer of vaccine produced at the highest passage from the Master Seed shall be established before the immunogenicity test is con ducted. The 20 dogs used as vaccinates VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00796 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 787 Animal and Plant Health Inspection Service, USDA § 113.317 shall be administered a predetermined quantity of vaccine virus. Five rep licate virus titrations shall be con ducted on a sample of the vaccine virus dilution used to confirm the dosage ad ministered. If two doses are used, five replicate confirming titrations shall be conducted on each dose. (3) Three to 4 weeks after the final dose of vaccine, all dogs shall be bled for serum antibodies and nasal swabs shall be collected for canine parainfluenza virus isolation. On the same day, all vaccinates and controls shall be challenged with canine parainfluenza virus furnished or ap proved by Animal and Plant Health In spection Service. (4) The rectal temperature of each dog shall be taken and the presence of respiratory or other clinical signs of canine parainfluenza virus infection noted and recorded each day for 14 con secutive days postchallenge. Nasal swabs shall be collected from each dog each day for at least 10 consecutive days postchallenge. Individual swabs shall be tested for virus isolation by culture in canine parainfluenza virus susceptible cells for at least 7 days. Re sults shall be evaluated according to the following criteria: (i) If five of five controls have not re mained seronegative at a final serum dilution of 1:2 during the prechallenge period, the test is a No Test and may be repeated. (ii) If more than one vaccinate shows febrile response, respiratory or other clinical signs of canine parainfluenza virus infection; or, if less than 19 of 20 vaccinates show serum neutralization titers of 1:4 or greater; or, if there is not a significant reduction in virus iso lation rate in vaccinates when com pared with controls, the Master Seed is unsatisfactory. (5) An Outline of Production change shall be made before authority for use of a new lot of Master Seed shall be granted by Animal and Plant Health Inspection Service. (c) Test requirements for release. Each serial and subserial shall meet the ap plicable general requirements pre scribed in § 113.300 and the require ments in this paragraph. Any serial or subserial found unsatisfactory by a prescribed test shall not be released. (1) Virus titer requirements. Final con tainer samples of completed product shall be tested for virus titer using the titration method used in paragraph (b)(2) of this section. To be eligible for release, each serial and each subserial shall have a virus titer sufficiently greater than the titer of vaccine virus used in the immunogenicity test pre scribed in paragraph (b) of this section to assure that, when tested at any time within the expiration period, each se rial and subserial shall have a virus titer at least 10 0.7 greater than that used in the immunogenicity test but not less than 10 2.5 TCID50 per dose. (2) [Reserved] [50 FR 436, Jan. 4, 1985. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66784, 66786, Dec. 26, 1991; 72 FR 72564, Dec. 21, 2007] § 113.317 Parvovirus Vaccine (Canine). Parvovirus Vaccine recommended for use in dogs shall be prepared from virus-bearing cell culture fluids. Only Master Seed which has been estab lished as pure, safe, and immunogenic shall be used for preparing seeds for vaccine production. All serials of vac cine shall be prepared from the first through the fifth passage from the Master Seed. (a) The Master Seed shall meet the applicable general requirements pre scribed in § 113.300 and the require ments in this section. (b) The Master Seed shall be tested for reversion to virulence in dogs using a method acceptable to Animal and Plant Health Inspection Service. If a significant increase in virulence is seen within five backpassages, the Master Seed is unsatisfactory. (c) Each lot of Master Seed shall be tested for immunogenicity. The se lected virus dose shall be established as follows: (1) Twenty-five canine parvovirus susceptible dogs (20 vaccinates and 5 controls) shall be used as test animals. Blood samples drawn from each dog shall be individually tested for neutral izing antibody against canine parvovirus to determine susceptibility. Dogs shall be considered susceptible if there is no neutralization at a 1:2 final serum dilution in a constant virus varying serum neutralization test in VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00797 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 788 9 CFR Ch. I (1–1–18 Edition) § 113.318 cell culture using 50 to 300 TCID50 of canine parvovirus. (2) A geometric mean titer of the vac cine produced at the highest passage from the Master Seed shall be estab lished before the immunogenicity test is conducted. The 20 dogs used as vac cinates shall be administered a pre determined quantity of vaccine virus by the method recommended on the label. To confirm the dosage calcula tions, five replicate virus titrations shall be conducted on a sample of the vaccine virus dilution used. If two doses are used, five replicate con firming titrations shall be conducted on each dose. (3) Fourteen days or more after the final dose of vaccine the vaccinates and the controls shall be challenged with virulent canine parvovirus furnished or approved by Animal and Plant Health Inspection Service and the dogs ob served each day for 14 days. Rectal temperature, blood lymphocyte count, and feces for viral detection shall be taken from each dog each day for at least 10 days postchallenge and the presence or absence of clinical signs noted and recorded each day. (i) The immunogenicity of the Mas ter Seed shall be evaluated on the fol lowing criteria of infection: tempera ture ≥103.4 °F; lymphopenia of ≥50 per cent of prechallenge normal; clinical signs such as diarrhea, mucus in feces, or blood in feces; and viral hemagglutinins at a level of ≥1:64 in a 1:5 dilution of feces or a test of equal sensitivity. If at least 80 percent of the controls do not show at least three of the four criteria of infection during the observation period, the test is a No Test and may be repeated. (ii) If at least 19 of the 20 vaccinates do not survive the observation period without showing more than one cri terion of infection described in para graph (c)(3)(i), of this section, the Mas ter Seed is unsatisfactory. (4) An Outline of Production change shall be made before authority for use of a new lot of Master Seed shall be granted by Animal and Plant Health Inspection Service. (d) Test requirements for release. Each serial and subserial shall meet the ap plicable general requirements pre scribed in § 113.300 and the require ments in this paragraph. Any serial or subserial found unsatisfactory by a prescribed test shall not be released. (1) Virus titer requirements. Final con tainer samples of completed product shall be tested for virus titer using the titration method used in paragraph (c)(2) of this section. To be eligible for release, each serial and each subserial shall have a virus titer sufficiently greater than the titer of vaccine used in the immunogenicity test in para graph (c) of this section to assure that, when tested at any time within the ex piration period, each serial and sub serial shall have a virus titer of 10 0.7 greater than that used in the immunogenicity test, but not less than 10 2.5 ID50 per dose. [50 FR 436, Jan. 4, 1985. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66784, 66786, Dec. 26, 1991; 72 FR 72564, Dec. 21, 2007] § 113.318 Pseudorabies Vaccine. Pseudorabies Vaccine shall be pre pared from virus-bearing cell culture fluids. Only Master Seed which has been established as pure, safe, and immunogenic shall be used for pre paring seeds for vaccine production. All serials of vaccine shall be prepared from the first through the fifth passage from the Master Seed. (a) The Master Seed shall meet the applicable general requirements pre scribed in § 113.300 and the require ments in this section. (b) Each lot of Master Seed shall be tested for immunogenicity. The se lected virus dose shall be established as follows: (1) Twenty-five pseudorabies suscep tible pigs (20 vaccinates and 5 controls) of the youngest age for which the vac cine is recommended, shall be used as test animals. Blood samples shall be taken from each pig and the serums in activated and individually tested for neutralizing antibody against pseudorabies virus. Pigs shall be con sidered susceptible if there is no neu tralization at a 1:2 final serum dilution in a constant virus-varying serum neu tralization test using 50 to 300 TCID50 pseudorabies virus. (2) A geometric mean titer of the vac cine produced at the highest passage VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00798 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 789 Animal and Plant Health Inspection Service, USDA § 113.325 from the Master Seed shall be estab lished before the immunogenicity test is conducted. The 20 pigs used as vac cinates shall be administered a pre determined quantity of vaccine virus by the method recommended on the label. To confirm the dosage adminis tered, five replicate virus titrations shall be conducted on a sample of the vaccine virus dilution used. (3) Fourteen to 28 days postvaccination, the vaccinates and controls shall be challenged with viru lent pseudorabies virus furnished or ap proved by Animal and Plant Health In spection Service and observed each day for 14 days. (i) If at least four of the five controls do not develop severe central nervous system signs or die, the test is a No Test and may be repeated. (ii) If at least 19 of the 20 vaccinates in a valid test do not remain free of signs of pseudorabies, the Master Seed is unsatisfactory. (4) An Outline of Production change shall be made before authority for use of a new lot of Master Seed shall be granted by Animal and Plant Health Inspection Service. (c) Test requirements for release. Each serial and subserial shall meet the ap plicable general requirements pre scribed in § 113.300 and the require ments in this paragraph. (2) Virus titer requirements. Final con tainer samples of completed product shall be titrated by the method used in paragraph (b)(2) of this section. To be eligible for release, each serial and sub serial shall have a virus titer suffi ciently greater than the titer of the vaccine used in the immunogenicity test prescribed in paragraph (b) of this section to assure that, when tested at any time within the expiration period, each serial and subserial shall have a virus titer at least 10. 0.7 greater than that used in the immunogenicity test, but not less than 10 2.5 TCID50 per dose. [50 FR 437, Jan. 4, 1985. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66784, 66786, Dec. 26, 1991; 72 FR 72564, Dec. 21, 2007] §§ 113.319–113.324 [Reserved] § 113.325 Avian Encephalomyelitis Vaccine. Avian Encephalomyelitis Vaccine shall be prepared from virus-bearing tissues or fluids from embryonated chicken eggs. Only Master Seed Virus which has been established as pure, safe, and immunogenic in accordance with the requirements in paragraphs (a), (b), and (c) of this section shall be used for preparing the production seed virus for vaccine production. All serials shall be prepared from the first through the fifth passage from the Master Seed Virus. (a) The Master Seed Virus shall meet the applicable requirements prescribed in § 113.300 and the requirements pre scribed in this section. (b) Each lot of Master Seed Virus shall be tested for pathogens by the chicken embryo inoculation test pre scribed in § 113.37, except that, if the test is a No Test because of a vaccine virus override, the test may be re peated and if the repeat test is incon clusive for the same reason, the chick en inoculation test prescribed in § 113.36 may be conducted and the virus judged accordingly. (c) Each lot of Master Seed Virus shall be tested for immunogenicity and the selected virus dose to be used shall be established as follows: (1) Avian encephalomyelitis suscep tible chickens, all of the same age (eight weeks or older) and from the same source, shall be used. Twenty or more chickens shall be used as vac cinates for each method of administra tion recommended on the label. Ten additional chickens of the same age and from the same source shall be held as unvaccinated controls. (2) A geometric mean titer of the vac cine produced from the highest passage of the Master Seed Virus shall be es tablished before the immunogenicity test is conducted. Each vaccinate shall receive a predetermined quantity of vaccine virus. Five replicate virus ti trations shall be conducted on an ali quot of the vaccine virus to confirm the amount of virus administered to each chicken used in the test. At least VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00799 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 790 9 CFR Ch. I (1–1–18 Edition) § 113.326 three appropriate (not to exceed ten fold) dilutions shall be used and the test conducted as follows: (i) For each dilution, inoculate at least 10 embryos, 5 or 6 days old, in the yolk sac with 0.2 ml each. Twenty simi lar embryos obtained from the same source shall be kept as uninoculated negative controls. Disregard all deaths during the first 48 hours post-inocula tion. (ii) Eggs for each dilution shall be kept in separate containers and al lowed to hatch. Sufficient precaution shall be taken to assure that chickens from each dilution remain separated. To be a valid test, at least 75 percent of the uninoculated eggs shall hatch. (iii) On the third day after normal hatching time, count all unhatched eggs and all dead, paralyzed and ataxic chickens as positive evidence of viral infection. (iv) A satisfactory titration shall have at least one dilution with between 50 and 100 percent positives and at least one dilution with between 50 and 0 per cent positives. (v) Calculate the EID50 by the Spearman-Karber or Reed-Muench method. (3) At least 21 days post-vaccination, the vaccinates and the controls shall be challenged intracerebrally with a virulent avian encephalomyelitis virus and observed each day for 21 days. (4) If at least 80 percent of the con trols do not show signs of avian encephalomyelitis or die, the test is a No Test and may be repeated. If at least 19 of 20, or 27 of 30, or 36 of 40 of the vaccinates in each group do not re main free from clinical signs of avian encephalomyelitis during the observa tion period, the Master Seed Virus is unsatisfactory. (5) An Outline of Production change shall be made before authority for use of a new lot of Master Seed Virus shall be granted by Animal and Plant Health Inspection Service. (d) After a lot of Master Seed Virus has been established as prescribed in paragraphs (a), (b), and (c) of this sec tion, each serial and subserial shall meet the applicable requirements in § 113.300 and the requirements pre scribed in this paragraph. (1) Final container samples from each serial shall be tested for patho gens by the chicken embryo inocula tion test prescribed in § 113.37, except that, if the test is a No Test because of a vaccine virus override, the chicken inoculation test prescribed in § 113.36 may be conducted and the vaccine judged accordingly. (2) Safety test. Final container sam ples of completed product shall be test ed for safety as follows: (i) At least 25 AE susceptible birds (6 to 10 weeks of age) shall be vaccinated with the equivalent of 10 doses by each of all routes recommended on the label and be observed each day for 21 days. (ii) If unfavorable reactions attrib utable to the biological product occur during the observation period, the se rial is unsatisfactory. If unfavorable reactions occur which are not attrib utable to the product, the test shall be declared a No Test and repeated, except that, if the test is not repeated, the se rial shall be unsatisfactory. (3) Virus titer requirements. Final con tainer samples of completed product shall be tested for virus titer using the titration method used in paragraph (c)(2) of this section. To be eligible for release, each serial and each subserial shall have a virus titer sufficiently greater than the titer of vaccine virus used in the immunogenicity test pre scribed in paragraph (c) of this section to assure that when tested at any time within the expiration period, each se rial and subserial shall have a virus titer of 10 0.7 greater than that used in such immunogenicity test but not less than 10 .5 EID50 per dose. [39 FR 44723, Dec. 27, 1974, as amended at 40 FR 18405, Apr. 28, 1975; 40 FR 41089, Sept. 5, 1975; 42 FR 43617, Aug. 30, 1977; 48 FR 33473, July 22, 1983. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66784, 66786, Dec. 26, 1991; 72 FR 72564, Dec. 21, 2007; 79 FR 55969, Sept. 18, 2014] § 113.326 Avian Pox Vaccine. Fowl Pox Vaccine and Pigeon Pox Vaccine shall be prepared from virus bearing cell culture fluids or embryonated chicken eggs. Only Mas ter Seed Virus which has been estab lished as pure, safe, and immunogenic in accordance with the requirements in VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00800 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 791 Animal and Plant Health Inspection Service, USDA § 113.326 paragraphs (a), (b), and (c) of this sec tion shall be used for preparing the production seed virus for vaccine pro duction. All serials shall be prepared from the first through the fifth passage from the Master Seed Virus. (a) The Master Seed Virus shall meet the applicable requirements prescribed in § 113.300 except paragraph (c) of this section and shall meet the require ments prescribed in this section. (b) Each lot of Master Seed Virus shall be tested for pathogens by the chicken inoculation test prescribed in § 113.36. (c) Each lot of Master Seed Virus shall be tested for immunogenicity and the selected virus dose to be used shall be established as follows: (1) Fowl pox susceptible birds all of the same age and from the same source, shall be used as test birds. Twenty or more birds shall be used as vaccinates for each method of adminis tration recommended on the label. Ten additional birds of the same age and from the same source as the vaccinates shall be held as unvaccinated controls. (2) A geometric mean titer of the dried vaccine produced from the high est passage of the Master Seed Virus shall be established before the immunogenicity test is conducted. Each vaccinate shall receive a pre determined quantity of vaccine virus. Five replicate virus titrations shall be conducted on an aliquot of the vaccine virus to confirm the amount of virus administered to each bird used in the test. At least three appropriate (not to exceed tenfold) dilutions shall be used and the test conducted as follows: (i) For each dilution, inoculate at least five embryos, 9 to 11 days old, on the chorioallantoic membrane with at least 0.2 ml each. Disregard all deaths during the first 24 hours post-inocula tion. To be a valid test, at least four embryos in each dilution shall remain viable beyond 24 hours. (ii) Examine the surviving embryos for evidence of infection 5 to 7 days post-inoculation. (iii) A satisfactory titration shall have at least one dilution with between 50 and 100 percent positives and at least one dilution with between 50 and 0 per cent positives. (iv) Calculate the EID50 by the Spearman-Karber or Reed-Muench method. (3) Fourteen to twenty-one days post vaccination, all vaccinates and con trols shall be challenged by the wing web method and observed each day for 10 days. If the wing web method was used for vaccination, the opposite wing shall be used for challenge. Challenge virus shall be provided or approved by Animal and Plant Health Inspection Service. (4) If at least 90 percent of the con trols do not develop fowl pox during the observation period, the test is a No Test and may be repeated. If at least 19 of 20, or 27 of 30, or 36 of 40 of the vac cinates in each group do not remain free from clinical signs of fowl pox dur ing the observation period, the Master Seed Virus is unsatisfactory. (5) An Outline of Production change shall be made before authority for use of a new lot of Master Seed Virus shall be granted by Animal and Plant Health Inspection Service. (d) After a lot of Master Seed Virus has been established as prescribed in paragraphs (a), (b), and (c) of this sec tion, each serial and subserial shall meet the requirements in § 113.36, in § 113.300 except paragraph (c), and in this paragraph. (1) Safety test. Final container sam ples of completed product from each se rial shall be tested. Vaccines rec ommended for use in birds 10 days of age or younger shall be tested in ac cordance with paragraphs (d)(1)(i), (ii), and (iii) of this section. (i) Each of 25 susceptible birds 5 days of age or younger, properly identified and obtained from the same source and hatch, shall be vaccinated with the equivalent of 10 doses of vaccine by each of all routes recommended on the label and observed each day for 14 days. Severe clinical signs or death shall be counted as failures. Two-stage sequen tial testing may be conducted if the first test (which then becomes stage one) has three failures. (ii) The results shall be evaluated ac cording to the following table: VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00801 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 792 9 CFR Ch. I (1–1–18 Edition) § 113.327 CUMULATIVE TOTALS Stage Number of birds Failures for satisfactory serials Failures for unsatisfactory serials 1 ................... 25 ................ 2 or less ...... 4 or more. 2 ................... 50 ................ 5 or less ...... 6 or more. (iii) If unfavorable reactions occur which are not attributable to the prod uct, the test shall be declared a No Test and may be repeated or, in lieu thereof, the serial declared unsatisfac tory. (iv) Vaccines not recommended for use in birds 10 days of age or younger shall be tested for safety as follows: Each of twenty-five 3- to 5-week-old, fowl-pox susceptible birds shall be vac cinated with the equivalent of 10 doses of vaccine by each of all routes rec ommended on the label and observed each day for 14 days. If any of the birds show severe clinical signs of disease or death during the observation period due to causes attributable to the prod uct, the serial is unsatisfactory. If un favorable reactions occur which are not attributable to the product, the test shall be declared a No Test and may be repeated or, in lieu thereof, the serial declared unsatisfactory. (2) Virus titer requirements. Final con tainer samples of completed product shall be tested for virus titer using the titration method used in paragraph (c)(2) of this section. To be eligible for release, each serial and each subserial shall have a virus titer sufficiently greater than the titer of vaccine virus used in the immunogenicity test pre scribed in paragraph (c) of this section to assure that when tested at any time within the expiration period, each se rial and subserial shall have a virus titer of 10 0.7 greater than that used in such immunogenicity test but not less than 10 2.0 EID50 per dose. [39 FR 44724, Dec. 27, 1974, as amended at 40 FR 18406, Apr. 28, 1975; 40 FR 41089, Sept. 5, 1975; 44 FR 33051, June 8, 1979; 48 FR 33473, July 22, 1983. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66784, 66786, Dec. 26, 1991; 72 FR 72564, Dec. 21, 2007] § 113.327 Bronchitis Vaccine. Bronchitis Vaccine shall be prepared from virus-bearing cell culture fluids or embryonated chicken eggs. Only Master Seed Virus which has been es tablished as pure, safe, and immunogenic in accordance with the requirements in paragraphs (a), (b), and (c) of this section shall be used for pre paring the production seed virus for vaccine production. All serials shall be prepared from the first through the fifth passage from the Master Seed Virus. (a) The Master Seed Virus shall meet the applicable requirements prescribed in § 113.300 and the requirements pre scribed in this section. (b) Each lot of Master Seed Virus shall be tested for pathogens by the chicken embryo inoculation test pre scribed in § 113.37, except that, if the test is a No Test because of a vaccine virus override, the test may be re peated and if the repeat test is a No Test for the same reason, the chicken inoculation test prescribed in § 113.36 may be conducted and the virus judged accordingly. (c) Each lot of Master Seed Virus used for vaccine production shall be tested for immunogenicity and the se lected virus dose to be used shall be es tablished as follows: (1) Bronchitis susceptible chickens, all of the same age and from the same source, shall be used in the virus-recov ery test. For each method of adminis tration recommended on the label for each serotype against which protection is claimed, twenty or more chickens shall be used as vaccinates. Ten addi tional chickens for each serotype against which protection is claimed shall be held as unvaccinated controls. (2) A geometric mean titer of the dried vaccine produced from the high est passage of the Master Seed Virus shall be established before the immunogenicity tests are conducted. Each vaccinate shall receive a pre determined quantity of vaccine virus. Five replicate virus titrations shall be conducted on an aliquot of the vaccine virus to confirm the amount of virus administered to each chicken used in such tests. At least three approved (not to exceed tenfold) dilutions shall be used and the test conducted as follows; (i) For each dilution, inject at least five embryos, 9 to 11 days old, in the allantoic cavity with 0.1 ml each. Deaths occurring during the first 24 hours shall be disregarded, but at least VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00802 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 793 Animal and Plant Health Inspection Service, USDA § 113.327 four viable embyros in each dilution shall survive beyond 24 hours of a valid test. After 5 to 8 days incubation, ex amine the surviving embryos for evi dence of infection. (ii) A satisfactory titration shall have at least one dilution with between 50 and 100 percent positives and at least one dilution with between 50 and 0 per cent positives. (iii) Calculate the EID50 by the Spearman-Karber or Reed-Muench method. (3) Twenty-one to twenty-eight days post-vaccination, all vaccinates and controls shall be challenged by eye drop with virulent bronchitis virus. A separate set of vaccinates and controls shall be used for each serotype against which protection is claimed. Each chal lenge virus shall be approved or pro vided by Animal and Plant Health In spection Service and shall titer at least 10 4.0 EID50 per ml. (i) Tracheal swabs shall be taken once, 5 days post-challenge, from each control and vaccinate. Each swab shall be placed in a test tube containing 3 ml of tryptose phosphate broth and anti biotics. The tube and swab shall be swirled thoroughly and if they are to be stored, be immediately frozen and be stored at below ¥40 °C. pending egg evaluation. For each chicken swab, at least five chicken embryos 9 to 11 days old shall be inoculated in the allantoic cavity with 0.2 ml each of broth from each tube. (ii) All embryos surviving the third day post-inoculation shall be used in the evaluation, except that, if a swab is not represented by at least four em bryos, the test of that swab is invalid and the results a No Test. A tracheal swab shall be positive for virus recov ery when any of the embryos in a valid test show typical infectious bronchitis virus lesions, such as but not limited to, stunting, curling, kidney urates, clubbed down, or death during the 4 to 7 day post-inoculation period. If less than 20 percent of the embryos which survive the third day post-inoculation die during the 4 to 7 day post-inocula tion period and show no gross lesions typical of infectious bronchitis, they may be disregarded. (iii) If less than 90 percent of the con trols are positive for virus recovery, the test is a No Test and may be re peated. (iv) If less than 90 percent of the vac cinates are negative for virus recovery, the Master Seed Virus is unsatisfac tory. (4) An Outline of Production change shall be made before authority for use of a new lot of Master Seed Virus shall be granted by Animal and Plant Health Inspection Service. (d) After a lot of Master Seed Virus has been established as prescribed in paragraphs (a), (b), and (c) of this sec tion, each serial and subserial shall meet the applicable requirements in § 113.300 and the requirements pre scribed in this paragraph, except that, if the vaccine contains more than one virus type, bulk samples taken from each type prior to mixing shall be used in the virus identity tests prescribed in § 113.300(c). The additional require ments in this paragraph shall also be met. (1) Final container samples from each serial shall be tested for patho gens by the chicken embryo inocula tion test prescribed in § 113.37, except that, if the test is a No Test because of a vaccine virus override, the chicken inoculation test prescribed in § 113.36 may be conducted and the vaccine judged accordingly. (2) Safety test. Final container sam ples of completed product shall be test ed to determine safety for use in bron chitis susceptible young chickens. (i) Twenty-five susceptible chickens, 5 days of age or younger, properly iden tified and obtained from the same source and hatch, shall be vaccinated by the eye-drop method with the equiv alent of 10 doses of vaccine and ob served each day for 21 days post-vac cination. Severe respiratory signs or death shall be counted as failures. Two stage sequential testing may be con ducted if the first test (which then be comes stage one) has three failures. (ii) The results shall be evaluated ac cording to the following table: CUMULATIVE TOTALS Stage Number of chickens Failures for satisfactory serials Failures for unsatisfactory serials 1 ................... 25 ................ 2 or less ...... 4 or more. 2 ................... 50 ................ 5 or less ...... 6 or more. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00803 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 794 9 CFR Ch. I (1–1–18 Edition) § 113.328 If unfavorable reactions occur which are not attributable to the product, the test shall be declared a No Test and re peated or, in lieu thereof, the serial de clared unsatisfactory. (3) Virus titer requirements. Final con tainer samples of completed product shall be tested for virus titer using the procedure prescribed in paragraph (c)(2) of this section and in this para graph. (i) The Newcastle disease virus frac tion of combined Newcastle-Bronchitis Vaccines shall be neutralized prior to titration of the bronchitis virus frac tion. Equal parts of heat-inactivated Newcastle disease antiserum shall be mixed with each appropriate serial ten fold dilution of the vaccine. After inac tivation, embryos shall be injected with 0.2 ml each and results calculated as a 0.1 ml dose to allow for serum dilu tion of the vaccine. The allantoic fluids, tested as prescribed in § 113.34 shall not show hemagglutinating activ ity in the lowest dilution used in the titration. (ii) Each bronchitis virus type shall be harvested separately and a sample of bulk harvested material shall be col lected prior to mixing with the other virus type(s). Each sample shall con tain not less than the minimum virus titer stated in the filed Outline of Pro duction. (iii) To be eligible for release, each serial and each subserial shall have a virus titer sufficiently greater than the titer of vaccine virus used in the immunogenicity test prescribed in paragraph (c) of this section to assure that when tested at any time within the expiration period, each serial and subserial shall have a virus titer of 10 0.7 greater than that used in such immunogenicity test but not less than 10 2.0 EID50 per dose. [39 FR 44724, Dec. 27, 1974, as amended at 40 FR 18406, Apr. 28, 1975; 40 FR 41089, Sept. 5, 1975; 42 FR 43617, Aug. 30, 1977; 48 FR 33473, July 22, 1983. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66784, 66786, Dec. 26, 1991; 64 FR 43045, Aug. 9, 1999; 72 FR 72564, Dec. 21, 2007] § 113.328 Fowl Laryngotracheitis Vac cine. Fowl Laryngotracheitis Vaccine shall be prepared from virus-bearing cell culture fluids or embryonated chicken eggs. Only Master Seed Virus which has been established as pure, safe, and immunogenic in accordance with the requirements in paragraphs (a), (b), and (c) of this section shall be used for preparing the production seed virus for vaccine production. All serials shall be prepared from the first through the fifth passage from the Master Seed Virus. (a) The Master Seed Virus shall meet the applicable requirements prescribed in § 113.300 and the requirements pre scribed in this section. (b) Each lot of Master Seed Virus shall be tested for pathogens by the chicken embryo inoculation test pre scribed in § 113.37, except that, if the test is a No Test because of vaccine virus override, the test may be re peated and if the repeat test is a No Test for the same reason, the chicken inoculation test prescribed in § 113.36 may be conducted and the virus judged accordingly. Each lot shall also be test ed for safety as follows: (1) Each of at least ten 3 to 4 week old susceptible chickens obtained from the same source and hatch as those used in the immunogenicity test pre scribed in paragraph (c) of this section shall be injected intratracheally with 0.2 ml of the virus as used in the vac cine and the chickens observed each day for 14 days. (2) If more than 20 percent of the chickens die during the observation pe riod, the virus is unsatisfactory. (c) Each lot of Master Seed Virus used for vaccine production shall be tested for immunogenicity and the se lected virus dose to be used shall be es tablished as follows: (1) Fowl laryngotracheitis suscep tible chickens all of the same age and from the same source shall be used. Twenty or more chickens shall be used as vaccinates for each method of ad ministration recommended on the label. Ten additional chickens of the same age and from the same source shall be held as unvaccinated controls. (2) A geometric mean titer of the dried vaccine produced from the high est passage of the Master Seed Virus shall be established before the immunogenicity test is conducted. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00804 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 795 Animal and Plant Health Inspection Service, USDA § 113.328 Each vaccinate shall receive a pre determined quantity of vaccine virus. Five replicate virus titrations shall be conducted on an aliquot of the vaccine virus to confirm the amount of virus administered to each chicken used in the test. At least three appropriate (not to exceed tenfold) dilutions shall be used for vaccine of chicken embryo origin and the test conducted as fol lows: (i) For each dilution, inject at least five embryos, 9 to 11 days old, on the chorioallantoic membrane with 0.2 ml each. Disregard all deaths during the first 24 hours post-injection. To be a valid test, at least four embryos in each dilution shall remain viable be yond 24 hours. (ii) Examine the surviving embryos for evidence of infection 5 to 8 days post-injection. (iii) A satisfactory titration shall have at least one dilution with between 50 and 100 percent positives and at least one dilution with between 50 and 0 per cent positives. (iv) Calculate the EID50 by the Spearman-Karber or Reed-Muench method. (3) Tissue culture origin vaccine may be titrated by a tissue culture method approved by Animal and Plant Health Inspection Service and written into the filed Outline of Productions. (4) Ten to fourteen days post-vaccina tion, all vaccinates and controls shall be challenged intratracheally or in the orbital sinus with infectious fowl laryngotracheitis virus and observed each day for 10 days. Challenge virus shall be provided or approved by Ani mal and Plant Health Inspection Serv ice. (5) If at least 80 percent of the con trols do not die or show clinical signs of fowl laryngotracheitis during the ob servation period, the test is a No Test and may be repeated. If at least 19 of 20, 27 of 30, or 36 of 40 of the vaccinates in each group do not remain free of clinical signs of fowl laryngotracheitis during the observation period, the Mas ter Seed Virus is unsatisfactory. (6) An Outline of Production change shall be made before authority for use of a new lot of Master Seed Virus shall be granted by Animal and Plant Health Inspection Service. (d) After a lot of Master Seed Virus has been established as prescribed in paragraphs (a), (b), and (c) of this sec tion, each serial and subserial shall meet the applicable requirements in § 113.300 and the requirements pre scribed in this paragraph. (1) Final container samples from each serial shall be tested for patho gens by the chicken embryo inocula tion test prescribed in § 113.37, except that, if the test is a No Test because of a vaccine virus override, the chicken inoculation test prescribed in § 113.36 may be conducted and the vaccine judged accordingly. (2) Safety test. Final container sam ples of completed product from each se rial of modified live virus vaccine shall be tested for safety as provided in this paragraph. Live virus vaccine not pre pared with modified live virus shall be tested for safety as provided in the filed Outline of Production. (i) Twenty-five 3 to 4 week old laryngotracheitis susceptible chickens shall be injected intratracheally with 0.2 ml of vaccine rehydrated at the rate of 30 ml for 1,000 doses. Chickens shall be observed each day for 14 days. Deaths shall be counted as failures. Two-stage sequential testing may be conducted if the first test (which then becomes stage one) has five, six, or seven failures. (ii) The results shall be evaluated ac cording to the following table: CUMULATIVE TOTALS Stage Number of chickens Failures for satisfactory serials Failures for unsatisfactory serials 1 ................... 25 ................ 4 or less ...... 8 or more. 2 ................... 50 ................ 10 or less .... 11 or more. (iii) If unfavorable reactions occur which are not attributable to the prod uct, the test shall be declared a No Test and repeated or in lieu thereof, the serial declared unsatisfactory. (3) Virus titer requirements. Final con tainer samples of completed product shall be tested for virus titer using the titration method provided in para graphs (c)(2) or (3) of this section. To be eligible for release, each serial and each subserial shall have a virus titer sufficiently greater than the titer of vaccine virus used in the VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00805 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 796 9 CFR Ch. I (1–1–18 Edition) § 113.329 immunogenicity test prescribed in paragraph (c) of this section to assure that when tested at any time within the expiration period, each serial and subserial shall have a virus titer of 10 0.7 greater than that used in such immunogenicity test but not less than 10 2.5 EID50 per dose for chicken embryo origin vaccine and 10 2.0 EID50 or 10 2.5 TCID50 per dose for tissue culture ori gin vaccine. [39 FR 44726, Dec. 27, 1974, as amended at 40 FR 18407, Apr. 28, 1975; 40 FR 41089, Sept. 5, 1975; 41 FR 44359, Oct. 8, 1976; 42 FR 43617, Aug. 30, 1977; 48 FR 33473, July 22, 1983. Re designated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66784, 66786, Dec. 26, 1991; 72 FR 72564, Dec. 21, 2007] § 113.329 Newcastle Disease Vaccine. Newcastle Disease Vaccine shall be prepared from virus-bearing cell cul ture fluids or embryonated chicken eggs. Only Master Seed Virus which has been established as pure, safe, and immunogenic in accordance with the requirements in paragraphs (a), (b), and (c) of this section shall be used for pre paring the production seed virus for vaccine production. All serials shall be prepared from the first through the fifth passage from the Master Seed Virus. (a) The Master Seed Virus shall meet the applicable requirements prescribed in § 113.300, except § 113.34, and the re quirements prescribed in this section. (b) Each lot of Master Seed Virus shall be tested for pathogens by the chicken embryo inoculation test pre scribed in § 113.37, except that, if the test is a No Test because of a vaccine virus override, the test may be re peated and if the repeat test is a No Test for the same reason, the chicken inoculation test prescribed in § 113.36 may be conducted and the virus judged accordingly. (c) Each lot of Master Seed Virus used for vaccine production shall be tested for immunogenicity and the se lected virus dose to be used shall be es tablished as follows: (1) Newcastle Disease susceptible chickens, all of the same age and from the same source, shall be used. Twenty or more chickens shall be used as vac cinates for each method of administra tion recommended on the label. Ten additional chickens of the same age and from the same source shall be held as unvaccinated controls. (2) A geometric mean titer of the dried vaccine produced from the high est passage of the Master Seed Virus shall be established before the immunogenicity test is conducted. Each vaccinate shall receive a pre determined quantity of vaccine virus. Five replicate virus titrations shall be conducted on an aliquot of the vaccine virus to confirm the amount of virus administered to each chicken used in the test. At least three appropriate (not to exceed tenfold) dilutions shall be used and the test conducted as fol lows: (i) For each dilution, inject at least five embryos, 9 to 11 days old, in the allantoic cavity with at least 0.1 ml each. Disregard all deaths during the first 24 hours post-injection. To be a valid test, at least four embryos in each dilution shall remain viable be yond 24 hours. (ii) Examine the surviving embryos for evidence of infection 5 to 7 days post-injection. (iii) A satisfactory titration shall have at least one dilution with between 50 and 100 percent positives and at least one dilution with between 50 and 0 per cent positives. (iv) Calculate the EID50 by the Spearman-Karber or Reed-Muench method. (3) Twenty to twenty-eight days postvaccination, all vaccinates and controls shall be challenged intramuscularly with at least 10 4.0 EID50 of virus per chicken and observed each day for 14 days. Challenge virus shall be provided or approved by Ani mal and Plant Health Inspection Serv ice. (4) If at least 90 percent of the con trols do not develop clinical signs of Newcastle disease during the observa tion period, the test is a No Test and may be repeated. If at least 19 of 20, or 27 of 30, or 36 of 40 of the vaccinates in each group do not remain free from clinical signs of Newcastle disease dur ing the observation period, the Master Seed Virus is unsatisfactory. (5) A strain identity test acceptable to Animal and Plant Health Inspection Service shall be conducted. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00806 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 797 Animal and Plant Health Inspection Service, USDA § 113.330 (6) An Outline of Production change shall be made before authority for use of a new lot of Master Seed Virus shall be granted by Animal and Plant Health Inspection Service. (d) After a lot of Master Seed Virus has been established as prescribed in paragraphs (a), (b), and (c) of this sec tion, each serial and subserial shall meet the applicable requirements in § 113.300, except § 113.34, and the require ments prescribed in this paragraph. (1) Final container samples from each serial shall be tested for patho gens by the chicken embryo inocula tion test prescribed in § 113.37, except that, if the test is a No Test because of a vaccine virus override, the chicken inoculation test prescribed in § 113.36 may be conducted and the vaccine judged accordingly. (2) Safety test: Final container sam ples of completed product from each se rial shall be tested to determine wheth er the vaccine is safe for use in suscep tible young chickens. Vaccines rec ommended for use in chickens 10 days of age or younger shall be tested in ac cordance with paragraphs (d)(2)(i), (ii), and (iii) of this section. (i) Twenty-five susceptible chickens, 5 days of age or younger, properly iden tified and obtained from the same source and hatch, shall be vaccinated by the eye drop method with the equiv alent of 10 doses of vaccine and the chickens observed each day for 21 days. Severe respiratory signs or death shall be counted as failures. Two-stage se quential testing may be conducted if the first test (which then becomes stage one) has 3 failures. (ii) The results shall be evaluated ac cording to the following table: CUMULATIVE TOTALS Stage Number of chickens Failures for satisfactory serials Failures for unsatisfactory serials 1 ................... 25 ................ 2 or less ...... 4 or more. 2 ................... 50 ................ 5 or less ...... 6 or more. (iii) If unfavorable reactions occur which are not attributable to the prod uct, the test shall be declared a No Test and may be repeated. (iv) Vaccines not recommended for use in chickens 10 days of age or younger shall be tested for safety as follows: Each of twenty-five 3 to 5 week old Newcastle disease susceptible chickens shall be vaccinated as recommended on the label with the equivalent of ten doses and observed each day for 21 days. If any of the birds show severe clinical signs of disease or death during the observation period due to causes attributable to the product, the serial is unsatisfactory. (3) Virus titer requirements. Final con tainer samples of completed product shall be tested for virus titer using the titration method used in paragraph (c)(2) of this section. To be eligible for release, each serial and each subserial shall have a virus titer per dose suffi ciently greater than the titer of vac cine virus used in the immunogenicity test prescribed in paragraph (c) of this section to assure that when tested at any time within the expiration period, each serial and subserial shall have a virus titer of 10 0.7 greater than that used in the immunogenicity test but not less than 10 5.5 EID50 per dose. [39 FR 44727, Dec. 27, 1974, as amended at 40 FR 18407, Apr. 28, 1975; 40 FR 23721, June 2, 1975; 40 FR 41090, Sept. 5, 1975; 42 FR 43618, Aug. 30, 1977; 48 FR 33473, July 22, 1983. Re designated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66784, 66786, Dec. 26, 1991; 72 FR 72564, Dec. 21, 2007] § 113.330 Marek’s Disease Vaccines. Marek’s disease vaccine shall be pre pared from virus-bearing tissue culture cells. Only Master Seed Virus which has been established as pure, safe, and immunogenic shall be used for pre paring the production seed virus for vaccine production. (a) The Master Seed Virus shall meet the applicable requirements prescribed in § 113.300, and the requirements pre scribed in this section. The identity test required in § 113.300(c) shall be con ducted in a serotype-specific manner by a method acceptable to APHIS. Each lot of Master Seed Virus shall also be tested for pathogens by the chicken embryo inoculation test pre scribed in § 113.37, except that, if the test is a No Test because of a vaccine virus override, the chicken inoculation VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00807 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 798 9 CFR Ch. I (1–1–18 Edition) § 113.330 test prescribed in § 113.36 may be con ducted and the virus judged accord ingly. (b) Safety test. The Master Seed Virus shall be nonpathogenic for chickens as determined by the following procedure: (1) Specific pathogen free chickens or embryos, negative for Marek’s disease virus antibodies, and from the same source, shall be isolated into the fol lowing groups: (i) Group 1. At least 50 test subjects shall be inoculated with 10 times as much viable virus as will be contained in one dose of vaccine, by the route recommended for vaccination. (ii) Group 2. At least 50 test subjects shall be injected with a very virulent Marek’s disease virus provided or ap proved by APHIS, at a dosage level that will cause gross lesions of Marek’s disease in at least 80 per cent of the chickens within 50 days. (iii) Group 3. Fifty uninoculated con trols. For in ovo studies, this group should receive a sham inoculation of diluent. (iv) Group 4. For studies evaluating Serotype 1 Master Seed Viruses, a group of 50 uninoculated control chick ens shall be housed in contact with the group 1 vaccinated chickens. (2) At least 40 chickens in each group shall survive to 5 days of age. All chickens that die shall be necropsied and examined for lesions of Marek’s disease and cause of death. The test shall be judged according to the fol lowing criteria: (i) At 50 days of age, the remaining chickens in group 2 shall be killed and examined for gross lesions of Marek’s disease. If at least 80 percent of this group do not develop Marek’s disease, the test is a No Test and may be re peated. (ii) At 120 days of age, the remaining chickens in groups 1, 3, and 4 shall be weighed, killed, and necropsied. If less than 30 of the chickens in group 3 sur vive the 120 day period, or if any of the chickens in group 3 have gross lesions of Marek’s disease at necropsy, the test is declared a No Test. If less than 30 chickens in groups 1 and 4 survive the 120 day period; or if any of the chickens in groups 1 and 4 have gross lesions of Marek’s disease at necropsy; or if the average body weight of the chickens in groups 1 or 4 is significantly (statis tically) different from the average in group 3 at the end of the 120 days, the lot of Master Seed Virus is unsatisfac tory. (3) For tests involving in ovo inocula tion, hatchability results shall also be reported for each group. (c) Immunogenicity. Each lot of Mas ter Seed Virus used for vaccine produc tion shall be tested for immunogenicity at the highest passage level allowed for the product, and the virus dose to be used shall be estab lished as follows: (1) Specific pathogen free chickens or embryos, negative for Marek’s disease antibodies, and from the same source, shall be isolated into the following groups: (i) Group 1. A minimum of 35 test sub jects shall be inoculated with the vac cine, using the recommended route, at 1 day of age for chicks or 18 days of embryonation for embryos. The dose used shall be established by 5 replicate virus titrations conducted by a cell culture system or other titration method acceptable to APHIS. (ii) Group 2. A minimum of 35 nonvac cinated test subjects shall be held as challenge controls. (iii) Group 3. A minimum of 25 non vaccinated test subjects shall be held as nonchallenge controls. (iv) Group 4. Except for studies evalu ating vaccines which contain only a Serotype 3 virus as the Marek’s disease fraction, a minimum of 35 chicks shall be vaccinated at 1 day of age with a li censed Serotype 3 vaccine, in order to document the severity of the very viru lent challenge. (2) At least 30 chickens in groups 1, 2, and 4, and at least 20 chickens in group 3, shall survive to 5 days of age. All chickens in groups 1, 2, and 4 shall be challenged at 5 days of age in the fol lowing manner: (i) For studies evaluating vaccines which contain only a Serotype 3 virus as the Marek’s disease fraction, groups 1 and 2 shall be inoculated with a standard virulent challenge virus pro vided or approved by APHIS. (ii) For all other Marek’s disease vac cines, groups 1, 2, and 4 shall be inocu lated with a very virulent challenge virus provided or approved by APHIS. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00808 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 799 Animal and Plant Health Inspection Service, USDA § 113.331 (3) All chickens shall be observed until 7 weeks of age, necropsied, and examined for grossly observable lesions consistent with Marek’s disease. All chickens dying before the end of the 7 week observation period shall be necropsied and evaluated for gross le sions of Marek’s disease. Any chickens not so examined shall be scored as posi tive for Marek’s disease. (4) For a valid test, at least 80 per cent of the chickens in group 2 must develop grossly observable lesions, none of the chickens in group 3 shall develop grossly observable lesions, and (when included) greater than 20 percent of the chickens in group 4 must develop grossly observable lesions. (5) For a valid test to be considered satisfactory, at least 80 percent of the chickens in group 1 must remain free of grossly observable lesions. The appro priate product claim resulting from a satisfactory test would be to aid in the prevention of Marek’s disease, for vac cines containing only a Serotype 3 virus as the Marek’s disease fraction, or to aid in the prevention of very viru lent Marek’s disease, for all other vac cines. (d) Test requirements for release. Each serial and subserial shall meet the ap plicable requirements prescribed in § 113.300. The identity test required in § 113.300(c) shall be conducted in a serotype-specific manner by a method acceptable to APHIS. Final container samples of completed product shall also meet the requirements in para graphs (d) (1), (2), and (3) of this sec tion. Any serial or subserial found un satisfactory by a prescribed test shall not be released. (1) Purity test. The chicken embryo inoculation test prescribed in § 113.37 shall be conducted, except that, if the test is a No Test because of a vaccine virus override, the chicken inoculation test prescribed in § 113.36 may be con ducted and the virus judged accord ingly. (2) Safety test. At least 25 one-day-old, specific pathogen free chickens shall be injected, by the subcutaneous route, with the equivalent of 10 chicken doses of virus (vaccine concentrated 10X). The chickens shall be observed each day for 21 days. Chickens dying during the period shall be examined, cause of death determined, and the results re corded. (i) If at least 20 chickens do not sur vive the observation period, the test is a No Test. (ii) If lesions of any disease or cause of death are directly attributable to the vaccine, the serial is unsatisfac tory. (iii) If less than 20 chicks survive the observation period and there are no deaths or lesions attributable to the vaccine, the test may be repeated one time, Provided, that if the test is not repeated, the serial shall be declared unsatisfactory. (3) Potency test. The samples shall be titrated using a cell culture system or other titration method acceptable to APHIS. For vaccines composed of more than one Marek’s disease virus serotype, each fraction shall be titrated in a serotype-specific manner. (i) Samples of desiccated vaccine shall be incubated at 37 °C for 3 days before preparation for use in the po tency test. Samples of desiccated or frozen vaccine shall be reconstituted in diluent according to the label rec ommendations, and held in an ice bath at 0 °C to 4 °C for 2 hours prior to use in the potency test. (ii) For a serial or subserial to be eli gible for release, each serotype con tained in the vaccine shall have a virus titer per dose which is at least 3 times greater than the number of plaque forming units (pfu) used in the immunogenicity test prescribed in paragraph (c) of this section, but not less than 1000 pfu per dose. (iii) When tested (without the pretest incubation of desiccated products) at any time within the expiration period, each serotype contained in the vaccine shall have a virus titer per dose which is at least 2 times the number of pfu used in the immunogenicity test, but not less than 750 pfu per dose. [61 FR 33841, July 1, 1996] § 113.331 Bursal Disease Vaccine. Bursal Disease Vaccine shall be pre pared from virus-bearing cell culture fluids or embryonated chicken eggs. Only Master Seed Virus which has been established as pure, safe, and immunogenic in accordance with the requirements in paragraphs (a), (b), and VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00809 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 800 9 CFR Ch. I (1–1–18 Edition) § 113.331 (c) of this section shall be used for pre paring the production seed virus for vaccine production. All serials shall be prepared from the first through the fifth passage from the Master Seed Virus. (a) The Master Seed Virus shall meet the applicable requirements prescribed in § 113.300 and the requirements pre scribed in this section. (b) Each lot of Master Seed Virus shall be tested for pathogens by the chicken embryo inoculation test pre scribed in § 113.37, except that, if the test is a No Test because of a vaccine virus override, the chicken inoculation test prescribed in § 113.36 may be con ducted and the virus judged accord ingly. Each lot of Master Seed Virus used in the preparation of modified live virus vaccines shall also be nonpatho genic to chickens as determined by the following procedures: (1) Each of twenty-five 1-day-old bursal disease susceptible chickens (vaccinates) shall be injected subcutaneously with 10 times the rec ommended dose of vaccine virus and observed for 21 days. Fifteen chickens of the same source and hatch shall be kept isolated as controls. (i) Seventeen days postvaccination, each of five controls shall be adminis tered at least 10 2.0 EID50 of a virulent bursal disease virus by eye-drop, iso lated, and used as positive controls. The remaining controls shall be used as negative controls. (ii) If the vaccinates do not remain free of clinical signs of bursal disease, the Master Seed Virus is unsatisfac tory. If unfavorable reactions which are not attributable to the Master Seed Virus occur in more than two of the vaccinates, the test shall be declared a No Test and may be repeated. (iii) Twenty-one days postvaccination, the vaccinates and the controls shall be necropsied and ex amined for gross lesions of bursal dis ease. If more than two of the vac cinates have such lesions, the Master Seed Virus is unsatisfactory, except that, if any of the negative controls or less than four of the positive controls have such lesions, the test is a No Test and may be repeated. For purposes of this test, gross lesions shall include ob vious pathological processes and/or ob vious reduction in size of the bursa from normal. (2) Each of thirty-five 3- to 4-week old bursal disease susceptible chickens (vaccinates) shall be vaccinated with approximately one minimum protec tive dose of vaccine virus as deter mined in paragraph (c) of this section. Each of 10 chickens of the same source and hatch shall be administered at least 10 2.0 EID50 of a virulent bursal disease virus by eye-drop, isolated, and used as positive controls. Also, each of 20 additional chickens of the same source and hatch shall be isolated and held as negative controls. (i) Three or four days postvaccination, 10 of the vaccinates, the 10 positive controls, and 10 of the negative controls shall be necropsied and examined for gross lesions of bursal disease. If any of the vaccinates have such lesions, the Master Seed Virus is unsatisfactory, except that, if any of the negative controls or less than 8 of the positive controls have such lesions, the test is a No Test and may be repeated. For purposes of this test, gross lesions shall include peri bursal edema and/or edema and/or mac roscopic hemorrhage in the bursal tis sue. (ii) Fourteen days post-vaccination, the remaining vaccinates and negative controls shall be necropsied and exam ined for obvious bursal atrophy. If any of the vaccinates have such atrophy, the Master Seed Virus is unsatisfac tory, except that, if any of the negative controls have such atrophy, the test is a No Test and may be repeated. (c) Each lot of Master Seed Virus shall be tested for immunogenicity and the selected virus dose to be used shall be established as follows: (1) Bursal Disease susceptible chick ens, all of the same age (3 weeks or younger) and from the same source, shall be used. Twenty or more chickens shall be used as vaccinates for each method of administration rec ommended on the label. Ten additional chickens of the same age and from the same source shall be held as unvaccinated controls. (2) A geometric mean titer of the vac cine produced from the highest passage of the Master Seed Virus shall be es tablished before the immunogenicity VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00810 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 801 Animal and Plant Health Inspection Service, USDA § 113.332 test is conducted. Each vaccinate shall receive a predetermined quantity of vaccine virus. Five replicate virus ti trations shall be conducted on an ali quot of the vaccine virus to confirm the amount of virus administered to each chicken used in the test. At least three appropriate (not to exceed ten fold) dilutions shall be used to conduct the titrations by a method acceptable to Animal and Plant Health Inspection Service. (3) When the test chickens are 28 to 35 days of age but not less than 14 days postvaccination, each vaccinate and each control shall be challenged by eye-drop with a virulent bursal disease virus provided or approved by Animal and Plant Health Inspection Service. (i) Three to five days postchallenge, all vaccinates and controls shall be necropsied and examined for gross le sions of bursal disease as described in paragraph (b)(2)(i) of this section. (ii) If at least 19 of 20, or 27 of 30, or 36 of 40 vaccinates in each group are not free from such lesions, the Master Seed Virus is unsatisfactory, except that, if less than 90 percent of the con trols have such lesions, the test is a No Test and may be repeated. (4) An Outline of Production change shall be made before authority for use of a new lot of Master Seed Virus shall be granted by Animal and Plant Health Inspection Service. (d) After a lot of Master Seed Virus has been established as prescribed in paragraphs (a), (b), and (c) of this sec tion, each serial and subserial shall meet the applicable requirements in § 113.300 and the requirements pre scribed in this paragraph. (1) Tests for pathogens. Final con tainer samples from each serial shall be tested for pathogens by the chicken embryo inoculation test prescribed in § 113.37, except that, if the test is a No Test because of a vaccine virus over ride, the chicken inoculation test pre scribed in § 113.36 may be conducted and the serial judged accordingly. (2) Safety tests. (i) Final container samples of completed product from each serial shall be tested to determine whether the vaccine is safe as follows: (A) For vaccines intended for paren teral administration, each of twenty five 1-day-old bursal disease suscep tible chickens shall be vaccinated with the equivalent of 10 doses by subcuta neous injection. (B) For vaccines intended for drink ing water administration, each of twenty-five 4- to 5-week-old bursal dis ease susceptible chickens shall be vac cinated orally with the equivalent of 10 doses. (C) Ten chickens of the same source and hatch shall be maintained in isola tion as negative controls. The vac cinates and controls shall be observed each day for 21 days. (ii) If unfavorable reactions which are attributable to the biological prod uct occur during the observation pe riod, the serial is unsatisfactory. If un favorable reactions occur in more than one of the controls or if unfavorable re actions which are not attributable to the biological product occur in more than two of the vaccinates, the test shall be declared a No Test and re peated, except that, if the test is not repeated, the serial shall be unsatisfac tory. (3) Virus titer requirements. Final con tainer samples of completed product shall be tested for virus titer using the titration method used in paragraph (c)(2) of this section. To be eligible for release, each serial and each subserial shall have a virus titer sufficiently greater than the titer of vaccine virus used in the immunogenicity test pre scribed in paragraph (c) of this section to assure that when tested at any time within the expiration period, each se rial and subserial shall have a virus titer of 10 0.7 times greater than that used in such immunogenicity test, but not less than 10 2.0 titration units (PFU or ID50’s) per dose. [44 FR 60263, Oct. 19, 1979, as amended at 44 FR 67087, Nov. 23, 1979; 48 FR 33473, July 22, 1983. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66784, 66786, Dec. 26, 1991; 64 FR 43045, Aug. 9, 1999; 72 FR 72564, Dec. 21, 2007] § 113.332 Tenosynovitis Vaccine. Tenosynovitis Vaccine shall be pre pared from virus-bearing cell culture fluids or embryonated chicken eggs. Only Master Seed which has been es tablished as pure, safe, and immunogenic shall be used for pre paring seeds for vaccine production. All VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00811 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 802 9 CFR Ch. I (1–1–18 Edition) § 113.332 serials of vaccine shall be prepared from the first through the fifth passage from the Master Seed. (a) The Master Seed shall meet the applicable general requirements pre scribed in § 113.300, except (a)(3)(ii) and (c), and the special requirements in this section. (b) Each lot of Master Seed shall be tested for: (1) Pathogens by the chicken inocula tion test prescribed in § 113.36. (2) Lymphoid leukosis virus contami nation as follows: (i) Each of at least 10 3-week-old or older lymphoid leukosis free chickens from the same source and hatch shall be injected intra-muscularly with an amount of Master Seed equal to 100 label doses of vaccine. At least 15 chickens of the same source and hatch shall be used as controls; 5 or more shall be unvaccinated and serve as neg ative controls; 5 or more shall be in jected with subgroup A lymphoid leu kosis virus; and 5 or more with sub group B lymphoid leukosis virus. Each group of control chickens shall be held isolated from each other and from the vaccinates. (ii) Twenty-one to 28 days postinoculation, blood samples shall be taken from each chicken and the serum separated using a technique conducive to virus preservation. These serums shall be used as inocula in the com plement fixation for avian lymphoid leukosis (COFAL) test prescribed in § 113.31. (iii) Serums from the vaccinates shall be tested separately, but serums within each control group may be pooled. A valid test shall have positive COFAL reactions from each virus in oculated group and negative reactions from the uninoculated controls. If any of the chickens injected with the Mas ter Seed have positive COFAL test re actions in a valid test, the Master Seed is unsatisfactory. (3) Identity using the following agar gel immunodiffusion test. The undi luted Master Seed may be used as test antigen or the Master Seed may be in oculated onto the chorioallantoic membrane (CAM) of fully susceptible chicken embryos and the infected CAMs ground and used as antigen. A known tenosynovitis antiserum and a known tenosynovitis antigen shall be used in the test. A precipitin line shall form between the test antigen and the known antiserum in the center well which shows identity with the line formed between the antiserum and the known antigen, or the Master Seed is unsatisfactory. (4) Safety using the following chick en test: (i) For vaccines intended for use in chickens less than 14 days of age, Mas ter Seed equal to 10 label doses shall be administered subcutaneously to each of 25 1-day-old tenosynovitis susceptible chickens. (ii) For vaccines intended for use only in chickens 14 days of age or older, Master Seed equal to 10 label doses shall be administered subcutaneously to each of 25 4-week-old or older tenosynovitis susceptible chickens. (iii) The vaccinates shall be observed each day for 21 days. If unfavorable re actions occur which are attributable to the vaccine, the Master Seed is unsat isfactory. If unfavorable reactions occur which are not attributable to the vaccine, the test is a No Test and may be repeated. (c) Each lot of Master Seed shall be tested for immunogenicity. The se lected virus dose shall be established as follows: (1) Tenosynovitis susceptible chick ens, of the same age and from the same source shall be used as test birds. Vac cines intended for use in very young chickens shall be administered to chickens of the youngest age for which the vaccine is recommended. Vaccines intended for use in older chickens shall be administered to 4-week-old or older chickens. Twenty or more vaccinates shall be used for each method of admin istration recommended on the label. Ten or more chickens shall be held as unvaccinated controls. (2) A geometric mean titer of the vac cine produced at the highest passage from the Master Seed shall be estab lished using a method acceptable to Animal and Plant Health Inspection Service before the immunogenicity test is conducted. A predetermined VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00812 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 803 Animal and Plant Health Inspection Service, USDA § 113.406 quantity of vaccine virus shall be ad ministered to each vaccinate. Five rep licate virus titrations shall be con ducted on an aliquot of the vaccine virus to confirm the dose. (3) Twenty-one to 28 days postvaccination, each vaccinate and control shall be challenged by injecting virulent virus furnished or approved by Animal and Plant Health Inspection Service into one foot pad. The vac cinates and controls shall be observed each day for 14 days. If at least 90 per cent of the controls do not develop swelling and discoloration in the pha langeal joint area of the injected foot pad typical of infection with tenosynovitis virus, the test is a No Test and may be repeated. If at least 19 of 20, 27 of 30, or 36 of 40 vaccinates do not remain free from these signs, dis regarding transient swelling which sub sides within 5 days postchallenge, the Master Seed is unsatisfactory. (4) An Outline of Production change shall be made before authority for use of a new lot of Master Seed shall be granted by Animal and Plant Health Inspection Service. (d) Test requirements for release. Each serial and subserial shall meet the ap plicable general requirements pre scribed in § 113.300, except (c), and the requirements in this paragraph. (1) Purity. Final container samples of completed product from each serial shall be tested for pathogens by the chicken inoculation test prescribed in § 113.36. (2) Safety. (i) Final container samples of completed product from each serial shall be safety tested as follows: (A) For vaccines intended for use in very young chickens, each of 25 1-day old tenosynovitis susceptible chickens shall be vaccinated with the equivalent of 10 doses by one method rec ommended on the label. (B) For vaccines intended for use in older chickens, each of 25 4-week-old or older tenosynovitis susceptible chick ens shall be vaccinated with the equiv alent of 10 doses by one method rec ommended on the label. (ii) The vaccinates shall be observed each day for 21 days. If unfavorable re actions occur which are attributable to the product, the serial is unsatisfac tory. If unfavorable reactions occur in more than two vaccinates which are not attributable to the product, the test is a No Test and may be repeated. If the test is not repeated, the serial is unsatisfactory. (3) Virus titer requirements. Final con tainer samples of completed product shall be titrated by the method used in paragraph (c)(2) of this section. To be eligible for release, each serial and sub serial shall have a virus titer suffi ciently greater than the titer of the vaccine virus used in the immunogenicity test prescribed in paragraph (c) of this section to assure that, when tested at any time within the expiration period, each serial and subserial shall have a virus titer 10 0.7 times greater than that used in the immunogenicity test, but not less than 10 2.0 titration units (PFU or ID 50) per dose. (4) Identity. Bulk or final container samples of completed product from each serial shall be tested for identity as prescribed in paragraph (b)(3) of this section and shall meet the criteria stated therein. [50 FR 438, Jan. 4, 1985. Redesignated at 55 FR 35562, Aug. 31, 1990, as amended at 56 FR 66784, 66786, Dec. 26, 1991; 64 FR 43045, Aug. 9, 1999; 72 FR 72564, Dec. 21, 2007] DIAGNOSTICS AND REAGENTS §§ 113.400–113.405 [Reserved] § 113.406 Tuberculin, Intradermic. Tuberculin, Intradermic, is a filtrate produced from cultures of Pn, C, and Dt strains of Mycobacterium tuberculosis (supplied by Animal and Plant Health Inspection Service) which has been in activated and is non-toxic. Each serial shall be tested for purity, safety, po tency, and special chemical tests in ac cordance with the conditions pre scribed for each test. A serial found un satisfactory by any prescribed test shall not be released. (a) Purity test. Each serial shall be tested for purity as provided in this paragraph. (1) Final container samples of com pleted product shall be tested for via ble bacteria and fungi as prescribed in § 113.26. (2) A 20 ml sample shall be centrifuged and the sediment examined VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00813 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 804 9 CFR Ch. I (1–1–18 Edition) § 113.406 microscopically for the presence of acidfast (Ziehl-Nielsen stain) or other microorganisms (Gram stain). A serial which contains microorganisms is un satisfactory for release. (b) Safety test. Final container sam ples of completed product from each se rial shall be tested for safety. Two ma ture guinea pigs shall be injected subcutaneously with 1 ml and observed for 10 days. If unfavorable reactions at tributable to the product occur during the observation period, the serial is un satisfactory. If unfavorable reactions occur which are not attributable to the product, the test shall be declared a No Test and repeated: Provided, That if the test is not repeated, the serial shall be declared unsatisfactory. (c) Potency test. Bulk or final con tainer samples of completed product from each serial shall be subjected to a comparison test using a Reference Tu berculin supplied by Animal and Plant Health Inspection Service. Test ani mals shall be 10 sensitized white female guinea pigs from one source which weigh 500–700 grams at the beginning of the test and which have not been used in a previous test. The comparison test shall be conducted in accordance with the procedures prescribed in para graphs (c)(1), (2), (3), (4), (5), (6), (7), and (8) of this section. (1) The guinea pigs shall be sensitized with a sterile heat-killed suspension of equal amounts of strains Pn, C, and Dt of Mycobacterium tuberculosis. The heat killed sensitizing agent shall be in jected in a volume of 0.5 ml per guinea pig. The guinea pigs shall be considered sensitized for testing not less than 30 days nor more than 120 days post-injec tion. (2) The guinea pigs shall be prepared for sensitivity testing at least 4 hours prior to the injection of tuberculin. The entire abdominal and flank areas shall be clipped, a depilatory agent ap plied for 5–10 minutes, the area rinsed with warm water, and dried. (3) Dilutions of 1:100, 1:200, and 1:400 shall be prepared with the Reference Tuberculin and the unknown tuber culin. Three test sites on each side of and equidistant from the abdominal midline shall be chosen on each guinea pig. Using a tuberculin syringe and needle, 0.05 ml of each dilution shall be injected intradermally at one of the test sites which has been randomly se lected for the dilution. (4) The sensitivity of the tuberculins shall be determined 24 hours after in jected by measuring the area of ery thema. Measurements in millimeters shall be made anterior of the greatest diameter and perpendicular to the first measurement. The square millimeter shall be calculated by multiplying the two measurements. (5) The total area of response for each tuberculin tested shall be determined by adding the areas of erythema for each dilution of each of the test ani mals in a group. The sums of the areas of erythema for all three dilutions of each tuberculin shall be added to give the total area of tuberculin response. (6) The total tuberculin response area of the serial being tested shall be ex pressed as a percentage of the total tu berculin response area of the Reference Tuberculin. (The total response area of the serial divided by the total response area of the Reference Tuberculin times 100.) (7) If the total tuberculin response area of the serial being tested does not fall between 75 percent and 125 percent of the total tuberculin response area of the Reference Tuberculin, the serial is unsatisfactory. (8) Two unsensitized guinea pigs are given 0.05 ml intradermal injections of 1:4 and 1:10 dilutions of both the serial being tested and the Reference Tuber culin as a control for nonspecific posi tive reactions. If positive reactions are observed with the Reference Tuber culin, the test is considered a ‘‘No Test’’ and repeated. If positive reac tions are observed with the serial being tested only, the serial is unsatisfac tory. (d) Special chemical tests and require ments. Final container samples of com pleted product from each serial shall be tested as follows: (1) Hydrogen ion concentration. The hydrogen ion concentration shall be de termined with a pH meter which has been standardized with a pH 7.0 buffer just prior to use. The pH of the product shall be 7.0 ±0.3. (2) Total nitrogen determination. The nitrogen content shall be determined by the Kjeldahl method on duplicate 15 VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00814 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 805 Animal and Plant Health Inspection Service, USDA § 113.407 ml samples consisting of 5 ml from each of three vials. The total nitrogen content of the product shall be 0.18 per cent ±0.06 percent. (3) Trichloroacetic acid precipitable ni trogen. The determination of precipi table nitrogen by a final concentration of 4 percent trichloroacetic acid shall be made by the Kjeldahl method on du plicate 15 ml samples, consisting of 5 ml from each of three vials. The trichloroacetic acid precipitable nitro gen content shall be 0.047 percent ±0.01 percent. (4) Phenol determination. The phenol content shall be determined by direct titration with a standardized bromide bromate solution. (A correction factor of 0.04 should be subtracted from the final value in the determination of phe nol in tuberculin.) The phenol content shall be 0.54 percent ±0.04 percent. (5) Clarity. The product shall be opti cally clear and free from any extra neous particles. [39 FR 16857, May 10, 1974. Redesignated at 39 FR 25463, July 11, 1974. Redesignated at 55 FR 35561, Aug. 31, 1990, as amended at 56 FR 66784, Dec. 26, 1991] § 113.407 Pullorum antigen. Pullorum Antigen shall be produced from a culture of representative strains of Salmonella pullorum which are of known antigenic composition, high agglutinability, but are not sensitive to negative and nonspecific serum. Each serial shall be tested for purity, density, preservative content, sensi tivity, homogeneity, and hydrogen ion concentration. A serial found unsatis factory by any prescribed test shall not be released. (a) Purity test. Final container sam ples of completed product shall be test ed for viable bacteria and fungi as pre scribed in § 113.26. In addition, each se rial shall be free from extraneous orga nisms as determined by Gram staining and microscopic examination. (b) Nephelometric determination of bac terial density. The bacterial density shall be 80 ±15 times McFarland No. 1 standard for stained antigen K’s and 50 ±10 times McFarland No. 1 standard for tube antigen. (c) Preservative requirements. (1) The formalin content of Pullorum Stained Antigen K shall be 1.0 ±0.2 percent as determined by a colorimetric method. (2) The phenol content for Pullorum Tube Antigen shall be 0.55 ±0.05 percent as determined by direct titration with a standardized bromide-bromate solu tion. (d) Sensitivity requirements. (1) Each serial of antigen shall be compared with a reference antigen of known sen sitivity using positive and negative chicken serum. The manufacturers’ recommendations for use on the ac companying label or package insert shall be followed. The recommended time limit specified for each antigen shall be carefully observed in the test. (2) A total of at least 12 serums shall be used. This shall include at least three definitely positive, at least three weakly positive, and at least six nega tive serums. At least three positive chicken serums diluted with negative chicken serum shall be used to further assay comparative sensitivity between test and reference plate antigens. All test antigens shall agree closely with the reference antigen. Tests in which variation of readings between the ref erence and test antigen would result in a different National Poultry Improve ment Plan classification shall be re garded as unsatisfactory. No unsatis factory tests among the six or more negative serums and not more than one unsatisfactory test among the six or more positive serums shall be per mitted. All tests performed shall be in cluded for evaluation of the sensitivity assay. In the event of an unsatisfactory test using positive serums, at least three additional definitely positive and three additional weakly positive se rums shall be tested. If not more than one unsatisfactory test is obtained with the additional serums, the anti gen shall be acceptable. (e) Homogeneity requirement. Antigens shall show no evidence of autoagglutination or unusual appear ance such as the presence of flakes, specks, or a preponderance of filament forms. Microscopic examination shall be made in this determination. (f) Hydrogen ion concentration. The hydrogen ion concentration shall be de termined with a pH meter which has been standardized with a pH 4.0 buffer just prior to use. The pH of Pullorum VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00815 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 806 9 CFR Ch. I (1–1–18 Edition) § 113.408 Stained Antigen K shall be 4.6 ±0.4. No pH level is specified for Pullorum Tube Antigen but after dilution as rec ommended for use, it shall have a pH of 8.2 to 8.5. [39 FR 16857, May 10, 1974. Redesignated at 39 FR 25463, July 11, 1974, and amended at 40 FR 760, Jan. 3, 1975. Redesignated at 55 FR 35561, Aug. 31, 1990] § 113.408 Avian mycoplasma antigen. Mycoplasma antigens shall be pre pared from organisms, grown in broth cultures, that are inactivated and standardized. Plate antigens shall be stained with a dye acceptable to Ani mal and Plant Health Inspection Serv ice (APHIS). Final container samples of completed product from each serial shall be tested for density, preservative content, homogeneity, hydrogen ion concentration, purity, sensitivity, and specificity in accordance with the con ditions prescribed for each test. A se rial found unsatisfactory by any pre scribed test shall not be released. (a) Density requirements. A 2.5 ml sam ple of completed antigen shall be di luted with 2.5 ml of buffer solution for mulated in the same manner as the ve hicle of the antigen being tested in a modified Hopkins tube and then sedimented at 1,000 × g in a refrigerated centrifuge at 20 °C for 90 minutes. If the packed cell volume of the com pleted antigen is not 1.2 percent (±0.4 percent), the serial is unsatisfactory. (b) Preservative requirements. Preserv atives shall be as specified in the Out line of Production filed with APHIS in accordance with 9 CFR 114.8. If phenol is used, a direct titration with a stand ardized bromide-bromate solution shall be made. If the final concentration of phenol is not 0.25 percent (±0.05 per cent), the serial is unsatisfactory. (c) Homogeneity requirements. (1) Plate antigen shall be checked on a plate for homogeneity and autoagglutination. If plate antigen is not homogeneous and free of large visible particles (strands or clumps) or if it autoagglutinates, the serial is unsatisfactory. (2) Stereo-microscopic examination shall be used when necessary to evalu ate a granular appearing antigen. (d) Hydrogen ion concentration. The hydrogen ion concentration shall be de termined with a pH meter which has been standardized with a pH buffer just prior to use. The pH of Mycoplasma Gallisepticum Antigen shall be 6.0±0.2. The pH of Mycoplasma Synoviae Anti gen and Mycoplasma Meleagridis Anti gen shall be 7.0±0.2. (e) Purity requirements. The antigen shall be tested for viable bacteria and fungi as prescribed in § 113.26. (f) Sensitivity requirements. The reac tivity of each antigen shall be tested by comparing the agglutination reac tions of each serial of antigen with the agglutination reactions of a standard reference antigen which is supplied by or acceptable to APHIS. A set con sisting of five known positive and five known negative serums shall be used. The negative serums shall be tested against the antigens undiluted and the positive serums shall be tested against the antigens diluted 1:4 in buffer solu tion formulated in the same manner as the vehicle of the antigen being tested. If negative serums do not have nega tive reactions in this test, the serial is unsatisfactory. If the test antigen and the reference antigen do not have the same agglutination reactions with at least four of the five positive serums used, the serial is unsatisfactory. (1) The sensitivity of Mycoplasma Gallisepticum Antigen shall be tested using a set of chicken and a set of tur key serums (the positive serums shall have varying degrees of reactivity from weakly positive to strongly positive). (2) The sensitivity of Mycoplasma Synoviae Antigen shall be tested using chicken serums. (3) The sensitivity of Mycoplasma Meleagridis Antigen shall be tested using turkey serums. (g) Specificity requirements. Myco plasma Synoviae Antigen shall be ex amined for cross-agglutination with five Mycoplasma gallisepticum antiserums (chicken origin); Myco plasma Meleagridis Antigen shall be examined for cross-agglutination with five Mycoplasma gallisepticum antiserums (turkey origin) and five Mycoplasma synoviae antiserums (tur key origin). Tests shall be conducted VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00816 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 807 Animal and Plant Health Inspection Service, USDA § 113.409 with undiluted antigen. If cross-agglu tination occurs, the serial is unsatis factory. [48 FR 33474, July 22, 1983. Redesignated at 55 FR 35561, Aug. 31, 1990, as amended at 56 FR 66784, Dec. 26, 1991] § 113.409 Tuberculin—PPD Bovis, Intradermic. Tuberculin—PPD Bovis, Intradermic is a purified protein derivative pro duced from cultures of Mycobacterium bovis Strain AN–5 (supplied by Animal and Plant Health Inspection Service), which has been inactivated and is nontoxic. Each serial shall be tested for purity, safety, potency, and special chemical characteristics in accordance with the conditions prescribed for each test. A serial found unsatisfactory by any prescribed test shall not be re leased. (a) Purity test. Each serial shall be tested for viable bacteria and fungi as prescribed in § 113.26. (b) Safety test. Final container sam ples of completed product from each se rial shall be tested for safety as pre scribed in § 113.38. (c) Potency test. Bulk or final con tainer samples of completed product from each serial shall be subjected to a comparison specificity test using a Reference PPD Tuberculin supplied by Animal and Plant Health Inspection Service. (1) Test animals. White female guinea pigs from one source, which weigh 500 to 700 grams at the beginning of the test, and which have not been used in a previous test, shall be used in the spec ificity test. Twenty-three guinea pigs (10 sensitized with M. bovis, 10 sen sitized with M. avium and three unsensitized) shall be required for each serial being tested, and 20 guinea pigs (10 sensitized with M. bovis and 10 sen sitized with M. avium) shall be required for the Reference PPD Tuberculin. Al lowance should be made for deaths dur ing the sensitization period. (2) Sensitization of guinea pigs. (i) Sen sitize one group of guinea pigs to M. bovis. Inject each animal intramuscularly with 0.5 ml of a sterile heat-killed suspension of M. bovis Strain AN–5 supplied by Animal and Plant Health Inspection Service. (ii) Sensitize one group of guinea pigs to M. avium. Inject each animal intramuscularly with 0.5 ml of a sterile heat-killed suspension of M. avium Strain D–4 supplied by Animal and Plant Health Inspection Service. (iii) Maintain an unsensitized group as control animals. (3) Thirty-five days post-injection, the guinea pigs shall be used for tuber culin testing. (4) The sensitized animals and con trols shall be prepared at least 4 hours prior to injection of PPD tuberculin by clipping the hair from the entire ab dominal and flank areas, applying a de pilatory agent for 5 to 10 minutes, then rinsing with warm water and drying. (i) Select four sites on each guinea pig for injection of PPD tuberculin. Two sites shall be on each side of the midline and spaced a sufficient dis tance from each other to avoid overlap ping of skin reactions. (ii) Prepare four dilutions of the Ref erence PPD Tuberculin and each serial of PPD tuberculin being tested so as to contain 0.6, 1.2, 2.4, and 4.8 micrograms of protein per 0.1 ml dose. Each of the four dilutions of the same tuberculin shall be randomly assigned a site on a guinea pig. (iii) Inject one dose of each dilution at the assigned site using a tuberculin syringe. (5) Measurement of skin reactions. Measure the area of erythema produced at each site on each guinea pig 24 hours following injection of PPD tuberculin. Measurements in millimeters shall be made anterior to posterior across the greatest diameter and perpendicular to the first measurement. Calculate the area of erythema in square millimeters at each site by multiplying the two measurements. (6) Calculation of average response per guinea pig. Obtain the total area of ery thema for each guinea pig by adding the areas of the four test sites. Add these composite areas of erythema from all guinea pigs with the same sen sitization and the same PPD tuberculin injection, then divide by the number of animals in the group. The number ob tained is the average response per guin ea pig to the PPD tuberculin for the given type of sensitization. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00817 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 808 9 CFR Ch. I (1–1–18 Edition) § 113.450 (7) Determination of specificity index. The specificity index of a PPD tuber culin is determined by subtracting the average response obtained on M. avium sensitized guinea pigs from the average response obtained on M. bovis sen sitized guinea pigs. (8) Validity of bioassay. The bioassay test results obtained on serials tested concurrently in a single test series are valid if the specificity index of the ref erence PPD tuberculin is at least 400 square millimeters. If the results are not valid, the bioassay test series must be repeated with a different set of sen sitized guinea pigs. (9) Reactions in unsensitized guinea pigs. If a positive reaction (erythema) is observed in one or more of the 3 unsensitized guinea pigs, the serial is unsatisfactory. (10) Interpretation of specificity index. When a bioassay is valid and reactions are not observed in unsensitized guinea pigs, the following interpretation of the specificity index will be used for classifying each serial of PPD tuber culin: Specificity index Classification 440 mm 2 or greater ............................... Satisfactory. Between 360 mm 2 and 440 mm 2 ......... Inconclusive. Less than 360 mm 2 ............................... Unsatisfactory. (11) Second stage test. If a serial is classified as inconclusive, it can be de clared unsatisfactory or undergo a sec ond stage test. The second stage shall be conducted in a manner identical to the first stage, except that unsensitized guinea pig controls are not necessary. The results are evalu ated by combining the results obtained on all guinea pigs tested in stages one and two. Calculate the average re sponse on the 20 M. bovis sensitized ani mals and on the 20 M. avium sensitized animals and determine the specificity index. An inconclusive serial is satis factory after the second stage test, if its specificity index is 400 square milli meters or more, and unsatisfactory if its specificity index is less than 400 square millimeters. (d) Special chemical tests and require ments. Final container samples of com pleted product from each serial shall be tested as follows: (1) Protein concentration. The final product shall contain a protein con centration of 1.0 ±0.1 mg/ml. The Micro kjeldahl Test for Nitrogen shall be used. (2) Phenol content. Phenol content of the final product shall be 0.50 percent plus or minus 0.04 percent. A direct ti tration with a standardized bromide bromate solution shall be conducted. [41 FR 8471, Feb. 27, 1976, as amended at 41 FR 21760, May 28, 1976; 41 FR 32883, Aug. 6, 1976. Redesignated at 55 FR 35561, Aug. 31, 1990, as amended at 56 FR 66784, Dec. 26, 1991] ANTIBODY PRODUCTS § 113.450 General requirements for antibody products. Unless otherwise prescribed in a Standard Requirement or in a filed Outline of Production, all antibody products shall meet the applicable re quirements of this section. (a) Terminology. The following terms in the regulations and standards con cerning antibody products shall mean: Antibody. An immunoglobulin mol ecule, having a precise glycoprotein structure, produced by certain cells of the B lymphocyte lineage in response to antigenic stimulation, and func tioning to specifically bind and influ ence the antigens that induced its syn thesis. IgG (Immunoglobulin G). One of the several recognized classes of struc turally related glycoproteins whose representatives include all known anti bodies. Monoclonal. Produced by, or derived from, the offspring of a single common progenitor cell. Failure of passive transfer. A condition of neonates characterized by an abnor mally low concentration of circulating maternal IgG. (b) Nomenclature. Antibody products shall be named as follows: (1) Virus-specific products. The true name of a virus-specific product shall: include the term ‘‘antibody,’’ specify the disease for which the product is in tended, and indicate the type of animal that supplied the component anti bodies. If the antibodies are monoclonal, the term ‘‘monoclonal’’ shall be used. Example: ‘‘Duck Virus Hepatitis Antibody, Duck Origin.’’ VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00818 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 809 Animal and Plant Health Inspection Service, USDA § 113.450 (2) Bacterium-specific products. The true name of a bacterium-specific prod uct shall: include the term ‘‘antibody’’ if the component antibodies are di rected against a nontoxin antigen or the term ‘‘antitoxin’’ if the component antibodies are directed against toxin, specify the organism against which the product is intended, and indicate the type of animal that supplied the com ponent antibodies. If the antibodies are monoclonal, the term ‘‘monoclonal’’ shall be used. Example: ‘‘Escherichia Coli Monoclonal Antibody, Murine Ori gin.’’ (3) Failure of passive transfer products. The true name of a product for treat ment of failure of passive transfer shall include the term ‘‘IgG’’ and indicate the type of animal that supplied the component IgG. Example: ‘‘Bovine IgG.’’ (4) Combination products. The true name of a product for treatment of fail ure of passive transfer as well as for the prevention and/or alleviation of a specific viral or bacterial disease shall be named according to the nomen clature prescribed above for virus-spe cific or bacterium-specific products. (c) Animals. All animals used in the production of antibody products shall be healthy. Their health status shall be determined by physical examination by, or under the direct supervision of, a licensed veterinarian and by tests for infectious diseases. Such animals shall be maintained at licensed establish ments: Provided, That cows maintained at Grade A dairies (or the equivalent) that are not injected with antigens for the purpose of stimulating the produc tion of specific antibodies and that are used only for the purpose of supplying lacteal secretions are exempt from being maintained at a licensed estab lishment. (1) No animal shall be used while showing clinical signs of disease. The presence of minor localized injuries or lesions (contusions, lacerations, burns, etc.) without body temperature ele vation and without significant pain and distress shall not be construed as clinical evidence of disease. (2) Before first use and on a regular basis, all animals used in the manufac ture of antibody products shall be indi vidually subjected to applicable tests for infectious diseases. Records of all test results shall be maintained. An animal which tests positive for an in fectious disease shall not be used in the manufacture of antibody products. Retests shall be conducted as deemed necessary by the Administrator. (i) Before first use, horses shall be tested as follows for: (A) Equine infectious anemia (EIA) at a laboratory approved by APHIS. (B) Piroplasmosis, dourine, and glan ders at the National Veterinary Serv ices Laboratories. (C) Brucellosis at a laboratory ap proved by APHIS. Horses with standard agglutination titers of 1:50 or less can be used for production. Horses with standard agglutination titers equal to or greater than 1:100 may be tested by the Rivanol or card tests. Reactors to these supplemental tests shall not be used for production. Nonreactors to the supplemental tests shall be retested after 30 days. If the supplemental tests are negative and the agglutination titer has not increased, the animal may be used for production. Otherwise, the animal is unsatisfactory for this purpose. (ii) Horses shall be retested annually for EIA and, if housed or pastured with any other species, shall be retested an nually for brucellosis. (iii) Before first use, cattle shall be tested as follows for: (A) Tuberculosis by an accredited veterinarian: Provided, That cattle at Grade A dairies supplying only lacteal secretions need only be tested for tu berculosis in accordance with applica ble Milk Ordinances or similar laws or regulations. (B) Brucellosis at a laboratory ap proved by APHIS. Cattle with standard agglutination titers of 1:50 or less can be used for production. Cattle with standard agglutination titers equal to or greater than 1:100 may be tested by the Rivanol or card tests. Reactors to these supplemental tests shall not be used for production. Nonreactors to the supplemental tests shall be retested after 30 days. If the supplemental tests are negative and the agglutination titer has not increased, the animal may be used for production; otherwise, the animal is unsatisfactory for this VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00819 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 810 9 CFR Ch. I (1–1–18 Edition) § 113.450 purpose. Cattle at Grade A dairies sup plying only lacteal secretions need not be tested individually for brucellosis if a portion of their secretions contribute to the herd milk pool tested as re quired by the brucellosis ring test. An animal of a herd testing positive by this test shall not be used in produc tion. (iv) Cattle shall be retested annually for both tuberculosis and brucellosis. Cattle at Grade A dairies supplying only lacteal secretions need only be tested for tuberculosis in accordance with applicable Milk Ordinances or similar laws or regulations. Cattle at Grade A dairies supplying only lacteal secretions need not be tested individ ually for brucellosis if a portion of their secretions contribute to the herd milk pool tested as required by the brucellosis ring test. An animal of a herd testing positive by this test shall not be used in production. (v) For other species, appropriate tests and the frequency with which they are applied shall be specified in the filed Outline of Production for the product. (vi) If a positive result is obtained on any prescribed test, the positive ani mal(s) shall be removed from the herd and the remaining animals retested. Production shall not be renewed until a negative herd test is obtained not less than 28 days following removal of the positive animal(s). (vii) Negative animals shall be main tained separate and apart from untest ed or positive animals of any species. Production animals shall not be used for any other purpose, such as testing, work, or recreation. (d) Collection procedures. Blood, lac teal secretions, and egg material shall be collected as described in the filed Outline of Production for the product. (e) Ingredient handling and processing. Blood derivatives (serum, plasma, etc.), lacteal secretions, and egg material used in the production of antibody products shall be subjected to an ap propriate procedure for the inactiva tion of potential contaminating micro organisms. The procedure shall be one of those described below and specified in the filed Outline of Production for the product: Provided, That another procedure may be substituted if dem onstrated to be at least as effective by data acceptable to APHIS and specified in the filed Outline of Production for the product. These data are expected to come from a study comparing the ef fectiveness of the established and sub stitute procedures against a satisfac tory battery of potential contami nating microorganisms. (1) Blood derivatives of equine origin shall be heated at 58.0–59.0 °C for 60 minutes, and blood derivatives of bo vine, porcine, or other origin shall be heated at 58.0–59.0 °C for 30 minutes. In lieu of heat treatment, blood deriva tives of any origin may be treated with at least 2.5 megarads of ionizing radi ation, with a maximum radiation dos age specified in the filed Outline of Production for the product. (2) Lacteal secretions shall be heated as described in paragraph (e)(1) of this section, or shall be pasteurized at ei ther 72 °C for 15 seconds or 89 °C for 1 second using appropriate equipment. In lieu of the heat treatment regimens prescribed, lacteal secretions may be treated with at least 2.5 megarads of ionizing radiation, with a maximum ra diation dosage specified in the Outline of Production for the product. (3) Egg material shall be heated at 58.0–59.0 °C for 30 minutes, or treated with at least 2.5 megarads of ionizing radiation, with a maximum radiation dosage specified in the filed Outline of Production for the product. (4) Blood derivatives, lacteal secre tions, and egg material shall not con tain preservatives at the time of heat treatment, and immediately after heat treatment shall be cooled to 7 °C or lower. (5) Licensees shall keep detailed records as to each batch treated and each serial of product prepared for marketing. Recording charts shall bear full information concerning the mate rial treated and tests made of the equipment used for treatment. (f) Preservatives. Liquid antibody products, except those immediately frozen following preparation and main tained in a frozen state until time of use, shall contain at least one preserv ative from the following list, within the range of concentration set forth: (1) Phenol 0.25 to 0.55 percent, or (2) Cresol 0.10 to 0.30 percent, and/or VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00820 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 811 Animal and Plant Health Inspection Service, USDA § 113.450 (3) Thimerosal 0.01 to 0.03 percent, or (4) Other preservative(s) specified in the filed Outline of Production for the product. (g) Antigens for hyperimmunization. If animals are hyperimmunized to gen erate antibodies for a product for the prevention and/or alleviation of a spe cific infectious disease, and a USDA-li censed veterinary biological product is not employed for this purpose, the fol lowing shall apply: (1) For each antigen, a Master Seed shall be established. (i) Bacterial Master Seeds shall be tested for purity and identity as pre scribed for live bacterial vaccines in § 113.64. (ii) Viral Master Seeds shall be tested for purity and identity as prescribed for live virus vaccines in § 113.300. (2) The maximum allowable passage level of the hyperimmunizing antigen shall be the passage level of the anti gen used to generate product shown to be efficacious and shall not exceed 10 passages from the Master Seed. (h) Purity tests. Final container sam ples of each serial and each subserial shall be tested for viable bacteria and fungi as follows: (1) Dried products for parenteral ad ministration and liquid products shall be tested as prescribed in § 113.26. (2) For dried products for oral admin istration, 10 final container samples shall be reconstituted with sterile water at the volume recommended on the label and tested for the following contaminants: (i) Coliforms. One milliliter of each re hydrated sample shall be pipetted into a 100 × 15 mm petri dish and 10–15 ml of violet red bile agar at 45–50 °C added. The plate shall be manipulated to coat its entirety with the agar-sample mix ture and allowed to stand until the mixture solidifies. The plate shall then be incubated at 35 °C for 24 hours. A positive control plate and a negative control plate shall be prepared at the same time and in the same manner as the plates containing samples of the se rial. All plates shall be examined at the end of the incubation period. If char acteristic growth is observed on the negative control plate, or no char acteristic growth is observed on the positive control plate, the test shall be considered a No Test and may be re peated. If characteristic growth is ob served on any of the 10 plates con taining samples of the serial, one retest to rule out faulty technique may be conducted on samples from 20 final containers. If characteristic growth is observed on any of the retest plates, or if a retest is not initiated within 21 days of the completion of the original test, the serial or subserial is unsatis factory. (ii) Salmonellae. One milliliter of each rehydrated sample shall be pipetted into a 100 × 15 mm petri dish and 10–15 ml of brilliant green agar at 45–50 °C added. The dish shall be manipulated to coat its entirety with the agar-sam ple mixture and allowed to stand until the mixture solidifies. The plate shall then be incubated at 35 °C for 24 hours. A positive control plate and a negative control plate shall be prepared at the same time and in the same manner as the plates containing samples of the se rial. All plates shall be examined at the end of the incubation period. If char acteristic growth is observed on the negative control plate, or no char acteristic growth is observed on the positive control plate, the test shall be considered a No Test and may be re peated. If characteristic growth is ob served on any of the 10 plates con taining samples of the serial, one retest to rule out faulty technique may be conducted on samples from 20 final containers. If characteristic growth is observed on any of the retest plates, or if a retest is not initiated within 21 days of the completion of the original test, the serial or subserial is unsatis factory. (iii) Fungi. One milliliter of each re hydrated sample shall be pipetted into a 100 × 15 mm petri dish and 10–15 ml of appropriately acidified potato dextrose agar at 45–50 °C added. The plate shall be manipulated to coat its entirety with the agar-sample mixture and al lowed to stand until the mixture solidi fies. The plate shall then be incubated at 20–25 °C for 5 days. A positive con trol plate and a negative control plate shall be prepared at the same time and in the same manner as the plates con taining samples of the serial. All plates shall be examined at the end of the in cubation period. If growth is observed VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00821 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 812 9 CFR Ch. I (1–1–18 Edition) § 113.451 on the negative control plate, or no growth is observed on the positive con trol plate, the test shall be considered a No Test and may be repeated. If growth is observed on any of the 10 plates containing samples of the serial, one retest to rule out faulty technique may be conducted on samples from 20 final containers. If growth is observed on any of the retest plates, or if a retest is not initiated within 21 days of the completion of the original test, the serial or subserial is unsatisfactory. (iv) Total bacterial count. One milli liter of each rehydrated sample, undi luted or diluted as prescribed in the Outline of Production, shall be pipetted into a 100 × 15 mm petri dish and 10–15 ml of tryptone glucose extract agar at 45–50 °C added. The plate shall be ma nipulated to coat its entirety with the agar-sample mixture and allowed to stand until the mixture solidifies. The plate shall then be incubated at 35 °C for 48 hours. A positive control plate and a negative control plate shall be prepared at the same time and in the same manner as the plates containing samples of the serial. All plates shall be examined at the end of the incuba tion period. If growth is observed on the negative control plate, or no growth is observed on the positive con trol plate, the test shall be considered a No Test and may be repeated. If the average number of bacterial colonies on the 10 plates containing samples of the serial exceeds that specified in the filed Outline of Production for the product, one retest to rule out faulty technique may be conducted on sam ples from 20 final containers. If the av erage number of bacterial colonies on the retest plates exceeds that specified in the filed Outline of Production for the product, or if a retest is not initi ated within 21 days of the completion of the original test, the serial or sub serial is unsatisfactory. (i) Safety tests. Bulk or final con tainer samples of each serial shall be tested as prescribed in § 113.33(b). Dried product shall be reconstituted as indi cated on the label and 0.5 ml injected per mouse. [61 FR 51774, Oct. 4, 1996] § 113.451 Tetanus Antitoxin. Tetanus Antitoxin is a specific anti body product containing antibodies di rected against the toxin of Clostridium tetani. Each serial shall meet the appli cable general requirements provided in § 113.450 and paragraph (a) of this sec tion, and be tested for potency as pro vided in paragraph (b) of this section. Any serial found unsatisfactory by a prescribed test shall not be released. (a) General requirements. The amount of antitoxin in a final container shall be the amount which is delivered from such container when opened and in verted until the flow stops. A grad uated volumetric cylinder which con forms to the National Institute of Standards and Technology require ments shall be used. The reading shall be made at the bottom of the meniscus. Volumes of 10 ml or less shall be re corded to the nearest 0.1 and volumes over 10 ml shall be recorded to the nearest ml. (1) All final containers of Tetanus Antitoxin shall yield not less than the labeled unitage of antitoxin through out the dating period. The minimum package size permitted for marketing in the United States shall be a 1,500 unit vial. (2) The expiration date of Tetanus Antitoxin shall be not more than 3 years after the date of a potency test which demonstrates that the recover able antitoxin from the final container provides at least 20 percent excess over the number of units claimed on the label or not more than 1 year after the date of a potency test which dem onstrates that the recoverable anti toxin from the final container provides 10 to 19 percent excess over the number of units claimed on the label. (b) Potency test. Bulk or final con tainer samples of completed product from each serial shall be assayed to calculate the units of Tetanus Anti toxin in each final container. A com parative toxin-antitoxin neutralization test shall be conducted using a stand ard antitoxin and a standard toxin. All dilutions shall be made in M/15 phos phate buffered (pH) 7.4 physiological saline with 0.2 percent gelatin. (1) One ml of the Standard Antitoxin shall be diluted before use so the final volume contains 0.1 unit per ml. The VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00822 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 813 Animal and Plant Health Inspection Service, USDA § 113.452 dilution shall be held at 20° to 25 °C for 30 minutes prior to combination with a test does of toxin. (2) The Standard Toxin test dose is that amount which when mixed with 0.1 unit of Standard Antitoxin, incu bated at 20° to 25 °C for 1 hour, and in jected subcutaneously into a 340 to 380 gram guinea pig, results in death of that guinea pig within 60 to 120 hours with clinical signs of tetanus. The toxin shall be diluted so the test dose shall be in 2.0 ml. (3) A mixture of diluted Standard Toxin and diluted Standard Antitoxin shall be made so that 0.1 unit of anti toxin in 1 ml is combined with a test dose of toxin. This Standard Toxin Antitoxin mixture shall be held at 20° to 25 °C for 1 hour before injections of guinea pigs are made. (4) A sample from each serial of anti toxin shall be prepared as was the Standard Toxin-Antitoxin mixture; ex cept the amount of antitoxin shall be based on an estimation of the expected potency. When testing is done on bulk material, the final container fill shall reflect the endpoint value plus 10 per cent overage for 1 year dating and 20 percent overage for 3 year dating. (5) Normal guinea pigs weighing within a range of 340 to 380 grams shall be used. Pregnant guinea pigs must not be used. (i) Each of two guinea pigs (controls) shall be injected subcutaneously with a 3 ml dose of the Standard Toxin-Anti toxin mixture. Injections shall be made in the same order that toxin is added to the dilutions of antitoxins. These shall be observed parallel with the ti tration of one or more unknown antitoxins. (ii) Two guinea pigs shall be used as test animals for each dilution of the unknown antitoxin. A 3.0 ml dose shall be injected subcutaneously into each animal. (6) Controls shall be observed until they are down and are unable to rise or stand under their own power. At this time they are euthanized and the time of death is recorded in hours. For a sat isfactory test, the controls must reach this point with clinical signs of tetanus within 24 hours of each other and with in an overall time of 60 to 120 hours. The clinical signs to be observed are in creased muscle tonus, curvature of the spine, asymmetry of the body outline when the resting animal is viewed from above, generalized spastic paralysis, particularly of the extensor muscles, inability to rise from a smooth surface when the animal is placed on its side, or any combination of these signs. If the control guinea pigs do not respond in this manner, the entire test shall be repeated. (7) Potency of an unknown antitoxin is determined by finding the mixture which will protect the test animal the same as the Standard Toxin-Antitoxin mixture. Test animals dying sooner than the controls indicate the unit value selected in that dilution was not present, whereas those living longer in dicate a greater unit value. [39 FR 16859, May 10, 1974. Redesignated at 39 FR 25463, July 11, 1974, and amended at 40 FR 760, Jan. 3, 1975; 40 FR 41996, Sept. 10, 1975; 43 FR 1479, Jan. 10, 1978; 50 FR 24905, June 14, 1985. Redesignated at 55 FR 35561, Aug. 31, 1990; 61 FR 51776, Oct. 4, 1996; 64 FR 43045, Aug. 9, 1999] § 113.452 Erysipelothrix Rhusiopathiae Antibody. Erysipelothrix Rhusiopathiae Anti body is a specific antibody product con taining antibodies directed against one or more somatic antigens of Erysipelothrix rhusiopathiae. Each serial shall be tested as provided in this sec tion. Any serial found unsatisfactory by a prescribed test shall not be re leased. (a) Each serial shall meet the appli cable general requirements provided in § 113.450. (b) Potency test. Bulk or final con tainer samples of completed product from each serial shall be tested using the two-stage test provided in this sec tion. (1) In the first stage, each of 40 Swiss mice, each weighing 16 to 20 grams, shall be injected subcutaneously with 0.1 ml of product (dried product shall be rehydrated according to label direc tions). Twenty-four hours postinjection, the injected mice and 10 additional mice designated controls shall be challenged subcutaneously with the same culture of Erysipelothrix rhusiopathiae. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00823 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 814 9 CFR Ch. I (1–1–18 Edition) § 113.453 (2) If less than eight of the 10 con trols die from erysipelas within 7 days post-challenge, the test is invalid. All dead mice shall be examined to deter mine if the cause of death was Erysipelothrix rhusiopathiae infection. (3) The mice injected with product shall be observed for 10 days postchallenge and all deaths recorded. The second stage shall be required when 7–10 of the mice injected with product die in the first stage. The sec ond stage shall be conducted in a man ner identical to the first stage. (4) The results of the test shall be evaluated according to the following table: Stage Number of vac-cinates Cumulative number of vac cinates Cumu lative total num ber of deaths for a satis factory test Cumu lative total num ber of deaths for an unsat isfac tory test 1 40 40 6 or less 11 or more. 2 40 80 12 or less 13 or more. [39 FR 16859, May 10, 1974. Redesignated at 39 FR 25463, July 11, 1974, as amended at 40 FR 20067, May 8, 1975; 40 FR 23989, June 4, 1975. Redesignated at 55 FR 35561, Aug. 31, 1990; 61 FR 51776, Oct. 4, 1996; 64 FR 43045, Aug. 9, 1999] § 113.453 [Reserved] § 113.454 Clostridium Perfringens Type C Antitoxin. Clostridium Perfringens Type C Anti toxin is a specific antibody product containing antibodies directed against the toxin of Clostridium perfringens Type C. Each serial shall be tested as provided in this section. Any serial found unsatisfactory by a prescribed test shall not be released. (a) Each serial shall meet the appli cable general requirements provided in § 113.450. (b) Potency test. Bulk or final con tainer samples of completed product from each serial shall be tested using the toxin-neutralization test for Beta Antitoxin provided in this section. Dried products shall be rehydrated ac cording to label directions. (1) When used in this test, the fol lowing words and terms shall mean: (i) International antitoxin unit. (I.U.) That quantity of Beta Antitoxin which reacts with L0 and L ∂ doses of Stand ard Toxin according to their defini tions. (ii) L0dose. The largest quantity of toxin which can be mixed with one unit of Standard Antitoxin and not cause sickness or death in injected mice. (iii) L ∂ dose. The smallest quantity of toxin which can be mixed with one unit of Standard Antitoxin and cause death in at least 80 percent of injected mice. (iv) Standard antitoxin. The Beta Antitoxin preparation which has been standardized as to antitoxin unitage on the basis of the International Clos tridium perfringens Beta Antitoxin Standard and which is either supplied by or acceptable to Animal and Plant Health Inspection Service. The anti toxin unit value shall be stated on the label. (v) Standard toxin. The Beta toxin preparation which is supplied by or is acceptable to Animal and Plant Health Inspection Service. (vi) Diluent. The solution used to make proper dilutions prescribed in this test. Such solution shall be made by dissolving 1 gram of peptone and 0.25 gram of sodium chloride in each 100 ml of distilled water; adjusting the pH to 7.2; autoclaving at 250 °F. for 25 min utes; and storing at 4 °C. until used. (2) The antitoxin content of the test sample shall be determined as follows: (i) Make a dilution of Standard Anti toxin to contain 10 International Units of antitoxin per ml. (ii) Make one dilution of Standard Toxin to contain 10 L0 doses per ml and make a second dilution of Standard Toxin to contain 10 L ∂ doses per ml. (iii) Dilute 1 ml of the test sample with 49 ml of diluent and combine 1 ml of this dilution with 1 ml of the Stand ard Toxin diluted to contain 10 L0 doses. (iv) Combine 10 International Units of Standard Antitoxin with 10 L0 doses of diluted Standard Toxin and combine 10 International Units of Standard Antitoxin with 10 L ∂ doses of diluted Standard Toxin. (v) Neutralize all toxin-antitoxin mixtures at room temperature for 1 VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00824 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 815 Animal and Plant Health Inspection Service, USDA § 113.455 hour and hold in ice water until injec tions of mice can be made. (vi) Five Swiss white mice, each weighing 16–20 grams, shall be used for each toxin-antitoxin mixture. A dose of 0.2 ml shall be injected intravenously into each mouse. Conclude the test 24 hours post-injection and record all deaths. (3) Test Interpretation. (i) If any mice inoculated with the mixture of 10 International Units of Standard Anti toxin and 10 L0 doses of Standard Toxin die, the results of the test are a No Test and shall be repeated: Provided, That, if the test is not repeated, the se rial shall be declared unsatisfactory. (ii) If less than 80 percent of the mice inoculated with the mixture of 10 International Units of Standard Anti toxin and 10 L ∂ doses of Standard Toxin die, the results of the test are a No Test and shall be repeated: Provided, That, if the test is not repeated, the se rial shall be declared unsatisfactory. (iii) If any mice inoculated with the mixture of Clostridium Perfringens Type C Antitoxin diluted 1:50 and 10 L0 doses of Standard Toxin die, the anti toxin is considered to contain less than 500 International Unit per ml and the serial is unsatisfactory. [39 FR 16859, May 10, 1974. Redesignated at 39 FR 25463, July 11, 1974. Redesignated at 55 FR 35561, Aug. 31, 1990, as amended at 56 FR 66784, Dec. 26, 1991; 61 FR 51777, Oct. 4, 1996] § 113.455 Clostridium Perfringens Type D Antitoxin. Clostridium Perfringens Type D Antitoxin is a specific antibody prod uct containing antibodies directed against the toxin of Clostridium perfringens Type D. Each serial shall be tested as provided in this section. Any serial found unsatisfactory by a pre scribed test shall not be released. (a) Each serial shall meet the appli cable general requirements provided in § 113.450. (b) Potency test. Bulk or final con tainer samples of completed product from each serial shall be tested using the toxin-neutralization test for Epsi lon Antitoxin provided in this section. Dried products shall be rehydrated ac cording to label directions. (1) When used in this test, the fol lowing words and terms shall mean: (i) International antitoxin unit. (I.U.) That quantity of Epsilon Antitoxin which reacts with L0 and L ∂ doses of Standard Toxin according to their defi nitions. (ii) L0dose. The largest quantity of toxin which can be mixed with one tenth unit of Standard Antitoxin and not cause sickness or death in injected mice. (iii) L ∂ dose. The smallest quantity of toxin which can be mixed with one tenth unit of Standard Antitoxin and cause death in at least 80 percent of in jected mice. (iv) Standard antitoxin. The Epsilon Antitoxin preparation which has been standardized as to antitoxin unitage on the basis of the International Clos tridium perfringens Epsilon Antitoxin Standard and which is either supplied by or acceptable to Animal and Plant Health Inspection Service. The anti toxin unit value shall be stated on the label. (v) Standard toxin. The Epsilon toxin preparation which is supplied by or is acceptable to Animal and Plant Health Inspection Service. (vi) Diluent. The solution used to make proper dilutions prescribed in this test. Such solution shall be made by dissolving 1 gram of peptone and 0.25 gram of sodium chloride in each 100 ml of distilled water; adjusting the pH to 7.2; autoclaving at 250 °F. for 25 min utes; and storing at 4 °C. until used. (2) The antitoxin content of the test sample shall be determined as follows: (i) Make a dilution of Standard Anti toxin to contain 1 International Unit of antitoxin per ml. (ii) Make one dilution of Standard Toxin to contain 10 L0 doses per ml and make a second dilution of Standard Toxin to contain 10 L ∂ doses per ml. (iii) Dilute 1 ml of the test sample with 33 ml of diluent and combine 1 ml of this dilution with 1 ml of the Stand ard Toxin diluted to contain 10 L0 doses. (iv) Combine 1 International Unit of Standard Antitoxin with 10 L0 doses of Standard Toxin and combine 1 Inter national Unit of Standard Antitoxin with 10 L ∂ doses of Standard Toxin. (v) Neutralize all toxin-antitoxin mixtures at room temperature for 1 VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00825 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 816 9 CFR Ch. I (1–1–18 Edition) §§ 113.456–113.498 hour, and hold in ice water until injec tions of mice can be made. (vi) Five Swiss white mice, each weighing 16–20 grams, shall be used for each toxin-antitoxin mixture. A dose of 0.2 ml shall be injected intravenously into each mouse. Conclude the test 24 hours post-injection and record all deaths. (3) Test Interpretation. (i) If any mice inoculated with the mixture of 1 International Unit of Standard Anti toxin and 10 L0 doses of Standard Toxin die, the results of the test are a No Test and shall be repeated: Provided, That, if the test is not repeated, the se rial shall be declared unsatisfactory. (ii) If less than 80 percent of the mice inoculated with mixture of 1 Inter national Unit of Standard Antitoxin and 10 L ∂ doses of Standard Toxin die, the results of the test are a No Test and shall be repeated: Provided, That, if the test is not repeated, the serial shall be declared unsatisfactory. (iii) If any mice inoculated with the mixture of Clostridium Perfringens Type D Antitoxin diluted 1:34 and 10 L0 doses of Standard Toxin die, the anti toxin is considered to contain less than 34 International Units per ml and the serial is unsatisfactory. [39 FR 16859, May 10, 1974. Redesignated at 39 FR 25463, July 11, 1974, as amended at 40 FR 760, Jan. 3, 1975. Redesignated at 55 FR 35561, Aug. 31, 1990, as amended at 56 FR 66784, Dec. 26, 1991; 61 FR 51777, Oct. 4, 1996] §§ 113.456–113.498 [Reserved] § 113.499 Products for treatment of failure of passive transfer. A product for the treatment of fail ure of passive transfer (FPT) shall con tain a specified minimum quantity of IgG per dose and shall be recommended for use only in neonates of the same species as that of antibody origin. A product for oral administration shall not be recommended for use in animals more than 24 hours of age, while one for parenteral administration shall only be recommended for use in neo natal animals. Each serial shall meet the applicable general requirements provided in § 113.450 and be tested for potency as provided in this section. Any serial found unsatisfactory by a prescribed test shall not be released. (a) Qualification of an IgG Reference Product. An IgG Reference Product (ref erence) shall be a serial of product that is manufactured according to the filed Outline of Production, properly quali fied, and used to assess the potency of subsequent product serials, as de scribed in paragraph (c) below. The ref erence shall be qualified as follows: (1) At least 20 newborn, colostrum-de prived animals of the species for which the product is recommended shall be randomly selected. (2) Blood samples shall be taken from each animal. (3) Each animal shall be administered one dose of reference by the rec ommended route and shall be observed for 24 hours. (i) Any adverse reactions shall be re corded. (ii) The dosage of reference adminis tered to each animal shall be in accord ance with label directions. Label direc tions may indicate a single dosage re gardless of weight, in which case the animals in the study shall be at or near the maximum weight for neonates of the species. (4) After 24 hours, blood samples shall be taken from each animal. (5) Pretreatment and post treatment serum IgG concentrations shall be con currently determined for each animal using a radial immunodiffusion (RID) method acceptable to APHIS and de scribed in the filed Outline of Produc tion for the product. (6) Concurrently, using the same method, five IgG measurements shall be made on an IgG Species Standard supplied or approved by APHIS. The IgG Species Standard shall be a prepa ration that contains IgG specific for the species in question at a concentra tion acceptable to APHIS. (7) For an IgG Reference Product to be satisfactory, all animals used to qualify the reference must remain free of unfavorable product-related reac tions and at least 90 percent of the paired serum samples must reflect an increase in IgG concentration (posttreatment minus pretreatment concentration) equal to or greater than the IgG concentration of the IgG Spe cies Standard. (b) Antibody functionality. Prior to li censure, the prospective licensee shall VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00826 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 817 Animal and Plant Health Inspection Service, USDA § 114.2 perform a neutralization study, or an other type of study acceptable to APHIS, to demonstrate functionality of product antibody. (c) Potency. Bulk or final container samples of completed product from each serial shall be tested for IgG con tent as provided in this paragraph. Samples of the test serial and of an IgG Reference Product established in ac cordance with paragraph (a) of this sec tion shall be concurrently tested for IgG content by the RID method re ferred to in paragraph (a)(5) of this sec tion. Five IgG measurements shall be made on each. If the IgG level per dose of the test serial does not meet or ex ceed that of the reference, one com plete retest, involving five IgG meas urements on both the reference and two samples of the test serial, may be conducted. If, upon retest, the average IgG level per dose of the two samples of the test serial does not meet or exceed that of the reference, or if a retest is not conducted, the serial is unsatisfac tory. [61 FR 51777, Oct. 4, 1996] PART 114—PRODUCTION REQUIRE MENTS FOR BIOLOGICAL PROD UCTS Sec. 114.1 Applicability. 114.2 Products not prepared under license. 114.3 Separation of establishments. 114.4 Identification of biological products. 114.5 Micro-organisms used as seed. 114.6 Mixing biological products. 114.7 Personnel at licensed establishments. 114.8 Outline of Production required. 114.9 Outline of Production guidelines. 114.10 Antibiotics as preservatives. 114.11 Storage and handling. 114.12 Expiration date required. 114.13 Expiration date determination. 114.14 Extension of expiration date for a se rial or subserial. 114.15 Disposal of unsatisfactory products and byproducts. 114.16 Producing subsidiaries. 114.17 Rebottling of biological products. 114.18 Reprocessing of biological products. AUTHORITY: 21 U.S.C. 151–159; 7 CFR 2.22, 2.80, and 371.4. SOURCE: 39 FR 16869, May 10, 1974, unless otherwise noted. § 114.1 Applicability. Unless exempted by regulation or otherwise authorized by the Adminis trator, all biological products prepared, sold, bartered or exchanged, shipped or delivered for shipment in or from the United States, the District of Colum bia, any Territory of the United States, or any place under the jurisdiction of the United States shall be prepared in accordance with the regulations in this part. The licensee or permittee shall adopt and enforce all necessary meas ures and shall comply with all direc tions the Administrator prescribes for carrying out such regulations. [52 FR 11026, Apr. 7, 1987, as amended at 56 FR 66784, Dec. 26, 1991] § 114.2 Products not prepared under li cense. (a) When an establishment license is issued, if biological products which were not prepared in compliance with the regulations are in the establish ment, such products shall not be shipped or delivered for shipment or otherwise dealt with as having been prepared under such regulations. (b) Except as provided in 9 CFR part 103, a biological product shall not be prepared in a licensed establishment unless the person to whom the estab lishment license is issued holds an un expired, unsuspended, and unrevoked product license issued by the Adminis trator to prepare such biological prod uct, or unless the products prepared are subject to the provisions of § 107.2 of this subchapter. (c) A biological product produced in a USDA-licensed establishment shall be produced under a U.S. Veterinary Bio logical Product License or a license granted by a State under § 107.2 (re ferred to as a State biological product license and the products prepared pur suant thereto as State-licensed biologi cal products, including autogenous bio logics), but not under both a U.S. Vet erinary Biological Product License and VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00827 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 818 9 CFR Ch. I (1–1–18 Edition) § 114.3 a State biological product license. Be fore a U.S. Veterinary Biological Prod uct License (including a conditional li cense) is issued, the licensee shall re linquish its State license for that prod uct: Provided, That autogenous bio logics shall not be subject to this pro vision when they are prepared in ac cordance with the provisions of para graph (c)(5) of this section. (1) State-licensed biological products (including autogenous biologics) shall only be distributed or shipped intra state, must not bear a U.S. Veterinary Biologics Establishment License Num ber, and must not otherwise be rep resented in any manner as having met the requirements for a U.S. Veterinary Biological Product license. Labeling of State- and USDA-licensed biological products produced in the same estab lishment must be distinctly different in color and design. (2) All biological products in USDA licensed establishments, whether li censed by USDA or by the State, shall be prepared only in locations indicated in legends filed in accordance with 9 CFR part 108. A description of each State-licensed product must be filed with the Animal and Plant Health In spection Service as part of the blue print legends and must be sufficient for Animal and Plant Health Inspection Service to determine any risk to the production of other products in the li censed establishment and to determine that adequate procedures are followed to prevent contamination during pro duction. (3) Records in such establishments must be maintained in accordance with §§ 116.1 and 116.2 of this subchapter and shall include all products licensed by the State or USDA. (4) Reports prescribed in § 116.5 of this subchapter for USDA-licensed estab lishments shall be submitted for all veterinary biological products in the establishment. (5) Under the following conditions, an autogenous biologic may be produced in a USDA-licensed establishment under either a State or U.S. Veterinary Biological Product License: (i) When a culture of microorga nisms, isolated from a herd in a State, is received at a USDA-licensed estab lishment that is in the same State but that holds both a State and a U.S. Vet erinary Biological Products License for autogenous biologics, the isolate shall be designated by the licensee for use in the production of an autogenous bio logical product under either the State product license, or the U.S. Veterinary Biological Product License: Provided, That the isolate meets the require ments of the respective regulatory au thority for an autogenous biologic. If, after producing the product pursuant to one license, the licensee elects to produce an autogenous biologic from the same isolate under provisions of the other license, the licensee may do so only with the approval of the other licensing authority. (ii) The true name of a State-licensed autogenous biologic shall specify the State of licensure: e.g. ‘‘ lllll Autogenous Bacterin’’ (State) or lllll Autogenous Vaccine’’. (State) [39 FR 16869, May 10, 1974, as amended at 60 FR 48021, Sept. 21, 1995] § 114.3 Separation of establishments. (a) Each licensed establishment shall be separate and distinct from any other establishment in which a biological product is prepared. (b) No biological products authorized to be prepared in a licensed establish ment shall be prepared in whole or in part by another licensed establishment except as provided in paragraphs (c) and (d) of this section. (c) When a partially prepared biologi cal product cannot be completed at a licensed establishment due to failure of essential equipment, the Administrator may authorize the use of similar equip ment at another licensed establish ment: Provided, That, such authoriza tion shall be limited to the duration of the emergency and to the phase of pro duction affected by the equipment fail ure. (d) Partially prepared products or se rials of completed products for further manufacture may be moved from one licensed establishment to another li censed establishment, imported under the provisions of § 104.5, or moved from a licensed establishment for purpose of VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00828 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 819 Animal and Plant Health Inspection Service, USDA § 114.8 being exported under conditions pre scribed in an Outline of Production filed with Animal and Plant Health In spection Service. Licensed products or products imported for distribution and sale may be prepared and recommended for final use, for further manufacturing purposes, or both. All serials shall be subject to the requirements for testing and release specified in § 113.5 or § 113.10 and to the requirements for identifica tion specified in § 114.4. [39 FR 16869, May 10, 1974, as amended at 40 FR 46093, Oct. 6, 1975; 49 FR 45846, Nov. 21, 1984; 56 FR 66784, Dec. 26, 1991] § 114.4 Identification of biological products. Suitable tags or labels of a distinct design shall be used for identifying all ingredients used in the preparation of biological products, all component parts to be combined to form a biologi cal product, all biological products while in the course of preparation and all completed biological products held in storage at licensed establishments: Provided, That, if such ingredients, components, or biological products are not so identified, they shall be disposed of as provided in § 114.15. § 114.5 Micro-organisms used as seed. Micro-organisms used in the prepara tion of biological products at licensed establishments shall be free from the causative agents of other diseases or conditions. A complete record of such micro-organisms shall be kept cur rently correct and a list submitted to Animal and Plant Health Inspection Service upon request of the Adminis trator. (Approved by the Office of Management and Budget under control number 0579–0059) [39 FR 16869, May 10, 1974, as amended at 48 FR 57473, Dec. 30, 1983; 56 FR 66784, Dec. 26, 1991] § 114.6 Mixing biological products. Each biological product, when in liq uid form, shall be mixed thoroughly in a single container. During bottling op erations, the product shall be con stantly mixed sufficient to maintain physical uniformity of the entire fill. A serial number, with any other mark ings that may be necessary for ready identification of the serial, shall be ap plied to identify it with the records of preparation and labeling. § 114.7 Personnel at licensed establish ments. (a) Each licensee shall designate a person(s) to make all official contacts with Animal and Plant Health Inspec tion Service on matters pertaining to the preparation of biological products under the Virus-Serum-Toxin Act. The licensee shall file three copies of bio graphical summary with Animal and Plant Health Inspection Service for such designated person and for each person responsible for any phase of preparation of a biological product. (b) All personnel employed in the preparation of biological products at a licensed establishment shall be com petent in good laboratory techniques through education or training, or both, so as to consistently prepare high qual ity products. (c) All biological products prepared at licensed establishments shall be pre pared and handled with due sanitary precautions. Good sanitary measures shall be practiced at all times by all personnel involved in such preparation and handling of biological products. (1) The clothing worn by persons while preparing biological products shall be clean. All persons, imme diately before entering laboratory rooms of a licensed establishment, shall change their outer clothing or ef fectively cover the same with gowns or other satisfactory clean garments. (2) Unsanitary practices such as, but not limited to, eating, smoking, or ex pectorating on the floors or otherwise creating a nuisance in any room, com partment, or place in which biological products are prepared, handled, or stored at licensed establishments are prohibited. (Approved by the Office of Management and Budget under control number 0579–0013) [39 FR 16869, May 10, 1974, as amended at 48 FR 57473, Dec. 30, 1983; 56 FR 66784, Dec. 26, 1991] § 114.8 Outline of Production required. An Outline of Production shall be on file with Animal and Plant Health In spection Service for each licensed bio logical product or for each biological VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00829 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 820 9 CFR Ch. I (1–1–18 Edition) § 114.9 product authorized to be imported into the United States for Distribution and Sale. Preparation of a biological prod uct in a licensed establishment shall be in accordance with the Outline of Pro duction for such product filed with Animal and Plant Health Inspection Service as provided in this section, but subject to changes as may be required under § 114.8(f). (a) The Outline of Production shall be prepared as prescribed in § 114.9 and submitted to Animal and Plant Health Inspection Service for filing. When ob jectionable features, if any, are cor rected and no further exceptions are taken by Animal and Plant Health In spection Service to an Outline of Pro duction for a biological product, such Outline of Production shall be ap proved for filing. (b) Each page shall be stamped as filed on the date such action was taken in the bottom right hand corner. Al though the filed outline may be re ferred to as an approved outline, ap proval for filing constitutes no en dorsement by Animal and Plant Health Inspection Service of such biological product or the methods and procedures used to prepare such biological prod uct. (c) One copy of the Outline of Produc tion shall be retained by the Animal and Plant Health Inspection Service and one copy returned to the licensee or permittee. (d) Each licensee shall review each Outline of Production for accuracy and sufficiency not less frequently than once a year. Revisions necessary to bring an Outline of Production into compliance with the regulations shall be submitted to Animal and Plant Health Inspection Service. (e) When a list of licensed products to be continued in production at a li censed establishment is requested by the Administrator in accordance with § 102.5(d) of this subchapter, the li censee shall supplement the list with information for each product as fol lows: (1) The Outline of Production cur rently being used shall be identified as to the date when last revised and filed with Animal and Plant Health Inspec tion Service and the date of the last re view made by the licensee. (2) The Outline of Production to be kept in the active file shall be des ignated. If more than one has been filed for a product, only the Outline of Pro duction currently being used shall be included. (f) The Administrator may, upon the basis of information not available to him at the time the current Outline of Production for a biological product was filed, object to the methods or proce dures being used in the preparation of such biological product and notify the licensee to modify the filed Outline of Production to eliminate such objec tions. If the licensee does not comply with the notice, the Administrator may, after affording opportunity for a hearing to the licensee, suspend the product license for the biological prod uct involved; in which case, the li censee shall not prepare such product until subsequent notice of withdrawal of the suspension is given to the li censee. (Approved by the Office of Management and Budget under control number 0579–0013) [39 FR 16869, May 10, 1974, as amended at 48 FR 57473, Dec. 30, 1983; 56 FR 66784, Dec. 26, 1991; 75 FR 20773, Apr. 21, 2010] § 114.9 Outline of Production guide lines. Each Outline of Production shall be prepared in accordance with the appli cable directions provided in this sec tion. (a) General requirements. (1) All copies of each Outline of Production or spe cial outline or revised pages of either shall be prepared on heavy paper (8.5″ × 11″) of a type receptive to permanent stamp ink. (2) The name of the biological prod uct (or component), the establishment license number, and the date prepared shall appear on a front cover page and each page of the Outline of Production or special outline. The name of the li censee (or foreign manufacturer) shall appear on the front cover page. (3) The pages shall be numbered in the upper center. At least 11⁄2 inch mar gin shall be left at the top of the first page and a 2 inch margin at the bottom of each page for the Animal and Plant Health Inspection Service stamp. (4) Amended pages shall be numbered the same as those being superseded. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00830 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 821 Animal and Plant Health Inspection Service, USDA § 114.9 They shall bear the date prepared and refer to the date on the pages being su perseded. If one replacement page su persedes more than one page, the new page shall indicate same, but if several replacement pages are added to super sede one page, the page number fol lowed by letters shall be used. (5) The last page of both copies of ei ther a new or a completely rewritten Outline of Production and each page revised separately shall be signed in the lower left corner by the authorized representative of the licensee (or for eign producer). Stamped or facsimile signatures are not acceptable. (6) A summary of changes shall ap pear on an attached page and refer to each page, paragraph, or subparagraph being changed. (7) Transmittal forms shall be used for the original and subsequent revi sions. Transmittal forms are available on the Internet at (http:// www.aphis.usda.gov/animallhealth/ vetlbiologics/vblforms.shtml). (b) Special outline. An outline describ ing the preparation of a component of a biological product or an operation performed in the preparation of a bio logical product may be required if such special outline could be referred to in Outlines of Production to eliminate repetition. Each special outline shall be identified by number and shall not be used until accepted and filed by Ani mal and Plant Health Inspection Serv ice. (c) Outline of Production for anti serum, antitoxin, and normal serum shall be written according to the fol lowing: OUTLINE GUIDE FOR PRODUCTION OF ANTI SERUM AND ANTITOXIN AND NORMAL SERUM License No. Name of Product Date I. Serum animals. A. Species, conditions, age, and general health. B. Examination, preparation, care, quar antine, tests, and treatment of animals be fore injections are started. C. Holding, handling, exercising, and moni toring the condition of animals after injec tions are started. II. Antigens. A. Composition and character of the antigen. 1. Micro-organisms. 2. Source and date of accession of each micro-organism. 3. Strains. 4. Proportions of each micro-organism and strain. B. Identification methods used for each micro-organism and frequency with which these methods are applied. C. Virulence and purity of cultures or anti gen and the determination and maintenance thereof. Range of subcultures or passages to be used in production. D. Attenuation, if any, before use for pro duction purposes. E. Character, size, and shape of containers used for growing micro-organisms. F. Media used for stock, seed, and antigen cultures (composition and reaction of). May refer to a special outline by number. G. Preparation of the antigen or toxin and toxoid. Complete and full description of each step and its manner of accomplishment and number these steps in sequence. Include all tests for each antigen, and the specifications for character, identity, virulence, concentra tion, and standardization. III. Immunization of animals. A. Outline fully with special attention given to the fol lowing: 1. Character and dose of the antigen. 2. Method and frequency of injections. 3. Time required for immunization or hyperimmunization. 4. Preliminary bleedings and tests, if any, to ascertain quality of serum. 5. All other similar matters, including treatments between bleedings. B. Period of time elapsing between last in jection and first bleeding; and between bleed ings. C. Technique of bleeding operations; vol ume of blood collected at each bleeding; and period of rest. IV. Preparation of the biological product. A. Describe fully and show each step of prepara tion from the first bleeding to the comple tion of the preserved product in bulk con tainers prior to filling of final containers. B. Composition of the preservative and proportions used. Indicate at which step of production, and the method used in adding the preservative. C. Agglutination and complement-fixation titers and the methods of their determina tions. D. Disposition of unsatisfactory biological products and infective materials not used in production. E. Assembly of units to make a serial; vol ume of the average serial; and the volume of the maximum serial. V. Testing. Indicate the stages in the prepa ration of the biological product at which samples are collected. Refer to all applicable Standard Requirements. Outline all addi tional tests in detail and state minimum re quirements for each satisfactory test. A. Purity. B. Safety. C. Potency. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00831 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 822 9 CFR Ch. I (1–1–18 Edition) § 114.9 D. Other tests. VI. Post preparatory steps. A. Form and size of final containers in which the product is to be distributed. B. Methods and techniques of filling final containers. Volume of fill for each size final container. C. Collection, storage, and submission of representative samples. Indicate at which steps in the production these samples are taken. D. Expiration date based on the earliest date of harvest and the date of the last satis factory potency test. E. Use, dosage, and route of administration for each animal species for which it is rec ommended. F. Include any additional pertinent infor mation. (d) Outline of Production for vaccines, bacterins, antigens, and toxoids shall be written according to the following: OUTLINE GUIDE FOR VACCINES, BACTERINS, ANTIGENS, AND TOXOIDS License No. Name of Product Date I. Composition, etc., of the product. A. Micro organisms used. Give the isolation and pas sage history. B. Source and date of accession of each micro-organism. C. Strains. D. Proportions of each strain. II. Cultures. A. Brief description of methods of identifying each micro-organism and the frequency with which these methods are ap plied. B. Virulence and purity of cultures and the determination and maintenance thereof. Range of subcultures or passages to be used in production. C. Composition and reaction of media used for seed and production cultures. Include the source of eggs, tissue, or cell cultures, and the tests to determine that eggs, tissues, and cells are free of contamination. D. Character, size, and shape of containers used for growing cultures. E. Storage conditions of seed cultures. F. Methods of preparing suspensions for seeding or inoculation. G. Technique of inoculating (1) seed media; (2) production media. Titer or concentration of inoculum, and the volume of medium for each size and type of culture container. H. Period of time and conditions for incu bation and degree of temperature used for each micro-organism or group of micro-orga nisms. I. Character and amount of growth; obser vation as to contamination of growth. J. Method of attenuation, if any, before used for production purposes. III. Harvest. A. Handling and preparation of cultures and media (including eggs) before removal of micro-organisms or tissues for production purposes. B. Minimum and maximum period of time elapsing from time of inoculation until har vest. C. Technique of harvesting micro-orga nisms or tissues (specify) for production pur poses. D. Specifications for acceptable harvest material. E. Handling of discarded material not used in production. F. Include any additional pertinent infor mation. IV. Preparation of the product. Describe fully and show each step of preparation from harvest of antigen containing tissues or pro duction cultures to the completion of the finished product in final containers. In de scribing the preparation of the product, em phasize the following: A. Method of inactivation, attenuation, or detoxification. B. Composition of preservative, adjuvant or stabilizer, and proportions used stated in such a manner that the concentration can be calculated; stage and method of addition. C. Method and degree of concentration. D. If product is standardized to give con centration of antigen, show procedures and calculations. E. 1. Assembly of units to make a serial (il lustrate by example). 2. Volume of average serial. 3. Volume of maximum serial. 4. Any other pertinent information. F. Volume of fill for each size vial. Type of vial if unusual. G. Method and technique of filling and sealing of final containers. H. Desiccation, including moisture control. Give maximum percent moisture. I. Amount of antigenic material per dose or doses in final container. V. Testing. Indicate the stages in the prepa ration of the biological product at which the samples are collected. Refer to all applicable Standard Requirements. Outline all addi tional tests in detail and state the minimum requirement for each satisfactory test. A. Purity. B. Safety. C. Potency. D. Moisture, if desiccated. E. Any other tests. VI. Post preparatory steps. A. Form and size of final containers in which the product is to be distributed. B. Collection, storage, and submission of representative samples. Indicate at which steps in the production these samples are taken. C. Expiration date based on the earliest date of harvest and the date of the last satis factory potency test. If applicable, give the date of lyophilization. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00832 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 823 Animal and Plant Health Inspection Service, USDA § 114.9 D. Use, dosage, and route of administration for each animal species for which the bio logical product is recommended. (e) Outlines of Production for aller genic extracts shall be written accord ing to the following: OUTLINE GUIDE FOR ALLERGENIC EXTRACTS License No. Name of Product Date I. Composition of the product. A. Source and type of raw material. B. Weight/volume concentration. II. Preparation of the product. A. Describe fully and show each step of preparation to the completion of the finished product in true containers. In describing the prepara tion of the product, emphasize the following: 1. Method of extraction. 2. Composition of preservative, adjuvant or stabilizer, and proportions used; stage and method of addition. 3. Method and degree of concentration. 4. Standardization of the product. 5. (a) Assembly of units to make a serial. (b) Volume of average serial. (c) Maximum serial. 6. Volume of fill for each size vial. 7. Method and technique of filling and seal ing of final containers. 8. Amount material per dose or doses in final container. III. Testing. Indicate the stages in the prep aration of the biological product at which the samples are collected. Refer to all appli cable Standard Requirements. Outline all ad ditional tests in detail and state the min imum requirement for each satisfactory test. A. Purity. B. Safety. C. Potency. D. Any other tests. E. Include any additional pertinent infor mation. IV. Post preparatory steps. A. Form and size of final containers in which the product is to be distributed. B. Collection, storage, and submission of representative samples. Indicate at which steps in the production these samples are taken. C. Expiration date based on the earliest date of harvest and the date of the last satis factory potency test. D. Use, dosage, and route of administration for each animal species for which the bio logical product is recommended. (f) Outlines of Production for diag nostic test kits based on antigen-anti body reactions, and other diagnostics whose production methods are ame nable to description as described herein shall be written according to the fol lowing requirements: OUTLINE GUIDE FOR DIAGNOSTIC TEST KITS License No. Name of product Date Introduction Provide a brief description of the kit as fol lows: 1. Principle of the test (ELISA, latex ag glutination, etc.). 2. Antigen or antibody detection test. 3. Sample(s) used for testing (serum, whole blood, tears, etc.). 4. List reagents, references, and equipment included. 5. Identify materials obtained under split manufacturing agreements. 6. General description of test interpreta tions and their limitations, including fol lowup tests. I. Antibody Components A. Production of polyclonal antibody com ponents. 1. If purchased, list suppliers, criteria for acceptability, and describe all tests per formed after receipt to determine that speci fications have been met. 2. If produced in-house, describe the spe cies, age, weight, conditions, and general health of all animals used in antiserum pro duction. a. Preinjection considerations: Describe the examination, preparation, care, quarantine procedures, and treatments administered before immunization(s). De scribe all tests used to determine suitability for use. Describe the preparation of any standard negative serum(s) collected prior to immunization. b. Immunization of animals. i. Describe the character and dose of the antigen; if adjuvant is used provide details on its preparation. If commercial product is used include its true name as shown on the label, the manufacturer, serial number, and expiration date. ii. Describe the method and schedule for immunizations. iii. Describe the method for harvesting and evaluating the immunization product, in cluding tests for acceptability. iv. Provide number and intervals between harvests, volume obtained, and any other pertinent information. B. Production of Monoclonal Antibody Components. 1. Hybridoma components: a. If hybridoma components are purchased, list suppliers and criteria for acceptability; if tests are performed after receipt, describe fully. b. If hybridomas are prepared inhouse, identify the antigen(s) used, describe the im munization scheme, and the species of ani mal used. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00833 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 824 9 CFR Ch. I (1–1–18 Edition) § 114.9 c. Identify the tissue of origin, and the pro cedures for harvesting, isolating, and identi fying the immune cells. d. Describe the source, identity, and the product secreted (light or heavy chain) by the parent Myeloma Cell Line. e. Summarize cloning and recloning proce dures, including clone characterization and propagation, if appropriate. f. If appropriate, describe procedures for establishing and maintaining seed lots. g. Describe any other pertinent tests or procedures performed on the hybridoma cell line. 2. Antibody production: a. Describe the production method. If pro duced in cell culture, animal serum additives must conform to 9 CFR 113.53. If produced in animals, describe fully including husbandry practices and passage procedures. b. Provide the criteria for acceptable monoclonal antibody, including tests for pu rity. c. Describe all tests or other methods used to ensure uniformity between production lots of monoclonal antibody. Include all re action conditions, equipment used, and reac tivity of the component. d. Describe all characterization procedures and include the expected reactivity of all ref erence monoclonal antibodies. II. Antigen Preparation A. Identify the microorganism(s) or anti gen being used. If previously approved Mas ter Seed virus, bacteria, or antigen derived therefrom is used, provide pertinent infor mation on the testing performed, and details of dates of United States Department of Ag riculture confirmatory tests and approval, as appropriate. B. Describe all propagation steps, includ ing identification of cell cultures, media in gredients, cell culture conditions, and har vest methods. For antigen produced in eggs, give the egg source, age, and route of inocu lation. If cell lines are being used, give dates of testing and approval as specified in 9 CFR 113.52. C. Describe procedures used for extracting and characterizing the antigen. D. Describe the method used to standardize the antigen. E. If the antigen is purchased, identify the supplier and describe the criteria for accept able material, including all tests performed by the producer and/or the recipient to deter mine acceptability. III. Preparation of Standard Reagents A. Describe the positive and negative con trols included in the kit. If purchased, list suppliers and criteria for acceptance. B. Describe the preparation and standard ization of the conjugate(s). If purchased, list suppliers and criteria for acceptance. C. Describe the preparation and standard ization of the substrate(s). If purchased, list suppliers and criteria for acceptance. D. Identify buffers, diluents, and other re agents included in the kit. The preparation of these components may be described in this section or in filed Special Outlines. IV. Preparation of the Product Fully describe methods used to standardize antigens, reference standards, positive con trol serum, negative control serum, and standard reagents from production/purchase to completion of finished product in final containers, including the following: 1. Composition and quantity of preserva tive in each. 2. Method of filling, plating, or attaching the antigen or antibody component to a solid phase. 3. Minimum and maximum acceptable fill volumes for each final container of reagent included in the kit. 4. The disposition of unsatisfactory mate rial. V. Testing Refer to all applicable standard require ments. A. Purity. Describe all tests of the kit for purity or specify the exemption as provided in 9 CFR 113.4. B. Safety. In vitro products are exempt from safety tests. C. Potency. Provide details of tests used to determine the relative reactivity of the kit including minimum requirements for a satisfactory test. Reference standards and control serum used for this purpose should be identified by unique codes or lot numbers. VI. Postpreparatory Steps A. Describe the form and size of final con tainers of each reagent/component included in the kit. B. Describe the collection, storage, and submission of representative samples. Refer to 9 CFR 113.3(b)(7). C. Specify the expiration date. Refer to 9 CFR 114.13. D. Provide details of recommendations for use, including all limitations, qualifications, and interpretation of results. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00834 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 825 Animal and Plant Health Inspection Service, USDA § 114.13 E. Submit confidentiality statement iden tifying specific parts of the outline con taining information, the release of which would cause harm to the submitter. (Approved by the Office of Management and Budget under control number 0579–0013) [39 FR 16869, May 10, 1974, as amended at 48 FR 57473, Dec. 30, 1983; 56 FR 20124, May 2, 1991; 56 FR 66784, Dec. 26, 1991; 75 FR 20773, Apr. 21, 2010] § 114.10 Antibiotics as preservatives. Antibiotics are authorized for use as preservatives for biological products if used within the limitations as to kinds and amounts prescribed in this section. (a) When an antibiotic or combina tion of antibiotics, with or without a fungistat is to be used in the prepara tion of a biological product, the kind(s) and amount(s) of each shall be speci fied in the outline for such product in such a way that the concentration in the final product may be calculated. Except as may be approved by the Ad ministrator, only those individual anti biotics or combinations of antibiotics listed in paragraphs (b) and (c) of this section shall be used. (b) Permitted individual antibiotics: (1) The antibiotic level of a specified individual antibiotic in one ml. of a bi ological product, when prepared as rec ommended for use, shall not exceed the amounts listed in this paragraph: Pro vided, That in the case a desiccated bio logical product is to be used with an in definite quantity of water or other menstruum, the determination shall be based on 30 ml. per 1,000 dose vial or equivalent. (2) Except as prescribed in paragraph (c) of this section, only one antibiotic shall be used as a preservative in a bio logical product. The kind and max imum amount per ml. of such anti biotic shall be restricted to: Amphotericin B ................................................. 2.5 mcg. Nystatin (Mycostatin) ........................................ 30.0 units Tetracyclines .................................................... 30.0 mcg. Penicillin ........................................................... 30.0 units Streptomycin .................................................... 30.0 mcg. Polymyxin B ..................................................... 30.0 mcg. Neomycin ......................................................... 30.0 mcg. Gentamicin ....................................................... 30.0 mcg. (c) Permitted combinations: (1) Penicillin and streptomycin. (2) Either amphotericin B or nys tatin, but not both, may be used with one of the other antibiotics listed in paragraph (b) of this section, or with a combination of penicillin and strepto mycin, or with a combination of poly myxin B and neomycin. (3) The maximum amount of each an tibiotic in a combination shall be the amount prescribed for such antibiotic in paragraph (b) of this section. (d) Antibiotics used in virus seed stock purification are not restricted as to kind or amounts provided carryover into the final product is controlled and specified in outlines of production. [39 FR 16869, May 10, 1974, as amended at 56 FR 66784, Dec. 26, 1991] § 114.11 Storage and handling. Biological products at licensed estab lishments must be protected at all times against improper storage and handling. Completed product must be kept under refrigeration at 35 to 46 °F (2 to 8 °C), unless the inherent nature of the product makes storage at dif ferent temperatures advisable, in which case, the proper storage tem perature must be specified in the filed Outline of Production. All biological products to be shipped or delivered must be securely packed. [81 FR 59436, Aug. 30, 2016] § 114.12 Expiration date required. Each serial or subserial of biological product prepared in a licensed estab lishment shall be given an expiration date determined in accordance with the requirements provided in § 114.13 or § 114.14. A licensed biological product shall be considered worthless under the Virus-Serum-Toxin Act subsequent to the expiration date appearing on the label. [41 FR 44687, Oct. 12, 1976] § 114.13 Expiration date determina tion. Unless otherwise provided for in a Standard Requirement of filed Outline of Production, the expiration date for each product shall be computed from the date of the initiation of the po tency test. Prior to licensure, stability of each fraction shall be determined by methods acceptable to Animal and Plant Health Inspection Service. Expi ration dates based on this stability data shall be confirmed as follows: VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00835 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 826 9 CFR Ch. I (1–1–18 Edition) § 114.14 (a) Products consisting of viable orga nisms. Each serial shall be tested for potency at release and at the approxi mate expiration date until a statis tically valid stability record has been established. (b) Nonviable biological products. Each serial presented in support of licensure shall be tested for potency at release and at or after the dating requested. (c) Subsequent changes in the dating period for a product may be granted, based on statistically valid data sub mitted to support a revision of the Out line of Production. [50 FR 24903, June 14, 1985, as amended at 56 FR 66784, Dec. 26, 1991] § 114.14 Extension of expiration date for a serial or subserial. (a) Unless otherwise provided for in a filed Outline of Production for the product, the expiration date shall not be extended: (1) If all fractions of the product are not evaluated for potency by tests des ignated in the filed Outline of Produc tion for such product in accordance with § 113.4(b) of this subchapter. (2) For any serial or portion of any serial which has left licensed premises: Provided, That product which has been shipped from one licensed premises to another licensed premises shall be ex empt from this requirement. (3) For a serial or portion of a serial if the expiration date has been ex tended previously, unless otherwise au thorized in accordance with § 114.1. (b) An extension of the expiration date may be granted by Animal and Plant Health Inspection Service if a re quest from the licensee is substan tiated by valid test data which dem onstrate the potency of the product meets or exceeds the requirements for release. The new expiration date shall be calculated from the date the latest satisfactory potency test was initiated. The extension of the expiration date shall not exceed the maximum dating allowed in the filed Outline of Produc tion. (1) Serials are approved for redating under the condition that Animal and Plant Health Inspection Service may require the firm to retest the redated serial for potency during the extended dating period and if found unsatisfac tory require it be removed from the market by the licensee. (2) [Reserved] [50 FR 24903, June 14, 1985, as amended at 56 FR 66784, Dec. 26, 1991] § 114.15 Disposal of unsatisfactory products and byproducts. All biological products found to be unsatisfactory for marketing, all bio logical products which have become worthless subsequent to the expiration date, all refuse, other materials deemed unsatisfactory for production purposes, all carcasses (part or whole) of production or test animals, and any undesirable byproducts of manufacture shall be disposed of as may be required by the Administrator. [41 FR 44687, Oct. 12, 1976, as amended at 56 FR 66784, Dec. 26, 1991] § 114.16 Producing subsidiaries. A serial or subserial of a biological product may be produced jointly by a licensee and one or more subsidiaries, or by two or more subsidiaries. The exact amount of each serial or sub serial credited to each participating producer shall be determined at the time of labeling and packaging and shall be noted in the records for such serial or subserial. [40 FR 46093, Oct. 6, 1975] § 114.17 Rebottling of biological prod ucts. The Administrator may authorize the rebottling of a completed product in liquid form subject to the conditions prescribed in this section. (a) All or part of a serial which has not left the licensed establishment may be aseptically returned to the mixing tank, thoroughly mixed, and re bottled in new final containers. (b) The rebottled product shall be adequately identified by serial number or subserial number, as the case may be. (c) Required purity tests for final container samples of the product shall be conducted on new samples selected from the rebottled product (serial or subserials). Rebottled product found to be unsatisfactory by such tests shall not be released. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00836 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 827 Animal and Plant Health Inspection Service, USDA § 115.2 (d) New test samples from each serial or subserial and copies of test reports of all tests conducted on the rebottled product shall be submitted to Animal and Plant Health Inspection Service. (e) The licensee shall not release the rebottled product unless notified by Animal and Plant Health Inspection Service that such product is eligible for release. Production records shall show the results of all tests conducted and shall accurately reflect the actions taken. [39 FR 16869, May 10, 1974, as amended at 56 FR 66784, Dec. 26, 1991] § 114.18 Reprocessing of biological products. The Administrator may authorize a licensee to reprocess a serial of com pleted product subject to the condi tions prescribed in this section. (a) Reprocessing shall not include any method or procedure which would be deleterious to the product. (b) All appropriate tests for purity, safety, potency, and efficacy for the product shall be conducted on the re processed product. A serial found un satisfactory by a required test shall not be released. (c) The reprocessed serial shall be identified by a new serial number and the records for the serial shall accu rately reflect the action taken. (d) Test samples of the reprocessed serial and test reports for all tests con ducted shall be submitted to Animal and Plant Health Inspection Service. The licensee shall not release the serial until notified by Animal and Plant Health Inspection Service that the se rial is eligible for release. [50 FR 24904, June 15, 1985, as amended at 56 FR 66784, Dec. 26, 1991] PART 115—INSPECTIONS Sec. 115.1 Inspections of establishments. 115.2 Inspections of biological products. AUTHORITY: 21 U.S.C. 151–159; 7 CFR 2.22, 2.80, and 371.4. § 115.1 Inspections of establishments. (a) Any inspector shall be permitted to enter any establishment where any biological product is prepared, at any hour during the day or night, and shall be permitted to inspect, without pre vious notification, the entire premises of the establishment, including all buildings, compartments, and other places, all biological products, and or ganisms and vectors in the establish ment, and all materials and equipment, such as chemicals, instruments, appa ratus, and the like, and the methods used in the manufacture of, and all records maintained relative to, biologi cal products produced at such estab lishment. (b) Each inspector will have in his or her possession a numbered USDA badge or identification card. Either shall be sufficient identification to entitle him/ her to admittance at all regular en trances and to all parts of such estab lishment and premises and to any place at any time for the purpose of making an inspection pursuant to paragraph (a) of this section. [52 FR 30134, Aug. 13, 1987] § 115.2 Inspections of biological prod ucts. (a) Any biological product, the con tainer of which bears a United States veterinary license number or a United States veterinary permit number or other mark required by these regula tions, may be inspected at any time or place. If, as a result of such inspection, it appears that any such product is worthless, contaminated, dangerous, or harmful, the Secretary shall give no tice to stop distribution and sale to the manufacturer (licensee) or importer (permittee) and may proceed against such product pursuant to the provi sions of part 118 of this subchapter. (b) When notified to stop distribution and sale of a serial or subserial of a veterinary biological product by the Secretary, veterinary biologics licens ees or permittees shall: (1) Stop the preparation, distribu tion, sale, barter, exchange, shipment, or importation of the affected serial(s) or subserial(s) of any such veterinary biological product pending further in structions from APHIS. (2) Immediately, but no later than 2 days, send stop distribution and sale notifications to any jobbers, whole salers, dealers, foreign consignees, or other persons known to have any such VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00837 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 828 9 CFR Ch. I (1–1–18 Edition) Pt. 116 veterinary biological product in their possession, which instruct them to stop the preparation, distribution, sale, bar ter, exchange, shipment, or importa tion of any such veterinary biological product. All notifications shall be doc umented in writing by the licensee or permittee. (3) Account for the remaining quan tity of each serial(s) or subserial(s) of any such veterinary biological product at each location in the distribution channel known to the manufacturer (li censee) or importer (permittee). (4) When required by the Adminis trator, submit complete and accurate reports of all notifications concerning stop distribution and sale actions to the Animal and Plant Health Inspec tion Service pursuant to § 116.5 of this subchapter. (c) Unless and until the Secretary shall otherwise direct, no persons so notified shall thereafter sell, barter, or exchange any such product in any place under the jurisdiction of the United States or ship or deliver for shipment any such product in or from any State, Territory, or the District of Columbia. However, failure to receive such notice shall not excuse any person from compliance with the Virus Serum-Toxin Act. (Approved by the Office of Management and Budget under control number 0579–0318) [72 FR 17798, Apr. 10, 2007] PART 116—RECORDS AND REPORTS Sec. 116.1 Applicability and general consider ations. 116.2 Inventory and disposition records. 116.3 Label records. 116.4 Sterilization and pasteurization records. 116.5 Reports. 116.6 Animal records. 116.7 Test records. 116.8 Completion and retention of records. AUTHORITY: 21 U.S.C. 151–159; 7 CFR 2.22, 2.80, and 371.4. § 116.1 Applicability and general con siderations. (a) Each licensee, permittee, and for eign manufacturer of biological prod ucts imported into the United States shall maintain, at the licensed or for eign establishment in which the prod ucts are prepared, detailed records of information necessary to give a com plete accounting of all the activities within each establishment. Such records shall include, but shall not be limited to, the items enumerated in this part. (1) Records shall be made concur rently with the performance of succes sive steps in the development and prep aration of biological products, includ ing new products under development. Such records shall include the date and where critical, the time that each es sential step was taken, the identity and quantity of ingredients added or removed at each step, and any gain or loss of product from the beginning to the end of product preparation. (2) Records shall be legible and indel ible; shall be as detailed as necessary for a clear understanding of each step by one experienced in the preparation of biological products; and shall be verified by initials or signature of the person immediately responsible for the action taken. (3) Records (other than disposition records) required by this part shall be completed by the licensee or the for eign manufacturer, as the case may be, before any portion of a serial of any product shall be marketed in the United States or exported. (b) In the case of imported products, each permittee shall maintain at the permittee’s place of business detailed and accurate records that are relevant to each imported product and that in clude, but are not limited to, importa tion documents, sampling records, test summaries, shipping records, and in ventory and disposition records as re quired in § 116.2. (c) When authorized by the Adminis trator, the licensee, permittee, or for eign manufacturer may maintain and retain records required under this part at an alternative location. Such au thorization shall be confirmed by the filing of an addendum to the plot plan legend. The addendum shall list the lo cation of the records and the condition of their storage and shall permit the VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00838 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 829 Animal and Plant Health Inspection Service, USDA § 116.6 inspection of the records by APHIS in spectors, or foreign inspectors acting on behalf of APHIS. (Approved by the Office of Management and Budget under control number 0579–0013) [39 FR 16872, May 10, 1974, as amended at 48 FR 57473, Dec. 30, 1983; 61 FR 52874, Oct. 9, 1996; 66 FR 21064, Apr. 27, 2001] § 116.2 Inventory and disposition records. (a) Records shall show the quantity and location of each biological product being prepared, in storage, and in dis tribution channels. (b) Detailed disposition records, in a form satisfactory to the Adminis trator, shall be maintained by each li censee, each distributor, and each per mittee showing the sale, shipment, or other disposition made of the biologi cal products handled by such person. (Approved by the Office of Management and Budget under control number 0579–0013) [39 FR 16872, May 10, 1974, as amended at 48 FR 57473, Dec. 30, 1983; 56 FR 66784, Dec. 26, 1991; 61 FR 52874, Oct. 9, 1996; 66 FR 21064, Apr. 27, 2001] § 116.3 Label records. (a) Each licensee and permittee shall maintain a list of all approved labels currently being used. Each label shall be identified as to: (1) Name and product code number as it appears on the product license or permit for the product; (2) Where applicable, the size of the package (doses, ml, cc, or units) on which the label shall be used; (3) Label number and date assigned; and (4) Name of licensee or subsidiary ap pearing on the label as the producer. (b) All labels printed shall be ac counted for and an inventory main tained. Records shall include the disposition of such labels including those not used in labeling a product. (Approved by the Office of Management and Budget under control number 0579–0013) [39 FR 16872, May 10, 1974, as amended at 48 FR 57473, Dec. 30, 1983; 61 FR 52874, Oct. 9, 1996; 66 FR 21064, Apr. 27, 2001] § 116.4 Sterilization and pasteurization records. Records shall be made by means of automatic recording devices or an equivalent accurate and reliable sys tem. Such records shall be identified with the ingredients, equipment, or bi ological product subjected to steriliza tion or pasteurization. (Approved by the Office of Management and Budget under control number 0579–0013) [39 FR 16872, May 10, 1974, as amended at 48 FR 57473, Dec. 30, 1983; 61 FR 52874, Oct. 9, 1996; 66 FR 21064, Apr. 27, 2001] § 116.5 Reports. (a) When required by the Adminis trator, reports containing accurate and complete information concerning bio logical products, including but not lim ited to, product development and prep aration, and market suspensions and recalls, shall be prepared and sub mitted to the Animal and Plant Health Inspection Service by the licensee, per mittee, or foreign manufacturer (whose products are being imported or offered for importation). Unless otherwise au thorized by the Administrator, records necessary to make such reports shall be maintained in each establishment. (b) If, at any time, there are indica tions that raise questions regarding the purity, safety, potency, or efficacy of a product, or if it appears that there may be a problem regarding the prepa ration, testing, or distribution of a product, the licensee, permittee, or for eign manufacturer must immediately notify the Animal and Plant Health In spection Service concerning the cir cumstances and the action taken, if any. Notification may be made by mail to Director, Center for Veterinary Bio logics, Inspection and Compliance, 1920 Dayton Avenue, P.O. Box 844, Ames, IA 50010; by electronic mail to (cvb@aphis.usda.gov); by fax to (515) 337 6120; or by telephone to (515) 337-6100. (Approved by the Office of Management and Budget under control number 0579–0013) [61 FR 52874, Oct. 9, 1996, as amended at 64 FR 43045, Aug. 9, 1999; 75 FR 20773, Apr. 21, 2010] § 116.6 Animal records. Complete records shall be kept for all animals at a licensed establishment. Results of tests performed, antigens or VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00839 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 830 9 CFR Ch. I (1–1–18 Edition) § 116.7 treatment administered, maintenance and production records, disposition records, necropsy records, if any, and all other pertinent records shall be in cluded. (Approved by the Office of Management and Budget under control number 0579–0013) [39 FR 16872, May 10, 1974, as amended at 48 FR 57473, Dec. 30, 1983; 61 FR 52874, Oct. 9, 1996; 66 FR 21064, Apr. 27, 2001] § 116.7 Test records. Detailed records of all tests con ducted on each serial and each sub serial shall be maintained by the li censee. Summaries of such tests shall be prepared from such records and sub mitted to the Animal and Plant Health Inspection Service using APHIS Form 2008 or an acceptable equivalent form prior to release of the serial or sub serial. Blank forms for such summaries shall be available from Animal and Plant Health Inspection Service upon request. (Approved by the Office of Management and Budget under control number 0579–0013) [39 FR 16872, May 10, 1974, as amended at 48 FR 57473, Dec. 30, 1983; 56 FR 66784, Dec. 26, 1991; 61 FR 52874, Oct. 9, 1996] § 116.8 Completion and retention of records. All records (other than disposition records) required by this part shall be completed by the licensee, permittee, or foreign manufacturer before any portion of a serial of any product may be marketed in the United States or exported. All records shall be retained at the licensed or foreign establish ment or permittee’s place of business for a period of two years after the expi ration date of a product, or for such longer period as may be required by the Administrator. (Approved by the Office of Management and Budget under control number 0579–0013) [61 FR 52874, Oct. 9, 1996] PART 117—ANIMALS AT LICENSED ESTABLISHMENTS Sec. 117.1 Applicability. 117.2 Animal facilities. 117.3 Admittance of animals. 117.4 Test animals. 117.5 Segregation of animals. 117.6 Removal of animals. AUTHORITY: 21 U.S.C. 151–159; 7 CFR 2.22, 2.80, and 371.4. SOURCE: 38 FR 15499, June 13, 1973, unless otherwise noted. § 117.1 Applicability. (a) All animals used in licensed es tablishments in the preparation or testing of biological products shall meet the regulations in this subchapter and special requirements as may be prescribed by the Administrator to pre vent the preparation, sale, and dis tribution of worthless, contaminated, dangerous, or harmful biological prod ucts. (b) Unless otherwise authorized or di rected by the Administrator, animals used in the preparation or testing of bi ological products shall be admitted to and maintained at the licensed estab lishment and ultimately disposed of in accordance with the regulations in this part, and with the Act of August 24, 1966 (Pub. L. 89–544) as amended by the Animal Welfare Act of 1970 (Pub. L. 91– 579) and the regulations in parts 1, 2, and 3 of this chapter. Personnel who supervise the care and welfare of such animals shall be qualified by edu cation, training, and experience to carry out the regulations in this part. [38 FR 15499, June 13, 1973, as amended at 56 FR 66784, Dec. 26, 1991] § 117.2 Animal facilities. Animal facilities shall comply with the requirements provided in part 108 of this chapter. § 117.3 Admittance of animals. (a) No animal which shows clinical signs or other evidence of disease shall be admitted to the premises of licensed establishments, except as provided in paragraphs (d) and (e) of this section. The health status of all animals offered for admission shall be determined by or under the direction of a veterinarian prior to admission. If the determina tion cannot be made prior to admis sion, the animals shall be kept sepa rate from animals already on the prem ises and in a quarantine area to be pro vided by the licensee for this purpose VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00840 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 831 Animal and Plant Health Inspection Service, USDA § 117.6 until the animal’s health status is de termined. (b) If special test requirements for admittance of the animals are specified in the Outline of Production for the product to be produced, the animals shall remain in the quarantine area until such tests have been performed and the results obtained. Animals which do not meet the requirements shall not be admitted to the production area or allowed to contact production animals. (c) All animals admitted to the prem ises of a licensed establishment shall be permanently identified either col lectively or individually by the li censee with tags, marks, or other means acceptable to the Adminis trator. (d) When an animal which has a dis ease is to be used to prepare a biologi cal product for control of such disease, the animal shall be admitted directly to the processing facilities in which the product is to be prepared but shall not be permitted contact with other ani mals on the premises. (e) The Administrator may authorize the maintenance of diagnostic facili ties at the licensed establishment: Pro vided, That safeguards proposed by the licensee are adequate to prevent dis eased or dead animals brought into such facilities from being a threat to biological products prepared in such es tablishment or to other animals on the premises used in the preparation of bio logical products. [38 FR 15499, June 13, 1973, as amended at 56 FR 66784, Dec. 26, 1991] § 117.4 Test animals. (a) All test animals shall be exam ined for clinical signs of illness, injury, or abnormal behavior prior to the start of a test and throughout the observa tion period specified in the test pro tocol. (b) All animals used for test purposes shall be identified either collectively or individually in a manner conducive to an accurate interpretation of the re sults of the test. (c) No test animals shall be given a biological product during the precondi tioning period which would affect its eligibility according to the test re quirements. No treatment, with a bio logical product or otherwise, shall be administered to a test animal during a test period which could interfere with a true evaluation of the biological prod uct being tested. (d) During the course of a test, ani mals that are injured or show clinical signs of illness or unfavorable reac tions that are not due to the test may be removed from the test and treated or humanely destroyed. If sufficient animals do not remain for the test to be evaluated, the test shall be declared inconclusive and may be repeated. (e) Test animals that show clinical signs of illness that are due to the test may be treated or humanely destroyed if the illness has progressed to a point (defined in the filed Outline of Produc tion) when death is certain to occur without therapeutic intervention. When interpreting the results of the test, the animals that were treated or humanely destroyed because of illness due to the test and the animals that have died from illness due to the test prior to being humanely destroyed shall be combined into a common sta tistic of mortality due to the test. [38 FR 15499, June 13, 1973, as amended at 60 FR 43356, Aug. 21, 1995] § 117.5 Segregation of animals. Animals which have been infected with or exposed to a dangerous, infec tious, contagious, or communicable disease shall be kept effectively seg regated at a licensed establishment until such time as they are humanely destroyed or successfully treated and removed as healthy animals. § 117.6 Removal of animals. Production animals or ex-test ani mals which are no longer useful at the licensed establishment may be re moved from the premises of the li censed establishment; provided, such removal is accomplished in a manner as shall preclude the dissemination of disease and in accordance with the fol lowing conditions: (a) Meat-producing animals which re ceived a biological product containing inactivated microorganisms and adju vants within 21 days shall not be re moved; or VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00841 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 832 9 CFR Ch. I (1–1–18 Edition) Pt. 118 (b) Animals which received virulent microorganisms within 30 days shall not be removed; or (c) Only animals that are in a healthy condition as determined by a veterinarian shall be removed, except as provided in paragraph (d) of this sec tion. (d) Other animals that are injured or otherwise unhealthy, except when af fected with a communicable disease, may be removed for immediate slaugh ter to an abattoir operated in accord ance with the Federal Meat Inspection Act of March 4, 1907, 34 Stat. 1260, as amended by the Wholesome Meat Act of 1967, 81 Stat. 585 (21 U.S.C. sec. 601 et seq.): Provided, That such animals shall be properly marked for identification and the inspector in charge of slaugh ter operations is given due notice in advance. (e) All animals on the premises shall be disposed of in accordance with the provisions of the regulations in this part and where specific provision is not made therefor shall be disposed of as required by the Administrator. [38 FR 15499, June 13, 1973, as amended at 56 FR 66784, Dec. 26, 1991] PART 118—DETENTION; SEIZURE AND CONDEMNATION Sec. 118.1 Administrative detention. 118.2 Method of detention; Notifications. 118.3 Movement of detained biological prod ucts; Termination of detention. 118.4 Seizure and condemnation. AUTHORITY: 21 U.S.C. 151–159; 7 CFR 2.22, 2.80, and 371.4. SOURCE: 52 FR 30135, Aug. 13, 1987, unless otherwise noted. § 118.1 Administrative detention. Whenever any biological product which is prepared, sold, bartered, ex changed, or shipped in violation of the Act or regulations is found by any au thorized representative of the Adminis trator upon any premises, it may be de tained by such representative for a pe riod not to exceed 20 days, pending ac tion under § 118.4, and shall not be moved by any person from the place at which it is located when so detained, until released by such representative. [52 FR 30135, Aug. 13, 1987, as amended at 56 FR 66784, Dec. 26, 1991] § 118.2 Method of detention; Notifica tions. An authorized representative of the Administrator shall detain any biologi cal product subject to detention under this part by: (a) Giving oral notification to the owner of the biological product if such owner can be ascertained, and, if not, to the agent representing the owner or to the immediate custodian of the bio logical product; and (b) Promptly furnishing the person so notified with a preliminary notice of detention which shall include identity and quantity of the product detained, the location where detained, the reason for the detention, and the name of the authorized representative of the Ad ministrator. (c) Within 48 hours after the deten tion of any biological product, an au thorized representative of the Adminis trator shall, if the detention is to con tinue, give written notification to the owner of the biological product de tained by furnishing a written state ment which shall include the identity and quantity of the product detained, the location where detained, specific description of the alleged noncompli ance including reference to the provi sions in the Act or the regulations which have resulted in the detention, and the identity of the authorized rep resentative of the Administrator; or, if such owner cannot be ascertained and notified within such period of time, furnish such notice to the agent rep resenting such owner, or the carrier or other person having custody of the bio logical product detained. The notifica tion, with a copy of the preliminary notice of detention shall be served by either delivering the notification to the owner or to the agent or to such other person, or by certifying and mail ing the notification, addressed to such owner, agent, or other person, at the last known residence or principal office or place of business. [52 FR 30135, Aug. 13, 1987, as amended at 56 FR 66784, Dec. 26, 1991] VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00842 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 833 Animal and Plant Health Inspection Service, USDA Pt. 121 § 118.3 Movement of detained biologi cal products; Termination of deten tion. Except as provided in paragraphs (a) and (b) of this section, no biological product detained in accordance with the provisions in this part shall be moved by any person from the place at which such product is located when it is detained. (a) A detained biological product may be moved from the place at which it is located when so detained for the purpose of providing proper storage conditions if such movement has been approved by an authorized representa tive of the Administrator; Provided, That, the biological product so moved shall be detained by an authorized rep resentative of the Administrator after such movement. (b) A detained biological product may be moved from the place at which it is detained on written notification by an authorized representative of the Administrator that the detention is terminated; Provided, That, the condi tions under which the detained biologi cal product may be moved will be spec ified in the written notification of the termination. The notification of termi nation shall be served by either person ally delivering the notification, or by certifying and mailing the notification addressed to such person at the last known residence or principal office or place of business of the owner, agent, or other person having custody of the biological product. [52 FR 30135, Aug. 13, 1987, as amended at 56 FR 66784, Dec. 26, 1991] § 118.4 Seizure and condemnation. Any biological product which is pre pared, sold, bartered, exchanged, or shipped in violation of the Act or regu lations shall be liable to be proceeded against and seized and condemned, at any time, on a libel of information in any United States district court or other proper court within the jurisdic tion of which the product is found. If the product is condemned, it shall, after entry of the decree, be disposed of by destruction or sale as the court may direct, and the proceeds, if sold, less the court costs and fees, and storage and other proper expenses, shall be paid into the Treasury of the United States, but the product shall not be sold contrary to the provisions of the Act or the laws of the jurisdiction in which it is sold; Provided, That, upon the execution and delivery of a good and sufficient bond conditioned that the product shall not be sold or other wise disposed of contrary to the provi sions of the Act or the laws or jurisdic tion in which disposal is made, the court may direct that such product be delivered to the owner thereof subject to such supervision by authorized rep resentatives of the Administrator as is necessary to ensure compliance with the applicable laws. When a decree of condemnation is entered against the product and it is released under bond, or destroyed, court costs and fees, and storage and other proper expenses shall be awarded against the person, if any, intervening as claimant of the product. The proceedings in such libel cases shall conform, as nearly as may be practicable, to the proceedings in ad miralty, except that either party may demand trial by jury of any issue of fact joined in any case, and all such proceedings shall be at the suit of and in the name of the United States. [52 FR 30135, Aug. 13, 1987, as amended at 56 FR 66784, Dec. 26, 1991] PART 121—POSSESSION, USE, AND TRANSFER OF SELECT AGENTS AND TOXINS Sec. 121.1 Definitions. 121.2 Purpose and scope. 121.3 VS select agents and toxins. 121.4 Overlap select agents and toxins. 121.5 Exemptions for VS select agents and toxins. 121.6 Exemptions for overlap select agents and toxins. 121.7 Registration and related security risk assessments. 121.8 Denial, revocation, or suspension of registration. 121.9 Responsible official. 121.10 Restricting access to select agents and toxins; security risk assessments. 121.11 Security. 121.12 Biosafety. 121.13 Restricted experiments. 121.14 Incident response. 121.15 Training. 121.16 Transfers. 121.17 Records. 121.18 Inspections. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00843 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 834 9 CFR Ch. I (1–1–18 Edition) § 121.1 121.19 Notification of theft, loss, or release. 121.20 Administrative review. AUTHORITY: 7 U.S.C. 8401; 7 CFR 2.22, 2.80, and 371.4. SOURCE: 70 FR 13284, Mar. 18, 2005, unless otherwise noted. § 121.1 Definitions. Administrator. The Administrator, Animal and Plant Health Inspection Service, or any person authorized to act for the Administrator. Animal and Plant Health Inspection Service (APHIS). The Animal and Plant Health Inspection Service of the U.S. Department of Agriculture. Attorney General. The Attorney Gen eral of the United States or any person authorized to act for the Attorney Gen eral. Biological agent. Any microorganism (including, but not limited to, bacteria, viruses, fungi, or protozoa), or infec tious substance, or any naturally oc curring, bioengineered, or synthesized component of any such microorganism or infectious substance, capable of causing: (1) Death, disease, or other biological malfunction in a human, an animal, a plant, or another living organism; (2) Deterioration of food, water, equipment, supplies, or material of any kind; or (3) Deleterious alteration of the envi ronment. Centers for Disease Control and Preven tion (CDC). The Centers for Disease Control and Prevention of the U.S. De partment of Health and Human Serv ices. Diagnosis. The analysis of specimens for the purpose of identifying or con firming the presence or characteristics of a select agent or toxin, provided that such analysis is directly related to protecting the public health or safety, animal health or animal products, or plant health or plant products. Entity. Any government agency (Fed eral, State, or local), academic institu tion, corporation, company, partner ship, society, association, firm, sole proprietorship, or other legal entity. HHS Secretary. The Secretary of the Department of Health and Human Services or his or her designee, unless otherwise specified. HHS select agent and/or toxin. A bio logical agent or toxin listed in 42 CFR 73.3. Import. To move into, or the act of movement into, the territorial limits of the United States. Information security. Protecting infor mation and information systems from unauthorized access, use, disclosure, disruption, modification, or destruc tion in order to provide: (1) Integrity, which means guarding against improper information modi fication or destruction, and includes ensuring information authenticity; (2) Confidentiality, which means pre serving authorized restrictions on ac cess and disclosure, including means for protecting personal privacy and proprietary information; and (3) Availability, which means ensuring timely and reliable access to and use of information. Interstate. From one State into or through any other State, or within the District of Columbia, Guam, the Virgin Islands of the United States, or any other territory or possession of the United States. Occupational exposure. Any reason ably anticipated skin, eye, mucous membrane, parenteral contact, or res piratory aerosol exposure to select agents or toxins that may result from the performance of an employee’s du ties. Overlap select agent and/or toxin. A bi ological agent or toxin that is listed in § 121.4 and 42 CFR 73.4. Permit. A written authorization by the Administrator to import or move interstate select agents or toxins, under conditions prescribed by the Ad ministrator. Principal investigator. The one indi vidual who is designated by the entity to direct a project or program and who is responsible to the entity for the sci entific and technical direction of that project or program. Proficiency testing. The process of de termining the competency of an indi vidual or laboratory to perform a speci fied test or procedure. Recombinant nucleic acids. (1) Mol ecules that are constructed by joining nucleic acid molecules and that can replicate in a living cell; or VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00844 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 835 Animal and Plant Health Inspection Service, USDA § 121.3 (2) Molecules that result from the replication of those described in para graph (1) of this definition. Responsible official. The individual designated by an entity with the au thority and control to ensure compli ance with the regulations in this part. Security barrier. A physical structure that is designed to prevent entry by unauthorized persons. Select agent and/or toxin. Unless other wise specified, all of the biological agents or toxins listed in §§ 121.3 and 121.4. Specimen. Samples of material from humans, animals, plants, or the envi ronment, or isolates or cultures from such samples, for diagnosis, verification, or proficiency testing. State. Any of the several States of the United States, the Commonwealth of the Northern Mariana Islands, the Commonwealth of Puerto Rico, the District of Columbia, Guam, the Virgin Islands of the United States, or any other territory or possession of the United States. Synthetic nucleic acids. (1) Molecules that are chemically or by other means synthesized or amplified, including those that are chemically or otherwise modified but can base pair with natu rally occurring nucleic acid molecules (i.e., synthetic nucleic acids); or (2) Molecules that result from the replication of those described in para graph (1) of this definition. Toxin. The toxic material or product of plants, animals, microorganisms (in cluding, but not limited to, bacteria, viruses, fungi, or protozoa), or infec tious substances, or a recombinant or synthesized molecule, whatever their origin and method of production, and includes: (1) Any poisonous substance or bio logical product that may be engineered as a result of biotechnology produced by a living organism; or (2) Any poisonous isomer or biologi cal product, homolog, or derivative of such a substance. United States. All of the States. USDA. The U.S. Department of Agri culture. Validated inactivation procedure. A procedure, whose efficacy is confirmed by data generated from a viability test ing protocol, to render a select agent non-viable but allows the select agent to retain characteristics of interest for future use; or to render any nucleic acids that can produce infectious forms of any select agent virus non-infectious for future use. Verification. The demonstration of ob taining established performance (e.g., accuracy, precision, and the analytical sensitivity and specificity) specifica tions for any procedure used for diag nosis. Viability testing protocol. A protocol to confirm the validated inactivation pro cedure by demonstrating the material is free of all viable select agent. VS. The Veterinary Services Pro grams of the Animal and Plant Health Inspection Service. VS select agent and/or toxin. A biologi cal agent or toxin listed in § 121.3. [70 FR 13284, Mar. 18, 2005, as amended at 77 FR 61077, Oct. 5, 2012; 82 FR 6206, Jan. 19, 2017] § 121.2 Purpose and scope. This part implements the provisions of the Agricultural Bioterrorism Pro tection Act of 2002 setting forth the re quirements for possession, use, and transfer of select agents and toxins. The biological agents and toxins listed in this part have the potential to pose a severe threat to public health and safety, to animal health, or to animal products. Overlap select agents and toxins are subject to regulation by both APHIS and CDC. § 121.3 VS select agents and toxins. (a) Except as provided in paragraphs (d) and (e) of this section, the Adminis trator has determined that the biologi cal agents and toxins listed in this sec tion have the potential to pose a severe threat to animal health or to animal products. The select agents and toxins marked with an asterisk (*) are des ignated as Tier 1 select agents and tox ins and are subject to additional re quirements as listed in this part. (b) VS select agents and toxins: Afri can horse sickness virus; African swine fever virus; Avian influenza virus; Clas sical swine fever virus; *Foot-and mouth disease virus; Goat pox virus; Lumpy skin disease virus; Mycoplasma VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00845 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 836 9 CFR Ch. I (1–1–18 Edition) § 121.3 1 A virulent Newcastle disease virus (avian paramyxovirus serotype 1) has an intracerebral pathogenicity index in day-old chicks (Gallus gallus) of 0.7 or greater or has an amino acid sequence at the fusion (F) pro tein cleavage site that is consistent with vir ulent strains of Newcastle disease virus. A failure to detect a cleavage site that is con sistent with virulent strains does not con firm the absence of a virulent virus. 2 The importation and interstate move ment of VS select agents or toxins listed in paragraphs (c)(1) through (c)(3) of this sec tion may be subject to the permit require ments under part 122 of this subchapter. 3 However, the importation and interstate movement of these nonviable select agents may be subject to the permit requirements under part 122 of this subchapter. capricolum; Mycoplasma mycoides; New castle disease virus; 1 Peste des petits ruminants virus; *Rinderpest virus; Sheep pox virus; Swine vesicular dis ease virus. (c) Genetic elements, recombinant and/or synthetic nucleic acids, and re combinant and/or synthetic organisms: (1) Nucleic acids that can produce in fectious forms of any of the select agent viruses listed in paragraph (b) of this section. 2 (2) Recombinant and/or synthetic nu cleic acids that encode for the func tional forms of any toxin listed in paragraph (b) of this section if the nu cleic acids: (i) Can be expressed in vivo or in vitro; or (ii) Are in a vector or recombinant host genome and can be expressed in vivo or in vitro. (3) VS select agents and toxins listed in paragraph (b) of this section that have been genetically modified. (d) VS select agents or toxins that meet any of the following criteria are excluded from the requirements of this part: (1) Any VS select agent or toxin that is in its naturally occurring environ ment, provided that the agent or toxin has not been intentionally introduced, cultivated, collected, or otherwise ex tracted from its natural source. (2) Nonviable VS select agents or nontoxic VS toxins.3 (3) A select agent or toxin that has been subjected to decontamination or a destruction procedure when intended for waste disposal. (4) A select agent or regulated nu cleic acids that can produce infectious forms of any select agent virus that has been subjected to a validated inac tivation procedure that is confirmed through a viability testing protocol. Surrogate strains that are known to possess equivalent properties with re spect to inactivation can be used to validate an inactivation procedure; however, if there are known strain-to strain variations in the resistance of a select agent to an inactivation proce dure, then an inactivation procedure validated on a lesser resistant strain must also be validated on the more re sistant strains. (5) Material containing a select agent that is subjected to a procedure that removes all viable select agent cells, spores, or virus particles if the mate rial is subjected to a viability testing protocol to ensure that the removal method has rendered the material free of all viable select agent. (6) A select agent or regulated nu cleic acids that can produce infectious forms of any select agent virus not sub jected to a validated inactivation pro cedure or material containing a select agent not subjected to a procedure that removes all viable select agent cells, spores, or virus particles if the mate rial is determined by the Adminis trator to be effectively inactivated or effectively removed. To apply for a de termination an individual or entity must submit a written request and sup porting scientific information to APHIS. A written decision granting or denying the request will be issued. (7) A VS select toxin identified in an original food sample or clinical sample. (8) Waste generated during the deliv ery of patient care by health care pro fessionals from a patient diagnosed with an illness or condition associated with a select agent, where that waste is decontaminated or transferred for destruction by complying with State and Federal regulations within 7 cal endar days of the conclusion of patient care. (9) Any low pathogenic strains of avian influenza virus, avian paramyxovirus serotype-1 (APMV–1) vi ruses which do not meet the criteria VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00846 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 837 Animal and Plant Health Inspection Service, USDA § 121.3 4 An APMV–1 virus isolated from poultry which has an intracerebral pathogenicity index in day-old chicks (Gallus gallus) of 0.7 or greater or has an amino acid sequence at the fusion (F) protein cleavage site that is consistent with virulent strains of Newcastle disease virus. A failure to detect a cleavage site that is consistent with virulent strains does not confirm the absence of a virulent virus. 5 Pigeon paramyxovirus (PPMV–1) is a spe cies-adapted APMV–1 virus which is endemic in pigeons and doves in the United States and can be identified through monoclonal antibody testing and demonstration of their characteristic amino acid signature at the fusion gene cleavage site. for Newcastle disease virus,4 including those identified as pigeon paramyxovirus-12 5 isolated from a non poultry species, all subspecies Myco plasma capricolum except subspecies capripneumoniae (contagious caprine pleuropneumonia), and all subspecies Mycoplasma mycoides except subspecies mycoides small colony (Mmm SC) (con tagious bovine pleuropneumonia), pro vided that the individual or entity can identify that the agent is within the exclusion category. (e) An attenuated strain of a select agent or a select toxin modified to be less potent or toxic may be excluded from the requirements of this part based upon a determination by the Ad ministrator that the attenuated strain or modified toxin does not pose a se vere threat to animal health or animal products. (1) To apply for exclusion, an indi vidual or entity must submit a written request and supporting scientific infor mation. A written decision granting or denying the request will be issued. An exclusion will be effective upon notifi cation to the applicant. Exclusions will be listed on the National Select Agent Registry Web site at http:// www.selectagents.gov/. (2) If an excluded attenuated strain or modified toxin is subjected to any manipulation that restores or enhances its virulence or toxic activity, the re sulting select agent or toxin will be subject to the requirements of this part. (3) An individual or entity may make a written request to the Administrator for reconsideration of a decision deny ing an application for the exclusion of an attenuated strain of a select agent or a select toxin modified to be less po tent or toxic. The written request for reconsideration must state the facts and reasoning upon which the indi vidual or entity relies to show the deci sion was incorrect. The Administrator will grant or deny the request for re consideration as promptly as cir cumstances allow and will state, in writing, the reasons for the decision. (f) Any VS select agent or toxin seized by a Federal law enforcement agency will be excluded from the re quirements of this part during the pe riod between seizure of the agent or toxin and the transfer or destruction of such agent or toxin provided that: (1) As soon as practicable, the Fed eral law enforcement agency transfers the seized agent or toxin to an entity eligible to receive such agent or toxin or destroys the agent or toxin by a rec ognized sterilization or inactivation process. (2) The Federal law enforcement agency safeguards and secures the seized agent or toxin against theft, loss, or release, and reports any theft, loss, or release of such agent or toxin. (3) The Federal law enforcement agency reports the seizure of the select agent or toxin to APHIS or CDC. (i) The seizure of any of the following VS select agents and toxins must be re ported within 24 hours by telephone, facsimile, or e-mail: African horse sickness virus, African swine fever virus, avian influenza virus (highly pathogenic), bovine spongiform encephalopathy agent, classical swine fever virus, foot-and-mouth disease virus, virulent Newcastle disease virus, rinderpest virus, and swine vesicular disease virus. This report must be fol lowed by submission of APHIS/CDC Form 4 within 7 calendar days after seizure of the select agent or toxin. (ii) For all other VS select agents or toxins, APHIS/CDC Form 4 must be submitted within 7 calendar days after seizure of the agent or toxin. (iii) A copy of APHIS/CDC Form 4 must be maintained for 3 years. (4) The Federal law enforcement agency reports the final disposition of the select agent or toxin by submission of APHIS/CDC Form 4. A copy of the VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00847 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 838 9 CFR Ch. I (1–1–18 Edition) § 121.4 6 The importation and interstate move ment of overlap select agents or toxins listed in paragraphs (c)(1) through (c)(3) of this sec tion may be subject to the permit require ments under part 122 of this subchapter. 7 However, the importation and interstate movement of these nonviable overlap select agents may be subject to the permit require ments under part 122 of this subchapter. completed form must be maintained for 3 years. [70 FR 13284, Mar. 18, 2005, as amended at 73 FR 61331, Oct. 16, 2008; 77 FR 61077, Oct. 5, 2012; 79 FR 26830, May 12, 2014; 82 FR 6207, Jan. 19, 2017] § 121.4 Overlap select agents and tox ins. (a) Except as provided in paragraphs (d) and (e) of this section, the Adminis trator has determined that the biologi cal agents and toxins listed in this sec tion have the potential to pose a severe threat to public health and safety, to animal health, or to animal products. The select agents and toxins marked with an asterisk (*) are designated as Tier 1 select agents and toxins and are subject to additional requirements as listed in this part. (b) Overlap select agents and toxins: *Bacillus anthracis; Bacillus anthracis (Pasteur strain); Brucella abortus; Brucella melitensis; Brucella suis; *Burkholderia mallei; *Burkholderia pseudomallei; Hendra virus; Nipah virus; Rift Valley fever virus; Venezuelan equine encephalitis virus. (c) Genetic elements, recombinant and/or synthetic nucleic acids, and re combinant and/or synthetic organisms: (1) Nucleic acids that can produce in fectious forms of any of the overlap se lect agent viruses listed in paragraph (b) of this section. 6 (2) Recombinant and/or synthetic nu cleic acids that encode for the toxic forms of any overlap toxin listed in paragraph (b) of this section if the nu cleic acids: (i) Can be expressed in vivo or in vitro; or (ii) Are in a vector or recombinant host genome and can be expressed in vivo or in vitro. (3) Overlap select agents and toxins listed in paragraph (b) of this section that have been genetically modified. (d) Overlap select agents or toxins that meet any of the following criteria are excluded from the requirements of this part: (1) Any overlap select agent or toxin that is in its naturally occurring envi ronment, provided that the agent or toxin has not been intentionally intro duced, cultivated, collected, or other wise extracted from its natural source. (2) Nonviable overlap select agents or nontoxic overlap toxins.7 (3) A select agent or toxin that has been subjected to decontamination or a destruction procedure when intended for waste disposal. (4) A select agent or regulated nu cleic acids that can produce infectious forms of any select agent virus that has been subjected to a validated inac tivation procedure that is confirmed through a viability testing protocol. Surrogate strains that are known to possess equivalent properties with re spect to inactivation can be used to validate an inactivation procedure; however, if there are known strain-to strain variations in the resistance of a select agent to an inactivation proce dure, then an inactivation procedure validated on a lesser resistant strain must also be validated on the more re sistant strains. (5) Material containing a select agent that is subjected to a procedure that removes all viable select agent cells, spores, or virus particles if the mate rial is subjected to a viability testing protocol to ensure that the removal method has rendered the material free of all viable select agent. (6) A select agent or regulated nu cleic acids that can produce infectious forms of any select agent virus not sub jected to a validated inactivation pro cedure or material containing a select agent not subjected to a procedure that removes all viable select agent cells, spores, or virus particles if the mate rial is determined by the Adminis trator or HHS Secretary to be effec tively inactivated or effectively re moved. To apply for a determination an individual or entity must submit a written request and supporting sci entific information to APHIS or CDC. A written decision granting or denying the request will be issued. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00848 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 839 Animal and Plant Health Inspection Service, USDA § 121.5 (7) An overlap select toxin identified in an original food sample or clinical sample. (8) Waste generated during the deliv ery of patient care by health care pro fessionals from a patient diagnosed with an illness or condition associated with a select agent, where that waste is decontaminated or transferred for destruction by complying with State and Federal regulations within 7 cal endar days of the conclusion of patient care. (9) Any subtypes of Venezuelan equine encephalitis virus except for Subtypes IAB or IC, provided that the individual or entity can identify that the agent is within the exclusion cat egory. (e) An attenuated strain of a select agent or a select toxin modified to be less potent or toxic may be excluded from the requirements of this part based upon a determination by the Ad ministrator that the attenuated strain or modified toxin does not pose a se vere threat to public health and safety, animal health, or animal products. (1) To apply for exclusion, an indi vidual or entity must submit a written request and supporting scientific infor mation. A written decision granting or denying the request will be issued. An exclusion will be effective upon notifi cation to the applicant. Exclusions will be listed on the National Select Agent Registry Web site at http:// www.selectagents.gov/. (2) If an excluded attenuated strain or modified toxin is subjected to any manipulation that restores or enhances its virulence or toxic activity, the re sulting select agent or toxin will be subject to the requirements of this part. (3) An individual or entity may make a written request to the Administrator or HHS Secretary for reconsideration of a decision denying an application for the exclusion of an attenuated strain of a select agent or a select toxin modi fied to be less potent or toxic. The written request for reconsideration must state the facts and reasoning upon which the individual or entity re lies to show the decision was incorrect. The Administrator or HHS Secretary will grant or deny the request for re consideration as promptly as cir cumstances allow and will state, in writing, the reasons for the decision. (f) Any overlap select agent or toxin seized by a Federal law enforcement agency will be excluded from the re quirements of this part during the pe riod between seizure of the agent or toxin and the transfer or destruction of such agent or toxin provided that: (1) As soon as practicable, the Fed eral law enforcement agency transfers the seized agent or toxin to an entity eligible to receive such agent or toxin or destroys the agent or toxin by a rec ognized sterilization or inactivation process. (2) The Federal law enforcement agency safeguards and secures the seized agent or toxin against theft, loss, or release, and reports any theft, loss, or release of such agent or toxin. (3) The Federal law enforcement agency reports the seizure of the over lap select agent or toxin to APHIS or CDC. (i) The seizure of any of the following overlap select agents and toxins must be reported within 24 hours by tele phone, facsimile, or e-mail: Bacillus anthracis, Burkholderia mallei, or Burkholderia pseudomallei. This report must be followed by submission of APHIS/CDC Form 4 within 7 calendar days after seizure of the overlap select agent or toxin. (ii) For all other overlap select agents or toxins, APHIS/CDC Form 4 must be submitted within 7 calendar days after seizure of the agent or toxin. (iii) A copy of APHIS/CDC Form 4 must be maintained for 3 years. (4) The Federal law enforcement agency reports the final disposition of the overlap select agent or toxin by submission of APHIS/CDC Form 4. A copy of the completed form must be maintained for 3 years. [70 FR 13284, Mar. 18, 2005, as amended at 73 FR 61331, Oct. 16, 2008; 77 FR 61078, Oct. 5, 2012; 79 FR 26830, May 12, 2014; 82 FR 6207, Jan. 19, 2017] § 121.5 Exemptions for VS select agents and toxins. (a) Diagnostic laboratories and other entities that possess, use, or transfer a VS select agent or toxin that is con tained in a specimen presented for di agnosis or verification will be exempt VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00849 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 840 9 CFR Ch. I (1–1–18 Edition) § 121.5 from the requirements of this part for such agent or toxin contained in the specimen, provided that: (1) Unless directed otherwise by the Administrator, within 7 calendar days after identification of the select agent or toxin, the select agent or toxin is transferred in accordance with § 121.16 or destroyed on-site by a recognized sterilization or inactivation process; (2) The agent or toxin is secured against theft, loss, or release during the period between identification of the agent or toxin and transfer or de struction of such agent or toxin, and any theft, loss, or release of such agent or toxin is reported; (3) Unless otherwise directed by the Administrator, the clinical or diag nostic specimens collected from a pa tient infected with a select agent are transferred in accordance with § 121.16 or destroyed on-site by a recognized sterilization or inactivation process within 7 calendar days after delivery of patient care by heath care profes sionals has concluded; and (4) The identification of the agent or toxin is reported to APHIS or CDC, the specimen provider, and to other appro priate authorities when required by Federal, State, or local law by tele phone, facsimile, or email. This report must be followed by submission of APHIS/CDC Form 4 to APHIS or CDC within 7 calendar days after identifica tion. (b) Diagnostic laboratories and other entities that possess, use, or transfer a VS select agent or toxin that is con tained in a specimen presented for pro ficiency testing will be exempt from the requirements of this part for such agent or toxin contained in the speci men, provided that: (1) Unless directed otherwise by the Administrator, within 90 calendar days of receipt, the agent or toxin is trans ferred in accordance with § 121.16 or de stroyed on-site by a recognized steri lization or inactivation process; (2) The agent or toxin is secured against theft, loss, or release during the period between identification of the agent or toxin and transfer or de struction of such agent or toxin, and any theft, loss, or release of such agent or toxin is reported; and (3) The identification of the agent or toxin, and its derivative, is reported to APHIS or CDC. To report the identi fication of a select agent or toxin, APHIS/CDC Form 4 must be submitted within 90 days of receipt of the agent or toxin. A copy of the completed form must be maintained for 3 years. (c) Diagnostic reagents and vaccines that are, bear, or contain VS select agents or toxins that are produced at USDA diagnostic facilities will be ex empt from the requirements of this part. (d) Unless the Administrator by order determines that additional regulation is necessary to protect animal health or animal products, products that are, bear, or contain VS select agents or toxins will be exempt from the require ments of this part if the products have been cleared, approved, licensed, or registered pursuant to: (1) The Federal Food, Drug, and Cos metic Act (21 U.S.C. 301 et seq.); (2) Section 351 of Public Health Serv ice Act (42 U.S.C. 262); (3) The Virus-Serum-Toxin Act (21 U.S.C. 151–159); or (4) The Federal Insecticide, Fun gicide, and Rodenticide Act (7 U.S.C. 131 et seq.). (e) The Administrator may exempt from the requirements of this part an experimental product that is, bears, or contains a VS select agent or toxin if such product is being used in an inves tigation authorized by any Federal law and the Administrator determines that additional regulation under this part is not necessary to protect animal health or animal products. To apply for an ex emption, an individual or entity must submit APHIS/CDC Form 5. A written decision granting or denying the ex emption will be issued. The applicant must notify APHIS when an authoriza tion for an investigation no longer ex ists. This exemption automatically ter minates when such authorization is no longer in effect. (f) In addition to the exemptions pro vided in paragraphs (a) through (e) of this section, the Administrator may grant a specific exemption upon a showing of good cause and upon his or her determination that such exemption is consistent with protecting animal VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00850 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 841 Animal and Plant Health Inspection Service, USDA § 121.6 health or animal products. An indi vidual or entity may request in writing an exemption from the requirements of this part. If granted, such exemptions are valid for a maximum of 3 years; thereafter, an individual or entity must request a new exemption. If a re quest for exemption is denied, an indi vidual or entity may request reconsid eration in writing to the Adminis trator. The request for reconsideration must state all of the facts and reasons upon which the individual or entity re lies to show that the exemption was wrongfully denied. The Administrator will grant or deny the request for re consideration as promptly as cir cumstances allow and will state, in writing, the reasons for the decision. [70 FR 13284, Mar. 18, 2005, as amended at 73 FR 61331, Oct. 16, 2008; 77 FR 61078, Oct. 5, 2012; 79 FR 26831, May 12, 2014; 82 FR 6208, Jan. 19, 2017] § 121.6 Exemptions for overlap select agents and toxins. (a) Clinical or diagnostic laboratories and other entities that possess, use, or transfer an overlap select agent or toxin that is contained in a specimen presented for diagnosis or verification will be exempt from the requirements of this part for such agent or toxin con tained in the specimen, provided that: (1) Unless directed otherwise by the Administrator, within 7 calendar days after identification of the select agent or toxin, the select agent or toxin is transferred in accordance with § 121.16 or destroyed on-site by a recognized sterilization or inactivation process; (2) The agent or toxin is secured against theft, loss, or release during the period between identification of the agent or toxin and transfer or de struction of such agent or toxin, and any theft, loss, or release of such agent or toxin is reported; (3) Unless otherwise directed by the Administrator or HHS Secretary, the clinical or diagnostic specimens col lected from a patient infected with a select agent are transferred in accord ance with § 121.16 or destroyed on-site by a recognized sterilization or inac tivation process within 7 calendar days after delivery of patient care by heath care professionals has concluded; and (4) The identification of the agent or toxin is reported to APHIS or CDC, the specimen provider, and to other appro priate authorities when required by Federal, State, or local law by tele phone, facsimile, or email. This report must be followed by submission of APHIS/CDC Form 4 to APHIS or CDC within 7 calendar days after identifica tion. (b) Clinical or diagnostic laboratories and other entities that possess, use, or transfer an overlap select agent or toxin that is contained in a specimen presented for proficiency testing will be exempt from the requirements of this part for such agent or toxin con tained in the specimen, provided that: (1) Unless directed otherwise by the Administrator or the HHS Secretary, within 90 days of receipt, the agent or toxin is transferred in accordance with § 121.16 or 42 CFR 73.16 or destroyed on site by a recognized sterilization or in activation process; (2) The agent or toxin is secured against theft, loss, or release during the period between identification of the agent or toxin and transfer or de struction of such agent or toxin, and any theft, loss, or release of such agent or toxin is reported; and (3) The identification of the agent or toxin, and its derivative, is reported to APHIS or CDC. To report the identi fication of an overlap select agent or toxin, APHIS/CDC Form 4 must be sub mitted within 90 calendar days of re ceipt of the agent or toxin. A copy of the completed form must be main tained for 3 years. (c) Unless the Administrator by order determines that additional regulation of a specific product is necessary to protect animal health or animal prod ucts, products that are, bear, or con tain overlap select agents or toxins will be exempt from the requirements of this part if the products have been cleared, approved, licensed, or reg istered pursuant to: (1) The Federal Food, Drug, and Cos metic Act (21 U.S.C. 301 et seq.); (2) Section 351 of Public Health Serv ice Act (42 U.S.C. 262); (3) The Virus-Serum-Toxin Act (21 U.S.C. 151–159); or VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00851 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 842 9 CFR Ch. I (1–1–18 Edition) § 121.7 8 These conditions may apply to more than one individual. (4) The Federal Insecticide, Fun gicide, and Rodenticide Act (7 U.S.C. 131 et seq.). (d) After consultation with the HHS Secretary, the Administrator may ex empt from the requirements of this part an investigational product that is, bears, or contains an overlap select agent or toxin if such product is being used in an investigation authorized by any Federal law and the Administrator determines that additional regulation under this part is not necessary to pro tect animal health or animal products. (1) To apply for an exemption, an in dividual or entity must submit APHIS/ CDC Form 5. (2) The Administrator will make a determination regarding an exemption within 14 calendar days after receipt of the application and notification that the investigation has been authorized under a Federal law. A written decision granting or denying the exemption will be issued. (3) The applicant must notify APHIS or CDC when an authorization for an investigation no longer exists. This ex emption automatically terminates when such authorization is no longer in effect. (e) If it is necessary to respond to a domestic or foreign agricultural emer gency involving an overlap select agent or toxin, the Administrator may ex empt an individual or entity from the requirements, in whole or in part, of this part for up to 30 calendar days. The Administrator may extend the ex emption once for an additional 30 days. (f) Upon request of the Secretary of Health and Human Services, the Ad ministrator may exempt an individual or entity from the requirements, in whole or in part, of this part for up to 30 calendar days if the Secretary of Health and Human Services has grant ed an exemption for a public health emergency involving an overlap select agent or toxin. The Administrator may extend the exemption once for an addi tional 30 days. [70 FR 13284, Mar. 18, 2005, as amended at 73 FR 61331, Oct. 16, 2008; 77 FR 61078, Oct. 5, 2012; 79 FR 26831, May 12, 2014; 82 FR 6208, Jan. 19, 2017] § 121.7 Registration and related secu rity risk assessments. (a) Unless exempted under § 121.5, an individual or entity shall not possess, use, or transfer any VS select agent or toxin without a certificate of registra tion issued by the Administrator. Un less exempted under § 121.6 or 42 CFR 73.6, an individual or entity shall not possess, use, or transfer any overlap se lect agent or toxin without a certifi cate of registration issued by the Ad ministrator and the HHS Secretary. (b) As a condition of registration, each entity is required to be in compli ance with the requirements of this part for select agents and toxins listed on the registration regardless of whether the entity is in actual possession of the select agent or toxin. With regard to toxins, the entity registered for posses sion, use, or transfer of a toxin must be in compliance with the requirements of this part regardless of the amount of toxins currently in its possession. (c) As a condition of registration, each entity must designate an indi vidual to be its responsible official. While most registrants are likely to be entities, in the event that an indi vidual applies for and is granted a cer tificate of registration, the individual will be considered the responsible offi cial. (d)(1) As a condition of registration, the following must be approved by the Administrator or the HHS Secretary based on a security risk assessment by the Attorney General: (i) The individual or entity; (ii) The responsible official; and (iii) Unless otherwise exempted under this section, any individual who owns or controls the entity. (2) Federal, State, or local govern mental agencies, including public ac credited academic institutions, are ex empt from the security risk assess ments for the entity and the individual who owns or controls such entity. (3) An individual will be deemed to own or control an entity under the fol lowing conditions: 8 (i) For a private institution of higher education, an individual will be deemed VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00852 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 843 Animal and Plant Health Inspection Service, USDA § 121.7 9 Depending on the change, a security risk assessment by the Attorney General may also be required (e.g., replacement of the re sponsible official, changes in ownership or control of the entity, new researchers or graduate students, etc.) to own or control the entity if the indi vidual is in a managerial or executive capacity with regard to the entity’s se lect agents or toxins or with regard to the individuals with access to the se lect agents or toxins possessed, used, or transferred by the entity. (ii) For entities other than institu tions of higher education, an individual will be deemed to own or control the entity if the individual: (A) Owns 50 percent or more of the entity, or is a holder or owner of 50 per cent or more of its voting stock; or (B) Is in a managerial or executive capacity with regard to the entity’s se lect agents or toxins or with regard to the individuals with access to the se lect agents or toxins possessed, used, or transferred by the entity. (4) An entity will be considered to be an institution of higher education if it is an institution of higher education as defined in section 101(a) of the Higher Education Act of 1965 (20 U.S.C. 1001(a)), or is an organization described in 501(c)(3) of the Internal Revenue Code of 1986, as amended (26 U.S.C. 501(c)(3)). (5) To obtain a security risk assess ment, an individual or entity must sub mit the information necessary to con duct a security risk assessment to the Attorney General. (e) To apply for a certificate of reg istration for only VS select agents or toxins, or for VS and PPQ select agents or toxins, an individual or entity must submit the information requested in the registration application package (APHIS/CDC Form 1) to APHIS. To apply for a certificate of registration for overlap select agents or toxins, overlap select agents or toxins and any combination of PPQ or VS select agents or toxins, or HHS select agents or toxins and any combination of PPQ or VS select agents or toxins, an indi vidual or entity must submit the infor mation requested in the registration application package (APHIS/CDC Form 1) to APHIS or CDC, but not both. (f) Prior to the issuance of a certifi cate of registration, the responsible of ficial must promptly provide notifica tion of any changes to the application for registration by submitting the rel evant page(s) of the registration appli cation. (g) The issuance of a certificate of registration may be contingent upon inspection or submission of additional information, such as the security plan, biosafety plan, incident response plan, or any other documents required to be prepared under this part. (h) A certificate of registration will be valid for one physical location (a room, a building, or a group of build ings) where the responsible official will be able to perform the responsibilities required in this part, for specific select agents or toxins, and for specific ac tivities. (i) A certificate of registration may be amended to reflect changes in cir cumstances (e.g., replacement of the responsible official or other personnel changes, changes in ownership or con trol of the entity, changes in the ac tivities involving any select agents or toxins, or the addition or removal of select agents or toxins). (1) Prior to any change, the respon sible official must apply for an amend ment to a certificate of registration by submitting the relevant page(s) of the registration application. 9 (2) The responsible official will be no tified in writing if an application to amend a certificate of registration has been approved. Approval of an amend ment may be contingent upon an in spection or submission of additional in formation, such as the security plan, biosafety plan, incident response plan, or any other documents required to be prepared under this part. (3) No change may be made without such approval. (j) An entity must immediately no tify APHIS or CDC if it loses the serv ices of its responsible official. In the event that an entity loses the services of its responsible official, an entity may continue to possess or use select agents or toxins only if it appoints as the responsible official another indi vidual who has been approved by the Administrator or the HHS Secretary following a security risk assessment by VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00853 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 844 9 CFR Ch. I (1–1–18 Edition) § 121.8 10 If registration is denied for this reason, we may provide technical assistance and guidance. the Attorney General and who meets the requirements of this part. (k) A certificate of registration will be terminated upon the written request of the entity if the entity no longer possesses or uses any select agents or toxins and no longer wishes to be reg istered. (l) A certificate of registration will be valid for a maximum of 3 years. [70 FR 13284, Mar. 18, 2005, as amended at 73 FR 61331, Oct. 16, 2008; 82 FR 6208, Jan. 19, 2017] § 121.8 Denial, revocation, or suspen sion of registration. (a) An application may be denied or a certificate of registration revoked or suspended if: (1) The individual or entity, the re sponsible official, or an individual who owns or controls the entity is within any of the categories described in 18 U.S.C. 175b; (2) The individual or entity, the re sponsible official, or an individual who owns or controls the entity is reason ably suspected by any Federal law en forcement or intelligence agency of: (i) Committing a crime set forth in 18 U.S.C. 2332b(g)(5); or (ii) Knowing involvement with an or ganization that engages in domestic or international terrorism (as defined in 18 U.S.C. 2331) or with any other orga nization that engages in intentional crimes of violence; or (iii) Being an agent of a foreign power as defined in 50 U.S.C. 1801; (3) The individual or entity does not meet the requirements of this part; 10 or (4) It is determined that such action is necessary to protect animal health or animal products. (b) Upon revocation or suspension of a certificate of registration, the indi vidual or entity must: (1) Immediately stop all use of each select agent or toxin covered by the revocation or suspension order; (2) Immediately safeguard and secure each select agent or toxin covered by the revocation or suspension order from theft, loss, or release; and (3) Comply with all disposition in structions issued by the Administrator for each select agent or toxin covered by the revocation or suspension. (c) Denial of an application for reg istration and revocation of registration may be appealed under § 121.20. How ever, any denial of an application for registration or revocation of a certifi cate of registration will remain in ef fect until a final agency decision has been rendered. [70 FR 13284, Mar. 18, 2005, as amended at 73 FR 61331, Oct. 16, 2008; 82 FR 6208, Jan. 19, 2017] § 121.9 Responsible official. (a) An individual or entity required to register under this part must des ignate an individual to be the respon sible official. The responsible official must. (1) Be approved by the Administrator or the HHS Secretary following a secu rity risk assessment by the Attorney General. (2) Be familiar with the requirements of this part. (3) Have authority and responsibility to act on behalf of the entity. (4) Ensure compliance with the re quirements of this part. (5) Have a physical (and not merely a telephonic or audio/visual) presence at the registered entity to ensure that the entity is in compliance with the select agent regulations and be able to re spond in a timely manner to onsite in cidents involving select agents and tox ins in accordance with the entity’s in cident response plan. (6) Ensure that annual inspections are conducted for each registered space where select agents or toxins are stored or used in order to determine compliance with the requirements of this part. The results of each inspec tion must be documented, and any defi ciencies identified during an inspection must be corrected and the corrections documented. (7) Ensure that individuals are pro vided the contact information for the USDA Office of Inspector General Hot line and the HHS Office of Inspector General Hotline so that they may anonymously report any biosafety/bio containment or security concerns re lated to select agents and toxins. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00854 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 845 Animal and Plant Health Inspection Service, USDA § 121.10 (8) Investigate to determine the rea son for any failure of a validated inac tivation procedure or any failure to re move viable select agent from mate rial. If the responsible official is unable to determine the cause of a deviation from a validated inactivation proce dure or a viable select agent removal method; or receives any report of any inactivation failure after the move ment of material to another location, the responsible official must report im mediately by telephone or email the inactivation or viable agent removal method failure to APHIS or CDC. (9) Review, and revise as necessary, each of the entity’s validated inactiva tion procedures or viable select agent removal methods. The review must be conducted annually or after any change in principal investigator, change in the validated inactivation procedure or viable select agent re moval method, or failure of the vali dated inactivation procedure or viable select agent removal method. The re view must be documented and training must be conducted if there are any changes to the validated inactivation procedure, viable select agent removal method, or viability testing protocol. (b) An entity may designate one or more individuals to serve as an alter nate responsible official who acts for the responsible official in his/her ab sence. These individuals must have the authority and control to ensure com pliance with the regulations when act ing as the responsible official. (c) The responsible official must re port the identification and final dis position of any select agent or toxin contained in a specimen presented for diagnosis or verification. (1) The identification of any of the following select agents or toxins must be immediately reported by telephone, facsimile, or email: African horse sick ness virus, African swine fever virus, avian influenza virus (highly patho genic), Bacillus anthracis, Burkholderia mallei, Burkholderia pseudomallei, clas sical swine fever virus, foot-and-mouth disease virus, virulent Newcastle dis ease virus, rinderpest virus, or swine vesicular disease virus. The final dis position of the agent or toxin must be reported by submission of APHIS/CDC Form 4 within 7 calendar days after identification. A copy of the completed form must be maintained for 3 years. (2) To report the identification and final disposition of any other select agent or toxin, APHIS/CDC Form 4 must be submitted within 7 calendar days after identification. A copy of the completed form must be maintained for 3 years. (3) Less stringent reporting may be required during agricultural emer gencies or outbreaks, or in endemic areas. (d) The responsible official must re port the identification and final dis position of any select agent or toxin contained in a specimen presented for proficiency testing. To report the iden tification and final disposition of a se lect agent or toxin, APHIS/CDC Form 4 must be submitted within 90 calendar days of receipt of the agent or toxin. A copy of the completed form must be maintained for 3 years. [70 FR 13284, Mar. 18, 2005, as amended at 73 FR 61332, Oct. 16, 2008; 77 FR 61078, Oct. 5, 2012; 79 FR 26831, May 12, 2014; 82 FR 6208, Jan. 19, 2017] § 121.10 Restricting access to select agents and toxins; security risk as sessments. (a) An individual or entity required to register under this part may not provide an individual access to a select agent or toxin, and an individual may not access a select agent or toxin, un less the individual is approved by the Administrator or the HHS Secretary following a security risk assessment by the Attorney General. (b) An individual will be deemed to have access at any point in time if the individual has possession of a select agent or toxin (e.g., carries, uses, or manipulates) or the ability to gain pos session of a select agent or toxin. (c) Each individual with access to se lect agents or toxins must have the ap propriate education, training, and/or experience to handle or use such agents or toxins. (d) To apply for access approval, each individual must submit the informa tion necessary to conduct a security risk assessment to the Attorney Gen eral. (e) A person with valid approval from the HHS Secretary or Administrator to VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00855 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 846 9 CFR Ch. I (1–1–18 Edition) § 121.11 have access to select agents or toxins may request, through his or her Re sponsible Official, that the HHS Sec retary or Administrator provide their approved access status to another reg istered individual or entity for a speci fied period of time. A responsible offi cial must immediately notify the re sponsible official of the visited entity if the person’s access to select agents and toxins has been terminated. (f) An individual’s security risk as sessment may be expedited upon writ ten request by the responsible official and a showing of good cause (e.g., pub lic health or agricultural emergencies, national security, or a short-term visit by a prominent researcher). A written decision granting or denying the re quest will be issued. (g) An individual’s access approval for VS select agents or toxins may be denied, limited, or revoked if: (1) The individual is within any of the categories described in 18 U.S.C. 175b; (2) The individual is reasonably sus pected by any Federal law enforcement or intelligence agency of committing a crime set forth in 18 U.S.C. 2332b(g)(5); knowing involvement with an organi zation that engages in domestic or international terrorism (as defined in 18 U.S.C. 2331) or with any other orga nization that engages in intentional crimes of violence; or being an agent of a foreign power as defined in 50 U.S.C. 1801; or (3) It is determined that such action is necessary to protect animal health or animal products. (h) For overlap select agents or tox ins, an individual’s access approval will be denied or revoked if the individual is within any of the categories de scribed in 18 U.S.C. 175b. An individ ual’s access approval may be denied, limited, or revoked for the reasons set forth in paragraphs (f)(2) through (f)(3) of this section. (i) An individual may appeal the Ad ministrator’s decision to deny, limit, or revoke access approval under § 121.20. (j) Access approval is valid for a max imum of 3 years. (k) The responsible official must im mediately notify APHIS or CDC when an individual’s access to select agents or toxins is terminated by the entity and the reasons therefore. [70 FR 13284, Mar. 18, 2005, as amended at 77 FR 61079, Oct. 5, 2012; 82 FR 6209, Jan. 19, 2017] § 121.11 Security. (a) An individual or entity required to register under this part must de velop and implement a written security plan. The security plan must be suffi cient to safeguard the select agent or toxin against unauthorized access, theft, loss, or release. (b) The security plan must be de signed according to a site-specific risk assessment and must provide graded protection in accordance with the risk of the select agent or toxin, given its intended use. A current security plan must be submitted for initial registra tion, renewal of registration, or when requested. (c) The security plan must: (1) Describe procedures for physical security, inventory control, and infor mation systems control; (2) Contain provisions for the control of access to select agents and toxins, including the safeguarding of animals (including arthropods) or plants inten tionally or accidentally exposed to or infected with a select agent, against unauthorized access, theft, loss or re lease. (3) Contain provisions for routine cleaning, maintenance, and repairs; (4) Establish procedures for removing unauthorized or suspicious persons; (5) Describe procedures for addressing loss or compromise of keys, keycards, passwords, combinations, etc. and pro tocols for changing access permissions or locks following staff changes; (6) Contain procedures for reporting unauthorized or suspicious persons or activities, loss or theft of select agents or toxins, release of select agents or toxins, or alteration of inventory records; (7) Contain provisions for ensuring that all individuals with access ap proval from the Administrator or the HHS Secretary understand and comply with the security procedures. (8) Describe procedures for how the responsible official will be informed of suspicious activity that may be crimi nal in nature and related to the entity, VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00856 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 847 Animal and Plant Health Inspection Service, USDA § 121.11 its personnel, or its select agents or toxins; and describe procedures for how the entity will notify the appropriate Federal, State, or local law enforce ment agencies of such activity. (9) Contain provisions for informa tion security that: (i) Ensure that all external connec tions to systems which manage secu rity for the registered space are iso lated or have controls that permit only authorized and authenticated users; (ii) Ensure that authorized and au thenticated users are only granted ac cess to select agent and toxin related information, files, equipment (e.g., servers or mass storage devices), and applications as necessary to fulfill their roles and responsibilities, and that access is modified when the user’s roles and responsibilities change or when their access to select agents and toxins is suspended or revoked; (iii) Ensure that controls are in place that are designed to prevent malicious code (such as, but not limited to, com puter viruses, worms, spyware) from compromising the confidentiality, in tegrity, or availability of information systems which manage access to spaces registered under this part or records as specified in § 121.17; (iv) Establish a robust configuration management practice for information systems to include regular patching and updates made to operating systems and individual applications; and (v) Establish procedures that provide backup security measures in the event that access control systems, surveil lance devices, and/or systems that manage the requirements of § 121.17 are rendered inoperable. (10) Contain provisions and policies for shipping, receiving, and storage of select agents and toxins, including doc umented procedures for receiving, monitoring, and shipping of all select agents and toxins. These provisions must provide that an entity will prop erly secure containers on site and have a written contingency plan for unex pected shipments. (d) An individual or entity must ad here to the following security require ments or implement measures to achieve an equivalent or greater level of security: (1) Allow access only to individuals with access approval from the Adminis trator or the HHS Secretary; (2) Allow individuals not approved for access by the Administrator or the HHS Secretary to conduct routine cleaning, maintenance, repairs, and other activities not related to select agents or toxins only when continu ously escorted by an approved indi vidual if the potential to access to se lect agents or toxins exists; (3) Provide for the control of select agents and toxins by requiring freezers, refrigerators, cabinets, and other con tainers where select agents or toxins are stored to be secured against unau thorized access (e.g., card access sys tem, lock boxes); (4) Inspect all suspicious packages before they are brought into or re moved from an area where select agents or toxins are used or stored; (5) Establish a protocol for intra-en tity transfers under the supervision of an individual with access approval from the Administrator or the HHS Secretary, including chain-of-custody documents and provisions for safe guarding against theft, loss, or release; and (6) Require that individuals with ac cess approval from the Administrator or the HHS Secretary refrain from sharing with any other person their unique means of accessing a select agent or toxin (e.g., keycards or pass words); (7) Require that individuals with ac cess approval from the Administrator or the HHS Secretary immediately re port any of the following to the respon sible official: (i) Any loss or compromise of keys, passwords, combinations, etc.; (ii) Any suspicious persons or activi ties; (iii) Any loss or theft of select agents or toxins; (iv) Any release of a select agent or toxin; (v) Any sign that inventory or use records for select agents or toxins have been altered or otherwise com promised; and (vi) Any loss of computer, hard drive or other data storage device containing information that could be used to gain access to select agents or toxins; and VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00857 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 848 9 CFR Ch. I (1–1–18 Edition) § 121.11 (8) Separate areas where select agents and toxins are stored or used from the public areas of the building. (e) Entities must conduct complete inventory audits of all affected select agents and toxins in long-term storage when any of the following occur: (1) Upon the physical relocation of a collection or inventory of select agents or toxins for those select agents or tox ins in the collection or inventory; (2) Upon the departure or arrival of a principal investigator for those select agents and toxins under the control of that principal investigator; or (3) In the event of a theft or loss of a select agent or toxin, all select agents and toxins under the control of that principal investigator. (f) In addition to the requirements contained in paragraphs (c) and (d) of this section, the security plan for an individual or entity possessing a Tier 1 select agent or toxin must also: (1) Describe procedures for con ducting a pre-access suitability assess ment of persons who will have access to a Tier 1 select agent or toxin; (2) Describe procedures for how an entity’s responsible official will coordi nate their efforts with the entity’s safety and security professionals to en sure security of Tier 1 select agents and toxins and share, as appropriate, relevant information; and (3) Describe procedures for the ongo ing assessment of the suitability of personnel with access to a Tier 1 select agent or toxin. The procedures must include: (i) Self- and peer-reporting of inci dents or conditions that could affect an individual’s ability to safely have ac cess to or work with select agents and toxins, or to safeguard select agents and toxins from theft, loss, or release; (ii) The training of employees with access to Tier 1 select agents and tox ins on entity policies and procedures for reporting, evaluation, and correc tive actions concerning the assessment of personnel suitability; and (iii) The ongoing suitability moni toring of individuals with access to Tier 1 select agents and toxins. (4) Entities with Tier 1 select agents and toxins must prescribe the following security enhancements: (i) Procedures that will limit access to a Tier 1 select agent or toxin to only those individuals who are approved by the HHS Secretary or Administrator following a security risk assessment by the Attorney General, have had an en tity-conducted pre-access suitability assessment, and are subject to the enti ty’s procedures for ongoing suitability assessment; (ii) Procedures that limit access to laboratory and storage facilities out side of normal business hours to only those specifically approved by the re sponsible official or designee; (iii) Procedures for allowing visitors, their property, and vehicles at the entry and exit points to the registered space, or at other designated points of entry to the building, facility, or com pound that are based on the entity’s site-specific risk assessment; (iv) A minimum of three security barriers where each security barrier adds to the delay in reaching secured areas where select agents and toxins are used or stored. One of the security barriers must be monitored in such a way as to detect intentional and unin tentional circumventing of established access control measures under all con ditions (day/night, severe weather, etc.) The final barrier must limit access to the select agent or toxin to personnel approved by the HHS Secretary or Ad ministrator, following a security risk assessment by the Attorney General. (v) All registered space or areas that reasonably afford access to the reg istered space must be protected by an intrusion detection system (IDS) un less physically occupied; (vi) Personnel monitoring the IDS must be capable of evaluating and in terpreting the alarm and alerting the designated security response force or law enforcement; (vii) For powered access control sys tems, describe procedures to ensure that security is maintained in the event of the failure of access control systems due to power disruption affect ing registered space; (viii) The entity must: (A) Determine that the response time for security forces or local police will not exceed 15 minutes where the re sponse time is measured from the time VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00858 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 849 Animal and Plant Health Inspection Service, USDA § 121.12 11 Technical assistance and guidance may be obtained by contacting APHIS. of an intrusion alarm, or report of a se curity incident, to the arrival of the re sponders at the first security barrier or; (B) Provide security barriers that are sufficient to delay unauthorized access until the response force arrives in order to safeguard the select agents and toxins from theft, intentional re lease, or unauthorized access. The re sponse time is measured from the time of an intrusion alarm, or report of a se curity incident, to the arrival of the re sponders at the first security barrier. (5) Entities that possess foot-and mouth disease virus and rinderpest virus must have the following addi tional security requirements: (i) A minimum of four barriers, one of which must be a perimeter security fence or equivalent which is monitored 24 hours a day, 7 days a week (24/7) to detect the presence of unauthorized persons, vehicles, materials, or unau thorized activities; (ii) Onsite 24/7 armed security re sponse force with roving patrol. Re sponse time must not exceed 5 minutes from the time of an intrusion alarm or report of a security incident; (iii) CCTV surveillance with 24/7 mon itoring and recording; and (iv) Transport vehicle with GPS tracking designed to serve as a con tainment vehicle. (g) In developing a security plan, an individual or entity should consider the document entitled, ‘‘Security Guid ance for Select Agent or Toxin Facili ties.’’ This document is available on the National Select Agent Registry at http://www.selectagents.gov/. (h) The plan must be reviewed annu ally and revised as necessary. Drills or exercises must be conducted at least annually to test and evaluate the effec tiveness of the plan. The plan must be reviewed and revised, as necessary, after any drill or exercise and after any incident. Drills or exercises must be documented to include how the drill or exercise tested and evaluated the plan, any problems that were identified and corrective action(s) taken, and the names of registered entity personal participants. [70 FR 13284, Mar. 18, 2005, as amended at 77 FR 61079, Oct. 5, 2012; 79 FR 26831, May 12, 2014; 82 FR 6209, Jan. 19, 2017] § 121.12 Biosafety. (a) An individual or entity required to register under this part must de velop and implement a written bio safety plan that is commensurate with the risk of the select agent or toxin, given its intended use.11 The biosafety plan must contain sufficient informa tion and documentation to describe the biosafety and containment procedures for the select agent or toxin, including any animals (including arthropods) or plants intentionally or accidentally ex posed to or infected with a select agent. The current biosafety plan must be submitted for initial registration, renewal of registration, or when re quested. The biosafety plan must in clude the following provisions: (1) The hazardous characteristics of each agent or toxin listed on the enti ty’s registration and the biosafety risk associated with laboratory procedures related to the select agent or toxin; (2) Safeguards in place with associ ated work practices to protect entity personnel, the public, and the environ ment from exposure to the select agent or toxin including, but not limited to: Personal protective equipment and other safety equipment; containment equipment including, but not limited to, biological safety cabinets, animal caging systems, and centrifuge safety containers; and engineering controls and other facility safeguards; (3) Written procedures for each vali dated method used for disinfection, de contamination, or destruction, as ap propriate, of all contaminated or pre sumptively contaminated materials in cluding, but not limited to: Cultures and other materials related to the propagation of select agents or toxins, items related to the analysis of select agents and toxins, personal protective equipment, animal caging systems and bedding (if applicable), animal car casses or extracted tissues and fluids (if applicable), laboratory surfaces and equipment, and effluent material; and (4) Procedures for the handling of se lect agents and toxins in the same spaces with non-select agents and tox ins to prevent unintentional contami nation. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00859 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 850 9 CFR Ch. I (1–1–18 Edition) § 121.13 12 Nothing in this section is meant to su persede or preempt incident response re quirements imposed by other statutes or reg ulations. 13 Technical assistance and guidance may be obtained by contacting APHIS. (b) The biosafety and containment procedures must be sufficient to con tain the select agent or toxin (e.g., physical structure and features of the entity, and operational and procedural safeguards). (c) In developing a biosafety plan, an individual or entity should consider the following: (1) The CDC/NIH publication, ‘‘Bio safety in Microbiological and Bio medical Laboratories.’’ This document is available on the National Select Agent Registry at http:// www.selectagents.gov/. (2) The ‘‘NIH Guidelines for Research Involving Recombinant or Synthetic Nucleic Acid Molecules.’’ This docu ment is available on the Internet at http://www.selectagents.gov/. (d) The biosafety plan must include an occupational health program for in dividuals with access to Tier 1 select agents and toxins, and those individ uals must be enrolled in the occupa tional health program. (e) The plan must be reviewed annu ally and revised as necessary. Drills or exercises must be conducted at least annually to test and evaluate the effec tiveness of the plan. The plan must be reviewed and revised, as necessary, after any drill or exercise and after any incident. Drills or exercises must be documented to include how the drill or exercise tested and evaluated the plan, any problems that were identified and corrective action(s) taken, and the names of registered entity personnel participants. [70 FR 13284, Mar. 18, 2005, as amended at 73 FR 61331, Oct. 16, 2008; 77 FR 61080, Oct. 5, 2012; 82 FR 6209, Jan. 19, 2017] § 121.13 Restricted experiments. (a) An individual or entity may not conduct, or possess products resulting from, the following experiments unless approved by and conducted in accord ance with the conditions prescribed by the Administrator: (1) Experiments that involve the de liberate transfer of, or selection for, a drug resistance trait to select agents that are not known to acquire the trait naturally, if such acquisition could compromise the control of disease agents in humans, veterinary medicine, or agriculture. (2) Experiments involving the delib erate formation of synthetic or recom binant DNA containing genes for the biosynthesis of select toxins lethal for vertebrates at an LD[50] <100 ng/kg body weight. (b) The Administrator may revoke approval to conduct any of the experi ments in paragraph (a) of this section, or revoke or suspend a certificate of registration, if the individual or entity fails to comply with the requirements of this part. (c) To apply for approval to conduct any of the experiments in paragraph (a) of this section, an individual or entity must submit a written request and sup porting scientific information. A writ ten decision granting or denying the request will be issued. [70 FR 13284, Mar. 18, 2005, as amended at 73 FR 61331, Oct. 16, 2008; 77 FR 61080, Oct. 5, 2012; 79 FR 26831, May 12, 2014] § 121.14 Incident response. 12 (a) An individual or entity required to register under this part must de velop and implement a written inci dent response plan 13 based upon a site specific risk assessment. The incident response plan must be coordinated with any entity-wide plans, kept in the workplace, and available to employees for review. The current incident re sponse plan must be submitted for ini tial registration, renewal of registra tion, or when requested. (b) The incident response plan must fully describe the entity’s response pro cedures for the theft, loss, or release of a select agent or toxin; inventory dis crepancies; security breaches (includ ing information systems); severe weather and other natural disasters; workplace violence; bomb threats and suspicious packages; and emergencies such as fire, gas leak, explosion, power outage, and other natural and man made events. (c) The response procedures must ac count for hazards associated with the select agent or toxin and appropriate VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00860 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 851 Animal and Plant Health Inspection Service, USDA § 121.15 actions to contain such select agent or toxin, including any animals (including arthropods) or plants intentionally or accidentally exposed to or infected with a select agent. (d) The incident response plan must also contain the following information: (1) The name and contact informa tion (e.g., home and work) for the indi vidual or entity (e.g., responsible offi cial, alternate responsible official(s), biosafety officer, etc.); (2) The name and contact informa tion for the building owner and/or man ager, where applicable; (3) The name and contact informa tion for tenant offices, where applica ble; (4) The name and contact informa tion for the physical security official for the building, where applicable; (5) Personnel roles and lines of au thority and communication; (6) Planning and coordination with local emergency responders; (7) Procedures to be followed by em ployees performing rescue or medical duties; (8) Emergency medical treatment and first aid; (9) A list of personal protective and emergency equipment, and their loca tions; (10) Site security and control; (11) Procedures for emergency evacu ation, including type of evacuation, exit route assignments, safe distances, and places of refuge; and (12) Decontamination procedures. (e) Entities with Tier 1 select agents and toxins must have the following ad ditional incident response policies or procedures: (1) The incident response plan must fully describe the entity’s response pro cedures for failure of intrusion detec tion or alarm system; and (2) The incident response plan must describe procedures for how the entity will notify the appropriate Federal, State, or local law enforcement agen cies of suspicious activity that may be criminal in nature and related to the entity, its personnel, or its select agents or toxins. (f) The plan must be reviewed annu ally and revised as necessary. Drills or exercises must be conducted at least annually to test and evaluate the effec tiveness of the plan. The plan must be reviewed and revised, as necessary, after any drill or exercise and after any incident. Drills or exercises must be documented to include how the drill or exercise tested and evaluated the plan, any problems that were identified and corrective action(s) taken, and the names of registered entity personnel participants. [70 FR 13284, Mar. 18, 2005, as amended at 73 FR 61331, Oct. 16, 2008; 77 FR 61080, Oct. 5, 2012; 82 FR 6209, Jan. 19, 2017] § 121.15 Training. (a) An individual or entity required to register under this part must pro vide information and training on bio containment, biosafety, security (in cluding security awareness), and inci dent response to: (1) Each individual with access ap proval from the Administrator or HHS Secretary. The training must address the particular needs of the individual, the work they will do, and the risks posed by the select agents or toxins. The training must be accomplished prior to the individual’s entry into an area where a select agent is handled or stored, or within 12 months of the date the individual was approved by the Ad ministrator or the HHS Secretary for access, whichever is earlier. (2) Each individual not approved for access to select agents and toxins by the Administrator or HHS Secretary before that individual enters areas under escort where select agents or toxins are handled or stored (e.g., lab oratories, growth chambers, animal rooms, greenhouses, storage areas, shipping/receiving areas, production fa cilities, etc.). Training for escorted personnel must be based on the risk as sociated with accessing areas where se lect agents and toxins are used and/or stored. The training must be accom plished prior to the individual’s entry into where select agents or toxins are handled or stored (e.g., laboratories, growth chambers, animal rooms, green houses, storage areas, shipping/receiv ing areas, production facilities, etc.). (b) Entities with Tier 1 select agents and toxins must conduct annual insider threat awareness briefings on how to identify and report suspicious behav iors. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00861 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 852 9 CFR Ch. I (1–1–18 Edition) § 121.16 14 The requirements of this section do not apply to transfers within a registered entity (i.e., the sender and the recipient are covered by the same certificate of registration). (c) Refresher training must be pro vided annually for individuals with ac cess approval from the HHS Secretary or Administrator or at such time as the registered individual or entity sig nificantly amends its security, incident response, or biosafety plans. (d) The responsible official must en sure a record of the training provided to each individual with access to select agents and toxins and each escorted in dividual (e.g., laboratory workers, visi tors, etc.) is maintained. The record must include the name of the indi vidual, the date of the training, a de scription of the training provided, and the means used to verify that the em ployee understood the training. (e) The responsible official must en sure and document that individuals are provided the contact information of the USDA Office of Inspector General Hotline and the HHS Office of Inspec tor General Hotline so that they may anonymously report any safety or se curity concerns related to select agents and toxins. [77 FR 61081, Oct. 5, 2012, as amended at 82 FR 6209, Jan. 19, 2017] § 121.16 Transfers. (a) Except as provided in paragraphs (c) and (d) of this section, a select agent or toxin may only be transferred to individuals or entities registered to possess, use, or transfer that agent or toxin. A select agent or toxin may only be transferred under the conditions of this section and must be authorized by APHIS or CDC prior to the transfer. 14 (b) A transfer may be authorized if: (1) The sender: (i) Has at the time of transfer a cer tificate of registration that covers the particular select agent or toxin to be transferred and meets all the require ments of this part; (ii) Meets the exemption require ments for the particular select agent or toxin to be transferred; or (iii) Is transferring the select agent or toxin from outside of the United States and meets all import require ments. (2) At the time of transfer, the recipi ent has a certificate of registration that includes the particular select agent or toxin to be transferred and meets all of the requirements of this part. (c) A select agent or toxin that is contained in a specimen for proficiency testing may be transferred without prior authorization from APHIS or CDC provided that, at least 7 calendar days prior to the transfer, the sender reports to APHIS or CDC the select agent or toxin to be transferred and the name and address of the recipient. (d) On a case-by-case basis, the Ad ministrator may authorize a transfer of a select agent or toxin not otherwise eligible for transfer under this part under conditions prescribed by the Ad ministrator. (e) To obtain authorization for a transfer, APHIS/CDC Form 2 must be submitted. (f) After authorization is provided by APHIS or CDC, the packaging of the select agent(s) and toxin(s) is per formed by an individual approved by the HHS Secretary or Administrator to have access to select agents and toxins and is in compliance with all applica ble laws concerning packaging. (g) The sender must comply with all applicable laws governing shipping. (h) Transportation in commerce starts when the select agent(s) or toxin(s) are packaged for shipment and ready for receipt by a courier trans porting select agent(s) or toxin(s) and ends when the package is received by the intended recipient who is an indi vidual approved by the HHS Secretary or Administrator to have access to se lect agents and toxins, following a se curity risk assessment by the Attorney General. (i) The recipient must submit a com pleted APHIS/CDC Form 2 within 2 business days of receipt of a select agent or toxin. (j) The recipient must immediately notify APHIS or CDC if the select agent or toxin has not been received within 48 hours after the expected de livery time or if the package con taining the select agent or toxin has been damaged to the extent that a re lease of the select agent or toxin may have occurred. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00862 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 853 Animal and Plant Health Inspection Service, USDA § 121.17 (k) An authorization for a transfer shall be valid only for 30 calendar days after issuance, except that such an au thorization becomes immediately null and void if any facts supporting the au thorization change (e.g., change in the certificate of registration for the send er or recipient, change in the applica tion for transfer). (l) Transfer the amounts only after the transferor uses due diligence and documents that the recipient has a le gitimate need (e.g., prophylactic, pro tective, bona fide research, or other peaceful purpose) to handle or use such toxins. Information to be documented includes, but is not limited, to the re cipient information, toxin and amount transferred, and declaration that the recipient has legitimate purpose to store and use such toxins. [70 FR 13284, Mar. 18, 2005, as amended at 73 FR 61331, Oct. 16, 2008; 77 FR 61081, Oct. 5, 2012; 82 FR 6210, Jan. 19, 2017] § 121.17 Records. (a) An individual or entity required to register under this part must main tain complete records relating to the activities covered by this part. Such records must include: (1) An accurate, current inventory for each select agent (including viral genetic elements, recombinant and/or synthetic nucleic acids, and organisms containing recombinant and/or syn thetic nucleic acids) held in long-term storage (placement in a system de signed to ensure viability for future use, such as in a freezer or lyophilized materials), including: (i) The name and characteristics (e.g., strain designation, GenBank Ac cession number, etc.); (ii) The quantity acquired from an other individual or entity (e.g., con tainers, vials, tubes, etc.), date of ac quisition, and the source; (iii) Where stored (e.g., building, room, and freezer or other storage con tainer); (iv) When moved from storage and by whom and when returned to storage and by whom; (v) The select agent used, purpose of use, and, when applicable, final disposi tion; (vi) Records created under § 121.16 or 42 CFR 73.16 (Transfers); (vii) For intra-entity transfers (send er and the recipient are covered by the same certificate of registration), the select agent, the quantity transferred, the date of transfer, the sender, and the recipient; and (viii) Records created under § 121.19 or 42 CFR 73.19 (Notification of theft, loss, or release); (2) An accurate, current accounting of any animals or plants intentionally or accidentally exposed to or infected with a select agent (including number and species, location, and appropriate disposition); (3) An accurate, current inventory for each toxin held, including: (i) The name and characteristics; (ii) The quantity acquired from an other individual or entity (e.g., con tainers, vials, tubes, etc.), date of ac quisition, and the source; (iii) The initial and current quantity amount (e.g., milligrams, milliliters, grams, etc.); (iv) The toxin used and purpose of use, quantity, date(s) of the use and by whom; (v) Where stored (e.g., building, room, and freezer or other storage container); (vi) When moved from storage and by whom and when returned to storage and by whom, including quantity amount; (vii) Records created under § 121.16 or 42 CFR 73.16 (Transfers); (viii) For intra-entity transfers (sender and the recipient are covered by the same certificate of registra tion), the toxin, the quantity trans ferred, the date of transfer, the sender, and the recipient; (ix) Records created under § 121.19 or 42 CFR 73.19 (Notification of theft, loss, or release); (x) If destroyed, the quantity of toxin destroyed, the date of such action, and by whom. (4) A current list of all individuals that have been granted access approval by the Administrator or the HHS Sec retary; (5) Information about all entries into areas containing select agents or tox ins, including the name of the indi vidual, name of the escort (if applica ble), and the date and time of entry; (6) Accurate, current records created under § 121.9 or 42 CFR 73.9 (Responsible VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00863 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 854 9 CFR Ch. I (1–1–18 Edition) § 121.18 official), § 121.11 or 42 CFR 73.11 (Secu rity), § 121.12 or 42 CFR 73.12 (Bio safety), § 121.14 or 42 CFR 73.14 (Incident response), and § 121.15 or 42 CFR 73.15 (Training); (7) A written explanation of any dis crepancies; and (8) For select agents or material con taining select agents or regulated nu cleic acids that can produce infectious forms of any select agent virus that have been subjected to a validated in activation procedure or a procedure for removal of viable select agent: (i) A written description of the vali dated inactivation procedure or viable select agent removal method used, in cluding validation data; (ii) A written description of the via bility testing protocol used; (iii) A written description of the in vestigation conducted by the entity re sponsible official involving an inactiva tion or viable select agent removal failure and the corrective actions taken; (iv) The name of each individual per forming the validated inactivation or viable select agent removal method; (v) The date(s) the validated inac tivation or viable select agent removal method was completed; (vi) The location where the validated inactivation or viable select agent re moval method was performed; and (vii) A certificate, signed by the prin cipal investigator, that includes the date of inactivation or viable select agent removal, the validated inactiva tion or viable select agent removal method used, and the name of the prin cipal investigator. A copy of the cer tificate must accompany any transfer of inactivated or select agent removed material. (b) The individual or entity must im plement a system to ensure that all records and databases created under this part are accurate and legible, have controlled access, and that their au thenticity may be verified. (c) The individual or entity must promptly produce upon request any in formation that is related to the re quirements of this part but is not oth erwise contained in a record required to be kept by this section. The location of such information may include, but is not limited to, biocontainment certifi cations, laboratory notebooks, institu tional biosafety and/or animal use committee minutes and approved pro tocols, and records associated with oc cupational health and suitability pro grams. All records created under this part must be maintained for 3 years. [70 FR 13284, Mar. 18, 2005, as amended at 77 FR 61081, Oct. 5, 2012; 82 FR 6210, Jan. 19, 2017] § 121.18 Inspections. (a) Without prior notification, APHIS must be allowed to inspect any site at which activities regulated under this part are conducted and must be al lowed to inspect and copy any records relating to the activities covered by this part. (b) Prior to issuing a certificate of registration to an individual or entity, APHIS may inspect and evaluate the premises and records to ensure compli ance with this part. § 121.19 Notification of theft, loss, or release. (a) An individual or entity must im mediately notify APHIS or CDC upon discovery of the theft or loss of a select agent or toxin. Thefts or losses must be reported even if the select agent or toxin is subsequently recovered or the responsible parties are identified. (1) The theft or loss of a select agent or toxin must be reported by telephone, facsimile, or e-mail. The following in formation must be provided: (i) The name of the select agent or toxin and any identifying information (e.g., strain or other characterization information); (ii) An estimate of the quantity sto len or lost; (iii) An estimate of the time during which the theft or loss occurred; (iv) The location (building, room) from which the theft or loss occurred; and (v) The list of Federal, State, or local law enforcement agencies to which the individual or entity reported, or in tends to report, the theft or loss. (2) A completed APHIS/CDC Form 3 must be submitted within 7 calendar days. (b) An individual or entity must im mediately notify APHIS or CDC upon discovery of a release of a select agent VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00864 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 855 Animal and Plant Health Inspection Service, USDA § 122.2 or toxin causing occupational exposure or a release of a select agent or toxin outside of the primary barriers of the biocontainment area. (1) The release of a select agent or toxin must be reported by telephone, facsimile, or e-mail. The following in formation must be provided: (i) The name of the select agent or toxin and any identifying information (e.g., strain or other characterization information); (ii) An estimate of the quantity re leased; (iii) The time and duration of the re lease; (iv) The environment into which the release occurred (e.g., in building or outside of building, waste system); (v) The location (building, room) from which the release occurred; and (vi) The number of individuals poten tially exposed at the entity; (vii) Actions taken to respond to the release; and (viii) Hazards posed by the release. (2) A completed APHIS/CDC Form 3 must be submitted within 7 calendar days. § 121.20 Administrative review. (a) An individual or entity may ap peal a denial, revocation, or suspension of registration under this part. The ap peal must be in writing, state the fac tual basis for the appeal, and be sub mitted to the Administrator within 30 calendar days of the decision. (b) An individual may appeal a de nial, limitation, or revocation of access approval under this part. The appeal must be in writing, state the factual basis for the appeal, and be submitted to the Administrator within 180 cal endar days of the decision. (c) The Administrator’s decision con stitutes final agency action. [77 FR 61081, Oct. 5, 2012] PART 122—ORGANISMS AND VECTORS Sec. 122.1 Definitions. 122.2 Permits required. 122.3 Application for permits. 122.4 Suspension or revocation of permits. AUTHORITY: 7 U.S.C. 8301–8317; 21 U.S.C. 151– 158; 7 CFR 2.22, 2.80, and 371.4. § 122.1 Definitions. The following words, when used in the regulations in this part 122, shall be construed, respectively, to mean: (a) Department. The U.S. Department of Agriculture. (b) Secretary. ‘‘Secretary’’ means the Secretary of Agriculture of the United States, or any officer or employee of the Department to whom authority has heretofore been delegated, or to whom authority may hereafter be delegated, to act in his stead. (c) Administrator. The Administrator, Animal and Plant Health Inspection Service, United States Department of Agriculture, or any person authorized to act for the Administrator. (d) Organisms. All cultures or collec tions of organisms or their derivatives, which may introduce or disseminate any contagious or infectious disease of animals (including poultry). (e) Vectors. All animals (including poultry) such as mice, pigeons, guinea pigs, rats, ferrets, rabbits, chickens, dogs, and the like, which have been treated or inoculated with organisms, or which are diseased or infected with any contagious, infectious, or commu nicable disease of animals or poultry or which have been exposed to any such disease. (f) Permittee. A person who resides in the United States or operates a busi ness establishment within the United States, to whom a permit to import or transport organisms or vectors has been issued under the regulations. (g) Person. Any individual, firm, part nership, corporation, company, society, association, or other organized group of any of the foregoing, or any agent, officer, or employee of any thereof. [31 FR 81, Jan. 5, 1966, as amended at 57 FR 30899, July 13, 1992] § 122.2 Permits required. No organisms or vectors shall be im ported into the United States or trans ported from one State or Territory or the District of Columbia to another State or Territory or the District of Columbia without a permit issued by the Secretary and in compliance with the terms thereof: Provided, That no permit shall be required under this sec tion for importation of organisms for VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00865 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 856 9 CFR Ch. I (1–1–18 Edition) § 122.3 which an import permit has been issued pursuant to part 102 of this sub chapter or for transportation of orga nisms produced at establishments li censed under part 102 of this sub chapter. As a condition of issuance of permits under this section, the per mittee shall agree in writing to observe the safeguards prescribed by the Ad ministrator for public protection with respect to the particular importation or transportation. (Approved by the Office of Management and Budget under control number 0579–0015) [28 FR 7896, Aug. 2, 1963. Redesignated at 31 FR 81, Jan. 5, 1966 and amended at 48 FR 57473, Dec. 30, 1983; 57 FR 30899, July 13, 1992; 59 FR 67134, Dec. 29, 1994] § 122.3 Application for permits. The Secretary may issue, at his dis cretion, a permit as specified in § 122.2 when proper safeguards are set up as provided in § 122.2 to protect the public. Application for such a permit shall be made in advance of shipment, and each permit shall specify the name and ad dress of the consignee, the true name and character of each of the organisms or vectors involved, and the use to which each will be put. (Approved by the Office of Management and Budget under control number 0579–0015) [23 FR 10065, Dec. 23, 1958. Redesignated at 31 FR 81, Jan. 5, 1966 and amended at 48 FR 57473, Dec. 30, 1983; 59 FR 67134, Dec. 29, 1994] § 122.4 Suspension or revocation of permits. (a) Any permit for the importation or transportation of organisms or vectors issued under this part may be formally suspended or revoked after opportunity for hearing has been accorded the per mittee, as provided in part 123 of this subchapter, if the Secretary finds that the permittee has failed to observe the safeguards and instructions prescribed by the Administrator with respect to the particular importation or transpor tation or that such importation or transportation for any other reason may result in the introduction or dis semination from a foreign country into the United States, or from one State, Territory or the District of Columbia to another, of the contagion of any contagious, infectious or commu nicable disease of animals (including poultry). (b) In cases of wilfulness or where the public health, interest or safety so re quires, however, the Secretary may without hearing informally suspend such a permit upon the grounds set forth in paragraph (a) of this section, pending determination of formal pro ceedings under part 123 of this sub chapter for suspension or revocation of the permit. [23 FR 10065, Dec. 23, 1958. Redesignated at 31 FR 81, Jan. 5, 1966, and amended at 57 FR 30899, July 13, 1992] PART 123—RULES OF PRACTICE GOVERNING PROCEEDINGS UNDER THE VIRUS-SERUM-TOXIN ACT AUTHORITY: 7 U.S.C. 8301–8317; 21 U.S.C. 151– 159; 7 CFR 2.22, 2.80, and 371.4. § 123.1 Scope and applicability of rules of practice. The Uniform Rules of Practice for the Department of Agriculture promul gated in subpart H of part 1, subtitle A, title 7, Code of Federal Regulations, are the Rules of Practice applicable to adjudicatory, administrative pro ceedings under the Virus-Serum-Toxin Act. [42 FR 10960, Feb. 25, 1977] PART 124—PATENT TERM RESTORATION Subpart A—General Provisions Sec. 124.1 Scope. 124.2 Definitions. Subpart B—Eligibility Assistance 124.10 APHIS liaison with PTO. Subpart C—Regulatory Review Period 124.20 Patent term extension calculation. 124.21 Regulatory review period determina tion. 124.22 Revision of regulatory review period determination. 124.23 Final action on regulatory review pe riod determination. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00866 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 857 Animal and Plant Health Inspection Service, USDA § 124.10 Subpart D—Due Diligence Petitions 124.30 Filing, format, and content of peti tions. 124.31 Applicant response to petition. 124.32 APHIS action on petition. 124.33 Standard of due diligence. Subpart E—Due Diligence Hearing 124.40 Request for hearing. 124.41 Notice of hearing. 124.42 Hearing procedure. 124.43 Administrative decision. AUTHORITY: 35 U.S.C. 156; 7 CFR 2.22, 2.80, and 371.4. SOURCE: 58 FR 11369, Feb. 25, 1993, unless otherwise noted. Subpart A—General Provisions § 124.1 Scope. (a) This parts sets forth procedures and requirements for APHIS review of applications for the extension of the term of certain patents for veterinary biological products pursuant to 35 U.S.C. 156—Extension of patent term. Responsibilities of APHIS include: (1) Assisting PTO in determining eli gibility for patent term restoration; (2) Determining the length of a prod uct’s regulatory review period; (3) If petitioned, reviewing and ruling on due diligence challenges to APHIS’s regulatory review period determina tions; and (4) Conducting hearings to review ini tial APHIS findings on due diligence challenges. (b) The regulations in this part are designed to be used in conjunction with regulations issued by PTO concerning patent term extension which may be found at 37 CFR 1.710 through 1.791. [58 FR 11369, Feb. 25, 1993, as amended at 64 FR 43045, Aug. 9, 1999] § 124.2 Definitions. Animal and Plant Health Inspection Service (APHIS). The agency in the De partment of Agriculture responsible for licensing veterinary biological prod ucts under the Virus-Serum-Toxin Act. Applicant. Any person who submits an application or an amendment or supplement to an application under 35 U.S.C. 156 seeking extension of the term of a patent. Due diligence petition. A petition sub mitted under § 124.30 of this part. Informal hearing. A hearing that is not subject to the provisions of 5 U.S.C. 554, 556, and 557 and that is conducted as provided in 21 U.S.C. 321(x). License applicant. Any person who, in accordance with part 102 of this chap ter, submits an application to the Ani mal and Plant Health Inspection Serv ice of the U.S. Department of Agri culture for a U.S. Veterinary Biologi cal Product License. Patent. A patent issued by the Patent and Trademark Office of the United States Department of Commerce. Person. Any individual, firm, partner ship, corporation, company, associa tion, educational institution, State or local government agency, or other or ganized group of any of the foregoing, or any agent, officer, or employee of any thereof. PTO. The Patent and Trademark Of fice of the United States Department of Commerce. [58 FR 11369, Feb. 25, 1993, as amended at 68 FR 6346, Feb. 7, 2003] Subpart B—Eligibility Assistance § 124.10 APHIS liaison with PTO. Upon receipt of a copy of an applica tion for extension of the term of a vet erinary biologic patent from PTO, APHIS will assist PTO in determining whether a patent related to a biologi cal product is eligible for patent term extension by: (a)(1) Verifying whether the product was subject to a regulatory review pe riod before its commercial marketing or use; (2) Determining whether the permis sion for commercial marketing or use of the product after the regulatory re view period was the first permitted commercial marketing or use of the product under the provision of law under which such regulatory review pe riod occurred, and, if so, whether it was the first permitted commercial mar keting or use of the veterinary biologi cal product for administration to a food-producing animal; (3) Ascertaining whether the patent term restoration application was sub mitted within 60 days after the product was approved for marketing or use; and VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00867 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 858 9 CFR Ch. I (1–1–18 Edition) § 124.20 (4) Providing such other information as may be necessary and relevant to PTO’s determination of whether a pat ent related to a product is eligible for patent term restoration. (b) APHIS will notify PTO of its find ings in writing, send a copy of this no tification to the applicant, and make a copy available for public inspection in room 1141, South Building, 14th Street and Independence Avenue SW., Wash ington, DC, between 8 a.m. and 4:30 p.m., Monday through Friday, except holidays. Subpart C—Regulatory Review Period § 124.20 Patent term extension calcula tion. (a) As provided in 37 CFR 1.779 of PTO’s regulations, in order to deter mine a product’s regulatory review pe riod, APHIS will review the informa tion in each application to determine the lengths of the following phases of the review period, and will then find their sum: (1) The number of days in the period beginning on the date authorization to prepare an experimental biological product under the Virus-Serum-Toxin Act became effective and ending on the date an application for a license was initially submitted under the Virus Serum-Toxin Act; and (2) The number of days in the period beginning on the date an application for a license was initially submitted for approval under the Virus-Serum Toxin Act and ending on the date such license was issued. (b) A license application is ‘‘initially submitted’’ on the date it contains suf ficient information to allow APHIS to commence review of the application. A product license is issued on the date of the APHIS letter informing the appli cant of the issuance. The issuance of a license releases the product for com mercial marketing or use. § 124.21 Regulatory review period de termination. (a) Not later than 30 days after the receipt of an application from PTO, APHIS shall determine the regulatory review period. Once the regulatory re view period for a product has been de termined, APHIS will notify PTO in writing of the determination, send a copy of the determination to the appli cant, and make a copy available for public inspection in room 1141, South Building, 14th Street and Independence Avenue SW., Washington, DC, between 8 a.m. and 4:30 p.m., Monday through Friday, except holidays. (b) APHIS will also publish a notice of the regulatory review period deter mination in the FEDERAL REGISTER. The notice will include the following: (1) The name of the applicant; (2) The trade name and true name of the product; (3) The number of the patent for which an extension of the term is sought; (4) The approved indications or uses for the product; (5) The regulatory review period de termination, including a statement of the length of each phase of the review period and the dates used in calcu lating each phase. § 124.22 Revision of regulatory review period determination. (a) Any interested person may re quest a revision of the regulatory re view period determination within the 30 day period beginning on its publica tion in the FEDERAL REGISTER. The re quest must be sent to the Director, Center for Veterinary Biologics, Pol icy, Evaluation, and Licensing, 1920 Dayton Avenue, P.O. Box 844, Ames, IA 50010. The request must specify the fol lowing: (1) The identity of the product; (2) The identity of the applicant for patent term restoration; (3) The docket number of the FED ERAL REGISTER notice announcing the regulatory review period determina tion; and (4) The basis for the request for revi sion, including any documentary evi dence. (b) If APHIS decides to revise its prior determination, APHIS will notify PTO of the decision, and will send a copy of notification to the applicant and the person requesting the revision (if different from the applicant) with a request for comments within 10 days of notification. If no comment on the pro posed revision is received, APHIS will VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00868 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 859 Animal and Plant Health Inspection Service, USDA § 124.32 publish the revision in the FEDERAL REGISTER, and include a statement giv ing the reasons for the revision. If com ment is received, APHIS will make a final determination regarding the revi sion based on such comment and will then publish the revision in the FED ERAL REGISTER, giving reasons for its determination. [59 FR 11369, Feb. 25, 1993, as amended at 59 FR 67617, Dec. 30, 1994; 64 FR 43045, Aug. 9, 1999; 75 FR 20773, Apr. 21, 2010] § 124.23 Final action on regulatory re view period determination. APHIS will consider its regulatory review period determination to be final upon expiration of the 180-day period for filing a due diligence petition under § 124.30 unless it receives: (a) New information from PTO records, or APHIS records, that affects the regulatory review period deter mination; (b) A request under § 124.22 for revi sion of the regulatory review period de termination; (c) A due diligence petition filed under § 124.30; or (d) A request for a hearing filed under § 124.40. [58 FR 11369, Feb. 25, 1993; 58 FR 29028, May 18, 1993] Subpart D—Due Diligence Petitions § 124.30 Filing, format, and content of petitions. (a) Any interested person may file a petition with APHIS, no later than 180 days after the publication of a regu latory review period determination under § 124.21, alleging that a license applicant did not act with due dili gence in seeking APHIS approval of the product during the regulatory review period. (b) The petition must be filed with APHIS under the docket number of the FEDERAL REGISTER notice of the agen cy’s regulatory review period deter mination. The petition must contain any additional information required by this subpart. (c) The petition must allege that the applicant failed to act with due dili gence sometime during the regulatory review period and must set forth suffi cient facts to merit an investigation by APHIS of whether the applicant acted with due diligence. (d) The petition must contain a cer tification that the petitioner has served a true and complete copy of the petition on interested parties by cer tified or registered mail (return receipt requested) or by personal delivery. § 124.31 Applicant response to petition. (a) The applicant may file with APHIS a written response to the peti tion no later than 20 days after the ap plicant’s receipt of a copy of the peti tion. (b) The applicant’s response may present additional facts and cir cumstances to address the assertions in the petition, but shall be limited to the issue of whether the applicant acted with due diligence during the regu latory review period. The applicant’s response may include documents that were not in the original patent term extension application. (c) If the applicant does not respond to the petition, APHIS will decide the matter on the basis of the information submitted in the patent term restora tion application, the due diligence peti tion, and APHIS records. § 124.32 APHIS action on petition. (a) Within 90 days after APHIS re ceives a petition filed under § 124.30, the Under Secretary for Marketing and Regulatory Programs shall make a de termination under paragraphs (b) or (c) of this section or under § 124.33 whether the applicant acted with due diligence during the regulatory review period. APHIS will publish its determination in the FEDERAL REGISTER together with factual and legal basis for the deter mination, notify PTO of the determina tion in writing, and send copies of the determination to PTO, the applicant, and the petitioner. (b) APHIS may deny a due diligence petition without considering the mer its of the petition if: (1) The petition is not filed in accord ance with § 124.30; (2) The petition does not contain in formation or allegations upon which APHIS may reasonably determine that VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00869 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 860 9 CFR Ch. I (1–1–18 Edition) § 124.33 the applicant did not act with due dili gence during the applicable regulatory review period; or (3) The petition fails to allege a suffi cient total amount of time during which the applicant did not exercise due diligence so that, even if the peti tion were granted, the petition would not affect the maximum patent term extension which the applicant is enti tled to under 35 U.S.C. 156. [59 FR 11369, Feb. 25, 1993, as amended at 64 FR 43045, Aug. 9, 1999] § 124.33 Standard of due diligence. (a) In determining the due diligence of an applicant, APHIS will examine the facts and circumstances of the ap plicant’s actions during the regulatory review period to determine whether the applicant exhibited the degree of atten tion, continuous directed effort, and timeliness as may reasonably be ex pected from, and are ordinarily exer cised by, a person during a regulatory review period. APHIS will take into consideration all relevant factors, such as the amount of time between the ap proval of an experimental use permit and licensure of the veterinary biologi cal product. (b) For purposes of this Part, the ac tions of the marketing applicant shall be imputed to the applicant for patent term restoration. The actions of an agent, attorney, contractor, employee, licensee, or predecessor in interest of the marketing applicant shall be im puted to the applicant for patent term restoration. Subpart E—Due Diligence Hearing § 124.40 Request for hearing. (a) Any interested person may re quest, within 60 days beginning on the date of publication of a due diligence determination by APHIS in accordance with § 124.32, that APHIS conduct an in formal hearing on the due diligence de termination. (b) The request for a hearing must: (1) Be in writing; (2) Contain the docket number of the FEDERAL REGISTER notice of APHIS’s regulatory review period determina tion; (3) Be delivered to the Director, Cen ter for Veterinary Biologics, Policy, Evaluation, and Licensing, 1920 Dayton Avenue, P.O. Box 844, Ames, IA 50010. (4) Contain a full statement of facts upon which the request for hearing is based; (5) Contain the name, the address, and the principal place of business of the person requesting the hearing; and (6) Contain a certification that the person requesting the hearing has served a true and complete copy of the request upon the petitioner of the due diligence determination and the appli cant for patent term extension by cer tified or registered mail (return receipt requested) or by personal service. (c) The request must state whether the requesting party seeks a hearing not later than 30 days after the date APHIS receives the request, or, at the request of the person making the re quest, not later than 60 days after such date. [58 FR 11369, Feb. 25, 1993, as amended at 59 FR 67617, Dec. 30, 1994; 64 FR 43045, Aug. 9, 1999; 75 FR 20773, Apr. 21, 2010] § 124.41 Notice of hearing. No later than ten days before the hearing, APHIS will notify the request ing party, the applicant, the petitioner, and any other interested person of the date, time, and location of the hearing. § 124.42 Hearing procedure. (a) The presiding officer shall be ap pointed by the Administrator of APHIS from officers and employees of the De partment who have not participated in any action of the Secretary which is the subject of the hearing and who are not directly responsible to an officer or employee of the Department who has participated in any such action. (b) Each party to the hearing shall have the right at all times to be ad vised and accompanied by an attorney. (c) Before the hearing, each party to the hearing shall be given reasonable notice of the matters to be considered at the hearing, including a comprehen sive statement of the basis for the ac tion taken or proposed by the Sec retary which is the subject of the hear ing and any general summary of the in formation which will be presented at the hearing in support of such action. VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00870 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B 861 Animal and Plant Health Inspection Service, USDA § 124.43 (d) At the hearing the parties to the hearing shall have the right to hear a full and complete statement of the ac tion which is the subject of the hearing together with the information and rea sons supporting such action, to con duct reasonable questioning, and to present any oral and written informa tion relevant to such action. (e) The presiding officer in such hear ing shall prepare a written report of the hearing to which shall be attached all written material presented at the hearing. The participants in the hear ing shall be given the opportunity to review and correct or supplement the presiding officer’s report of the hear ing. (f) The Secretary may require the hearing to be transcribed. A party to the hearing shall have the right to have the hearing transcribed at his ex pense. Any transcription of a hearing shall be included in the presiding offi cer’s report of the hearing. (g) The due diligence hearing will be conducted in accordance with rules of practice adopted for the proceeding. APHIS will provide the requesting party, the applicant, and the petitioner with an opportunity to participate as a party in the hearing. The standard of due diligence set forth in § 124.33 will apply at the hearing. The party re questing the due diligence hearing will have the burden of proof at the hear ing. § 124.43 Administrative decision. Within 30 days after completion of the due diligence hearing, the Under Secretary for Marketing and Regu latory Programs, taking into consider ation the recommendation of the Ad ministrator, will affirm or revise the determination made under § 124.32. APHIS will publish the due diligence redetermination in the FEDERAL REG ISTER, notify PTO of the redetermina tion, and send copies of the notice to PTO and the requesting party, the ap plicant, and the petitioner. [59 FR 11369, Feb. 25, 1993, as amended at 64 FR 43045, Aug. 9, 1999] VerDate Sep<11>2014 13:56 Feb 14, 2018 Jkt 244028 PO 00000 Frm 00871 Fmt 8010 Sfmt 8010 Q:\09\9V1.TXT 31kp a yn e o n D S K 5 4 D X V N 1 O F R w ith $ $ JO B