Modification of any particular test method or manufacturing process or the conditions under which it is conducted as required in this part or in the additional standards for specific biological products in parts 620 through 680 of this chapter shall be permitted only under the following conditions:

(a) The applicant presents evidence, in the form of a license application, or a supplement to the application submitted in accordance with § 601.12(b) or (c), demonstrating that the modification will provide assurances of the safety, purity, potency, and effectiveness of the biological product equal to or greater than the assurances provided by the method or process specified in the general standards or additional standards for the biological product; and

(b) Approval of the modification is received in writing from the Director, Center for Biologics Evaluation and Research or the Director, Center for Drug Evaluation and Research.

Tests for potency shall consist of either in vitro or in vivo tests, or both, which have been specifically designed for each product so as to indicate its potency in a manner adequate to satisfy the interpretation of potency given by the definition in § 600.3(s) of this chapter.

(a) The test. Except as provided in paragraph (h) of this section, manufacturers of biological products must perform sterility testing of each lot of each biological product's final container material or other material, as appropriate and as approved in the biologics license application or supplement for that product.

(b) Test requirements. (1) The sterility test must be appropriate to the material being tested such that the material does not interfere with or otherwise hinder the test.

(2) The sterility test must be validated to demonstrate that the test is capable of reliably and consistently detecting the presence of viable contaminating microorganisms.

(3) The sterility test and test components must be verified to demonstrate that the test method can consistently detect the presence of viable contaminating microorganisms.

(c) Written procedures. Manufacturers must establish, implement, and follow written procedures for sterility testing that describe, at a minimum, the following:

(1) The sterility test method to be used;

(i) If culture-based test methods are used, include, at a minimum:

(A) Composition of the culture media;

(B) Growth-promotion test requirements; and

(C) Incubation conditions (time and temperature).

(ii) If non-culture-based test methods are used, include, at a minimum:

(A) Composition of test components;

(B) Test parameters, including acceptance criteria; and

(C) Controls used to verify the method's ability to detect the presence of viable contaminating microorganisms.

(2) The method of sampling, including the number, volume, and size of articles to be tested;

(3) Written specifications for the acceptance or rejection of each lot; and

(4) A statement of any other function critical to the particular sterility test method to ensure consistent and accurate results.

(d) The sample. The sample must be appropriate to the material being tested, considering, at a minimum:

(1) The size and volume of the final product lot;

(2) The duration of manufacturing of the drug product;

(3) The final container configuration and size;

(4) The quantity or concentration of inhibitors, neutralizers, and preservatives, if present, in the tested material;

(5) For a culture-based test method, the volume of test material that results in a dilution of the product that is not bacteriostatic or fungistatic; and

(6) For a non-culture-based test method, the volume of test material that results in a dilution of the product that does not inhibit or otherwise hinder the detection of viable contaminating microorganisms.

(e) Verification. (1) For culture-based test methods, studies must be conducted to demonstrate that the performance of the test organisms and culture media are suitable to consistently detect the presence of viable contaminating microorganisms, including tests for each lot of culture media to verify its growth-promoting properties over the shelf-life of the media.

(2) For non-culture-based test methods, within the test itself, appropriate controls must be used to demonstrate the ability of the test method to continue to consistently detect the presence of viable contaminating microorganisms.

(f) Repeat test procedures. (1) If the initial test indicates the presence of microorganisms, the product does not comply with the sterility test requirements unless a thorough investigation by the quality control unit can ascribe definitively the microbial presence to a laboratory error or faulty materials used in conducting the sterility testing.

(2) If the investigation described in paragraph (f)(1) of this section finds that the initial test indicated the presence of microorganisms due to laboratory error or the use of faulty materials, a sterility test may be repeated one time. If no evidence of microorganisms is found in the repeat test, the product examined complies with the sterility test requirements. If evidence of microorganisms is found in the repeat test, the product examined does not comply with the sterility test requirements.

(3) If a repeat test is conducted, the same test method must be used for both the initial and repeat tests, and the repeat test must be conducted with comparable product that is reflective of the initial sample in terms of sample location and the stage in the manufacturing process from which it was obtained.

(g) Records. The records related to the test requirements of this section must be prepared and maintained as required by §§ 211.167 and 211.194 of this chapter.

(h) Exceptions. Sterility testing must be performed on final container material or other appropriate material as defined in the approved biologics license application or supplement and as described in this section, except as follows:

(1) This section does not require sterility testing for Whole Blood, Cryoprecipitated Antihemophilic Factor, Platelets, Red Blood Cells, Plasma, Source Plasma, Smallpox Vaccine, Reagent Red Blood Cells, Anti-Human Globulin, and Blood Grouping Reagents.

(2) A manufacturer is not required to comply with the sterility test requirements if the Director of the Center for Biologics Evaluation and Research or the Director of the Center for Drug Evaluation and Research, as appropriate, determines that data submitted in the biologics license application or supplement adequately establish that the route of administration, the method of preparation, or any other aspect of the product precludes or does not necessitate a sterility test to assure the safety, purity, and potency of the product.

Products shall be free of extraneous material except that which is unavoidable in the manufacturing process described in the approved biologics license application. In addition, products shall be tested as provided in paragraphs (a) and (b) of this section.

(a)(1) Test for residual moisture. Each lot of dried product shall be tested for residual moisture and shall meet and not exceed established limits as specified by an approved method on file in the biologics license application. The test for residual moisture may be exempted by the Director, Center for Biologics Evaluation and Research or the Director, Center for Drug Evaluation and Research, when deemed not necessary for the continued safety, purity, and potency of the product.

(2) Records. Appropriate records for residual moisture under paragraph (a)(1) of this section shall be prepared and maintained as required by the applicable provisions of §§ 211.188 and 211.194 of this chapter.

(b) Test for pyrogenic substances. Each lot of final containers of any product intended for use by injection shall be tested for pyrogenic substances by intravenous injection into rabbits as provided in paragraphs (b) (1) and (2) of this section: Provided, That notwithstanding any other provision of Subchapter F of this chapter, the test for pyrogenic substances is not required for the following products: Products containing formed blood elements; Cryoprecipitate; Plasma; Source Plasma; Normal Horse Serum; bacterial, viral, and rickettsial vaccines and antigens; toxoids; toxins; allergenic extracts; venoms; diagnostic substances and trivalent organic arsenicals.

(1) Test dose. The test dose for each rabbit shall be at least 3 milliliters per kilogram of body weight of the rabbit and also shall be at least equivalent proportionately, on a body weight basis, to the maximum single human dose recommended, but need not exceed 10 milliliters per kilogram of body weight of the rabbit, except that: (i) Regardless of the human dose recommended, the test dose per kilogram of body weight of each rabbit shall be at least 1 milliliter for immune globulins derived from human blood; (ii) for Streptokinase, the test dose shall be at least equivalent proportionately, on a body weight basis, to the maximum single human dose recommended.

(2) Test procedure, results, and interpretation; standards to be met. The test for pyrogenic substances shall be performed according to the requirements specified in United States Pharmacopeia XX.

(3) Retest. If the lot fails to meet the test requirements prescribed in paragraph (b)(2) of this section, the test may be repeated once using five other rabbits. The temperature rises recorded for all eight rabbits used in testing shall be included in determining whether the requirements are met. The lot meets the requirements for absence of pyrogens if not more than three of the eight rabbits show individual rises in temperature of 0.6 °C or more, and if the sum of the eight individual maximum temperature rises does not exceed 3.7 °C.

The contents of a final container of each filling of each lot shall be tested for identity after all labeling operations shall have been completed. The identity test shall be specific for each product in a manner that will adequately identify it as the product designated on final container and package labels and circulars, and distinguish it from any other product being processed in the same laboratory. Identity may be established either through the physical or chemical characteristics of the product, inspection by macroscopic or microscopic methods, specific cultural tests, or in vitro or in vivo immunological tests.

(a) Ingredients, preservatives, diluents, adjuvants. All ingredients used in a licensed product, and any diluent provided as an aid in the administration of the product, shall meet generally accepted standards of purity and quality. Any preservative used shall be sufficiently nontoxic so that the amount present in the recommended dose of the product will not be toxic to the recipient, and in the combination used it shall not denature the specific substances in the product to result in a decrease below the minimum acceptable potency within the dating period when stored at the recommended temperature. Products in multiple-dose containers shall contain a preservative, except that a preservative need not be added to Yellow Fever Vaccine; Poliovirus Vaccine Live Oral; viral vaccines labeled for use with the jet injector; dried vaccines when the accompanying diluent contains a preservative; or to an Allergenic Product in 50 percent or more volume in volume (v/v) glycerin. An adjuvant shall not be introduced into a product unless there is satisfactory evidence that it does not affect adversely the safety or potency of the product. The amount of aluminum in the recommended individual dose of a biological product shall not exceed:

(1) 0.85 milligrams if determined by assay;

(2) 1.14 milligrams if determined by calculation on the basis of the amount of aluminum compound added; or

(3) 1.25 milligrams determined by assay provided that data demonstrating that the amount of aluminum used is safe and necessary to produce the intended effect are submitted to and approved by the Director, Center for Biologics Evaluation and Research or the Director, Center for Drug Evaluation and Research (see mailing addresses in § 600.2(a) or (b) of this chapter).

(b) Extraneous protein; cell culture produced vaccines. Extraneous protein known to be capable of producing allergenic effects in human subjects shall not be added to a final virus medium of cell culture produced vaccines intended for injection. If serum is used at any stage, its calculated concentration in the final medium shall not exceed 1:1,000,000.

(c) Antibiotics. A minimum concentration of antibiotics, other than penicillin, may be added to the production substrate of viral vaccines.

(d) The Director of the Center for Biologics Evaluation and Research or the Director of the Center for Drug Evaluation and Research may approve an exception or alternative to any requirement in this section. Requests for such exceptions or alternatives must be in writing.

Except as otherwise provided by regulation, no liquid serum or antitoxin shall contain more than 20 percent total solids.

Licensed products may not be combined with other licensed products either therapeutic, prophylactic or diagnostic, except as a license is obtained for the combined product. Licensed products may not be combined with nonlicensable therapeutic, prophylactic, or diagnostic substances except as a license is obtained for such combination.

(a) Storage and maintenance. Cultures used in the manufacture of products shall be stored in a secure and orderly manner, at a temperature and by a method that will retain the initial characteristics of the organisms and insure freedom from contamination and deterioration.

(b) Identity and verification. Each culture shall be clearly identified as to source strain. A complete identification of the strain shall be made for each new stock culture preparation. Primary and subsequent seed lots shall be identified by lot number and date of preparation. Periodic tests shall be performed as often as necessary to verify the integrity of the strain characteristics and freedom from extraneous organisms. Results of all periodic tests for verification of cultures and determination of freedom from extraneous organisms shall be recorded and retained.

(c) Cell lines used for manufacturing biological products - (1) General requirements. Cell lines used for manufacturing biological products shall be:

(i) Identified by history;

(ii) Described with respect to cytogenetic characteristics and tumorigenicity;

(iii) Characterized with respect to in vitro growth characteristics and life potential; and

(iv) Tested for the presence of detectable microbial agents.

(2) Tests. Tests that are necessary to assure the safety, purity, and potency of a product may be required by the Director, Center for Biologics Evaluation and Research or the Director, Center for Drug Evaluation and Research.

(3) Applicability. This paragraph applies to diploid and nondiploid cell lines. Primary cell cultures that are not subcultivated and primary cell cultures that are subsequently subcultivated for only a very limited number of population doublings are not subject to the provisions of this paragraph (c).

(d) Records. The records appropriate for cultures under this section shall be prepared and maintained as required by the applicable provisions of §§ 211.188 and 211.194 of this chapter.