Comparative studies of the in vivo toxin neutralization and the in vitro Vero cell assay methods for use in potency testing of diphtheria component in combined vaccines/toxoids. 1: Standardization of a modified Vero cell assay for toxin-antitoxin titration of immunized guinea-pig sera.
The in vivo Toxin Neutralization (TN) test is currently used as an antibody limit test for regulatory control testing of diphtheria potency in combined vaccines/toxoids. This test requires the use of guinea-pigs for estimating diphtheria antitoxin titre in immunized sera. The acceptability criteria (minimum requirement) for adsorbed pediatric vaccines/toxoids in the TN test are the production of at least 2.0 IU/ml of antitoxin in the serum pool of immunized animals. The feasibility of employing the in vitro Vero cell assay as an alternative limit test to the TN test was investigated. Parallel titration of serum samples from guinea-pigs immunized with DPT (ads) vaccine (pediatric use) showed that although the in vitro antitoxin titre paralleled the in vivo values, a direct correlation between the two methods could not be established. The mean in vitro titres were lower (in terms of IU/ml) than those obtained by the in vivo TN test. This could possibly be explained by the differences between reference serum which is a hyperimmune horse serum and the test sera which are from guinea-pigs which have been immunized only once. The in vitro/in vivo ratio of the antitoxin titre ranged from 0.1 to 0.24 IU/ml with a mean value of 0.17 (SD, 0.06). Hence, for pediatric vaccines, a tentative in vitro limit of 0.3 IU/ml, which would correspond to the currently required limit of 2.0 IU/ml in the in vivo TN test, could be accepted as the minimum requirement for the antitoxin titre in the Vero cell assay.
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Bacterial Products Division, Health Protection Branch, Ottawa, Ontario, Canada.
D000818:Animals; D000917:Antibody Formation; D001681:Biological Assay / Q000379:methods / Q000592:standards*; D003602:Cytotoxicity, Immunologic; D004166:Diphtheria Antitoxin / Q000097:blood; D004168:Diphtheria Toxoid / Q000032:analysis* / Q000592:standards; D015721:Diphtheria-Tetanus-Pertussis Vaccine / Q000032:analysis* / Q000592:standards; D004305:Dose-Response Relationship, Drug; D006168:Guinea Pigs; D009500:Neutralization Tests / Q000379:methods / Q000592:standards; D011786:Quality Control; D012015:Reference Standards; D014709:Vero Cells
D004166:Diphtheria Antitoxin; D004168:Diphtheria Toxoid; D015721:Diphtheria-Tetanus-Pertussis Vaccine