Development, validation and acceptance of alternative methods in the quality control of vaccines: A case report

Information about levels of protection against tetanus is needed both for the assessment of immune status and for the estimation of the potency of batches of tetanus toxoid. Originally, levels of protective antibodies in human serum samples were titrated in an in vivo toxin neutralization (TN) test. Potency testing was based either on an indirect protection test or a direct challenge test. In the former test, rabbits or guinea pigs are immunized and bled, followed by titration of serum samples in a TN test. In the latter test, used in the quality control of tetanus toxoid for human use, protective immunity is induced by vaccination in guinea pigs or mice and subsequently challenging them directly with tetanus toxin. In the mid-1980s, an in vitro model was developed at the National Institute of Public Health and Environmental Protection (RIVM) for estimating levels of tetanus antitoxin in serum samples. This model, the toxin-binding inhibition (ToBI) test, was validated recently for both diagnostic testing and potency testing. Although accurate estimation of antitoxin levels is more important for diagnostic testing than for potency testing, the ToBI test has been adopted for determining the immune status but not for potency testing. One major reason is that there are no official guidelines for titration of human serum samples. By contrast, potency testing is performed in accordance with the monographs of national and international pharmacopoeias, which complicates acceptance for technical and bureaucratic reasons. This paper focuses on the validation studies performed at the RIVM. In particular, attention is paid to the various problems encountered. Suggestions for the role of the European Centre for Validation of Alternative Methods (ECVAM)( *) in the quality control of vaccines are also presented.