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Comparison of in vivo and in vitro methods for determining unitage of diphtheria antitoxin in adsorbed diphtheria-tetanus (DT) and diphtheria-tetanus-pertussis (DTP) vaccines.

In view of the current efforts to find a reliable in vitro method which can suitably act as an alternative for determining the potency of the diphtheria component in a combined vaccine, we have analysed experimental batches by the method proposed by WHO [1] i.e. challenge method in guinea pigs. The same batches were also analysed by the alternative antibody induction method as suggested in the Indian Pharmacopoeia (I.P.) [2] which is similar to the old method suggested in the British Pharmacopoeia (B.P.) 1973. As per I.P. the initial part of raising the antibodies remains unaltered but the actual titration of diphtheria antitoxin from the immunised guinea pigs was performed by using the following in vitro methods: a) indirect haemagglutination test using human "O" red blood cells to coat diphtheria toxoid using chromic chloride as the coupling agent [3]; b) toxin neutralisation test using Vero cells [4]; c) a double diffusion technique in agar gel for titration of diphtheria antitoxin [5]. Our findings show clearly that the results of two in vivo methods i.e. Challenge Test, Alternative I.P. Method and the above-mentioned three in vitro methods are comparable and would certainly reduce the number of animals required by making a combination of in vivo and in vitro techniques to give us an assessment of the potency of the vaccine to be tested.