A Competitive ELISA for the potency assay of vaccines and antitoxin sera against the epsilon toxin of Clostridium perfringens type D

PARREIRA, M.P., AURÉLIO, R., CAMPOS, P.C., JÚNIOR, A.D.C., LOBATO, F.C.F., HENEINE, L.G.D.

Immunology Laboratory, Fundação Ezequiel Dias, Belo Horizonte, MG, Brasil.

Competitive ELISA (cELISA) was standardized to detect antibodies to Clostridium perfringens Type D epsilon prototoxin as an alternative to the toxin neutralization test performed in mice. Epsilon prototoxin was purified by ion-exchange chromatography on DEAE-Sepharose column and detoxified by controlled iodination and used to coat microwell plates. Sera were tested for their ability to inhibit the standard ELISA reaction between rabbit anti-epsilon immunoglobulin and the epsilon prototoxin. The potency of the test serum was calculated from a standard curve produced with the International Standard anti-epsilon serum obtained from the National Institute for Biological Standards and Control (NIBSC). The applicabitility and repetibility of the test was measured by calculating the in vitro potency of sheep anti-epsilon toxin sera, in different days using standard curves prepared in the same day and previously elaborated. The potencies obtained were averaged and compared to the potency obtained with the neutralization test in mice. The correlation observed was equal to r=0.94. Cattle sera vaccinated with clostridial vaccines against C. perfringens type C and D, were assayed for antitoxin potency in mice and with the cELISA. The correlation was r= 0.87.The results indicate that the cELISA can reliably quantify the potency of anti-epsilon toxin sera and replace the neutralization test in mice.

Acknowledgements: FAPEMIG, CNPq.

CORRESPONDENCE TO:

HENEINE, L.G.D., Immunology Laboratory, Fundação Ezequiel Dias, Rua Conde Pereira Carneiro, 80, Gameleira, Belo Horizonte, MG, Brasil, Email: heneinel@funed.mg.gov.br