RESUMO
The structure and protective activity of tetanus antibodies elicited in rabbits after whole-cell pertussis diphtheria-tetanus vaccine (DTPw) vaccination was studied. ELISA antibody levels and toxin neutralisation activity (TNT) were measured in individual serum samples. The ratio of symmetric and asymmetric (functionally monovalent) IgG molecules was determined by concanavalin A (Con A) chromatography. This test is based on the fact that the carbohydrate group responsible for the molecular asymmetry has high affinity for the lectin Con A. Asymmetric molecule ratio was observed to increase with immunisation time, as well as differences between TNT and ELISA levels. All serum samples were overestimated by ELISA as compared to TNT assay, in line with the markedly higher proportion of asymmetric molecules which have lower toxin neutralising activity. Protective levels could not be predicted reasonably from ELISA results below 0. 222 IU/ml, because this methodology fails to discriminate between both types of antibodies and only an in vivo serum neutralisation procedure (TNT) reflects the true neutralising serum activity.
Assuntos
Anticorpos Antibacterianos/análise , Toxina Tetânica/imunologia , Animais , Vacina contra Difteria, Tétano e Coqueluche/imunologia , Ensaio de Imunoadsorção Enzimática , Fragmentos Fab das Imunoglobulinas/imunologia , Testes de Neutralização , CoelhosRESUMO
Symmetric and asymmetric IgGs having different neutralizing capacity are synthesized in variable proportions by the same clones during the course of immune response. The neutralizing activity of tetanus antibodies was studied in rabbits vaccinated with acellular (DTPa) or whole-cell pertussis (DTPw) vaccines. Symmetric and asymmetric F(ab)'2 fragments from the IgG fraction of the peak serum pools from each group of rabbits were purified by concanavalin A chromatography and measured by ELISA. After the third vaccine dose the asymmetric antibody percentage for DTPw (40%) was twice that for DTPa (20%). The neutralizing activity of asymmetric antibodies was roughly sixfold lower than symmetric ones. When antibody values titrated by ELISA approach minimal protective level, the proportion of symmetric antibodies with high toxin neutralizing activity acquires crucial importance.