Drug-protein conjugates--IX. Immunogenicity of captopril-protein conjugates

The immunogenicity of captopril (CP), conjugated to heterologous proteins, was investigated in male New Zealand White rabbits by monthly injections of CP-protein conjugates in Freund's Complete Adjuvant. Anti-CP antibody activity was readily detected by immunodiffusion in sera of rabbits immunized with the amide-linked CP-HSA (23:1) conjugate. Hapten inhibition studies revealed that the antigenic determinant contained CP and a lysine residue from the protein carrier. When rabbits were immunized with disulphide-linked CP-S-S-HSA (9:1) and CP-S-S-KLH (160:1) conjugates, anti-CP antibody activity was detected by a sensitive ELISA method, but not by immunodiffusion and radioligand binding assays. The specificity of the serum IgG anti-CP activity after immunization with disulphide-linked CP-S-S-protein conjugates was confirmed since anti-CP activity was inhibited by preincubation of the antisera with CP conjugated to an unrelated protein carrier (CP-S-S-OVA), but not by the corresponding unconjugated protein, nor by penicillamine-S-S-protein conjugates. These results show that disulphide-linked CP-protein conjugates are sufficiently stable to induce humoral (B lymphocyte) anti-hapten responses under the experimental conditions employed. In a separate study, delayed-type skin hypersensitivity reactions to topically applied CP were demonstrated in the guinea pig. The specific and sensitive immunochemical technique (ELISA) described here could be useful in future studies for determining whether or not patients taking CP produce antibodies to the drug.