Comparison of coating and immunizing antigen structure on the sensitivity and specificity of immunoassays for benzoylphenyl urea insecticides

Abstract

Several sensitive enzyme-linked immunosorbent assays (ELISAs) were developed for the benzoylphenylurea insect growth regulators diflubenzuron, BAY SIR 8514, and penfluron. The sensitivities and specificities of the assays have been investigated by using several different benzoylphenylurea derivatives as the immunogens as well as coating antigens. The following approach was used to solve the problem of bridge recognition: (a) homologous site systems, to use the same coupling site on the benzoylphenylurea molecule in the immunogen and coating antigen but to vary the bridge structure, and (b) heterologous site systems, coupling the immunogen and coating antigen from different positions on the benzoylphenylurea molecule. The heterologous systems yielded more sensitive assays, but antisera obtained from all the immunogens were used successfully in developing ELISA's for benzoylphenylureas. With these sensitive ELISA's, diflubenzuron was detected in milk at a level of 2 ppb without any sample extraction procedure, and sample cleanup led to still more sensitive assays. The reproducibility and sensitivity of the method are adequate for practical residue analysis. This work illustrates that a limited collection of antisera and coating antigens can yield a powerful library of assays capable of specifically detecting single compounds or a variety of members of a compound class.