An antigen based on molecular modeling resulted in the development of a monoclonal antibody-based immunoassay for the coccidiostat nicarbazin

Based on insights obtained from computer-assisted molecular modeling, p-nitrosuccinanilic acid was selected as a viable candidate to use as a hapten to produce antibodies to 4,4´dinitrocarbanilide, the active component of the coccidiostat nicarbazin. These antibodies would be used to quantify nicarbazin in feeds and tissues. The synthesis of p-nitrosuccinanilic acid is presented here. Due to the lack of water solubility of nicarbazin, a discussion is presented concerning the solubility of nicarbazin as it pertains to the immunoassay. DMF and ACN were both added to the assay buffer to make standard solutions of nicarbazin, and facilitate the use of the competition enzyme-linked immunosorbent assays (ELISAs). Monoclonal antibodies were isolated that compete with nicarbazin, and they were isotyped as IgM. However, this study has been complicated due to problems that developed during the study associated with the competition ELISA