Hapten design, monoclonal antibody preparation and immunoassay development for direct detection of the toxic metabolite of nitrovin without derivatization from fish

Abstract

Nitrovin, classified as a nitrofuran drug, has been banned in numerous countries due to its carcinogenic effect on human health. However, existing analytical techniques for its detection are laborious, requiring an overnight derivatization process lasting 16 h. In this study, novel haptens were designed to prepare antibodies against aminogunidine (AGD), the metabolite of nitrovin, and then developed an indirect competitive enzyme-linked immunosorbent assay (icELISA) method to detect AGD from fish samples without derivatization for the first time. Computer-assisted molecular modeling predicted the efficiency of novel haptens by conformational studies and electronic analysis. The results were consistent with the predicted results in producing high-affinity monoclonal antibody (mAb) against AGD. The half-inhibition concentration (IC50) value of icELISA was 0.80 ng mL−1 for AGD, with a limit of detection of 0.27 μg kg−1 and recoveries of 89.8%–97.2%. The results showed that this new hapten design and synthesis provided a great prospect to obtain high-affinity mAb for AGD. This developed icELISA substantially avoided the conventional and lengthy derivatization process and was an effective method for the direct detection of AGD in fish tissue without derivatization.