Hapten design to prepare monoclonal antibodies and establishment of immunoassay for direct screening of oxamichydrazide in chicken, the metabolite of nifuraldezone
Ghulam Mujtaba Mari, Weilin Wu, Aisha Talpur, Jan Muhammad Shah, Saqib Ali, FAIZ MUHAMAD KHAND, E. Bughio, Abdul Samad, Zhanhui Wang, Sergei A. Eremin
Abstract
Abstract In this study, two novel haptens of oxamichydrazide (OXZ), the metabolite of nifuraldezone, were designed and synthesized by derivatization with ethyl-4-bromobutanoate and ethyl-2-bromoacetate for the first time. The rationality of the haptens was enlightened by conformational alignment and electronic evaluations based on computional chemistry. The synthesized haptens were efficient in generating antibody response to OXZ. A monoclonal antibody (mAb) named 2D9 with high affinity was obtained and used to establish an indirect competitive enzyme-linked immunosorbent assay (icELISA) for the direct determination of OXZ in chicken samples without the need for derivatization step. Under optimum conditions, the developed icELISA showed an IC 50 value of 0.66 ng mL −1 , and limit of detection of 0.11 µg kg −1 in chicken. The average recoveries were found to be 80.4–91.3% with a coefficient of variation less than 9.0%. In conclusion, we have successfully desigened haptens, prepared mAbs and developed an immunoassay for the direct determination of OXZ in chicken samples without the need for a derivatization step for the first time. This provides a potentially rapid, accurate, and sensitive screening tool for nifuraldezone residue in animal-derived food.