氯丙嗪抗体制备与酶联免疫吸附测定方法的建立

本研究针对动物性食品中违禁药物氯丙嗪残留问题，建立了针对氯丙嗪的酶联免疫吸附测定方法。首次通过以氯丙嗪为原料，先去甲基化合成N-甲基氯丙嗪，再与丙烯酸叔丁酯反应及水解等步骤成功合成了氯丙嗪半抗原（CPZ-H）。半抗原CPZ-H分别与OVA和BSA通过活泼酯法偶联合成包被原与免疫原，采用免疫原CPZ-H-BSA免疫新西兰大白兔成功制备了特异性的抗氯丙嗪的多克隆抗体。基于该抗体建立了快速测定氯丙嗪的间接竞争酶联免疫吸附测定方法（ic-ELISA）。通过优化ic-ELISA方法的最佳工作条件，确定包被原和抗体稀释倍数均为1:8000时效果最好，此时标准曲线的IC50为1.1 ng/mL，线性范围为：0.13 ng/mL~8.8 ng/mL。制备的抗氯丙嗪的多克隆抗体与其它氯丙嗪类似物无交叉反应，特异性好。本研究为今后高特异性氯丙嗪快速检测试剂盒的制备提供依据。

Preparation of Conjugate Antigen and Antibody to Chlorpromazine in the Animal Food

In this study, an enzyme-linked immunosorbent assay (ELISA) for the determination of chlorpromazine residues in animal foods was established. Chlorpromazine hapten (CPZ-H) was synthesized from chlorpromazine by demethylation, reaction with tert-butyl acrylate and hydrolysis for the first time. Hapten CPZ-H was conjugated with OVA and BSA by active ester method to form the coated antigen and immunogen, respectively. New Zealand white rabbits were immunized with immunogen CPZ-H-BSA to successfully prepare specific polyclonal antibodies against chlorpromazine. An indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) for the rapid determination of chlorpromazine was established based on the antibody. By optimizing the working conditions of ic-ELISA method, it was determined that the dilution fold of coated antigen and antibody was 1:8000. At this time, the IC50 of standard curve was 1.1 ng/mL and the linear range was 0.13 ng/mL~8.8 ng/mL. The prepared polyclonal antibody against chlorpromazine has good specificity and no cross reaction with other chlorpromazine analogues. This study provides a basis for the preparation of specific and rapid detection kit for chlorpromazine in the future.