无色孔雀石绿人工抗原的制备及免疫原性鉴定

Synthesis and immunogenicity identification of artificial antigen of leucomalachite green

目的用免疫分析法检测水产品中无色孔雀石绿(LMG)的残留,研究LMG人工抗原及其多克隆抗体的制备和鉴定。方法通过混酸硝化还原法对LMG进行改造,获得氨基无色孔雀石绿(NH2-LMG),然后采用改良的碳二亚胺法制备LMG完全抗原。用该人工抗原免疫动物,获得多抗血清。采用酶联免疫法测定抗体效价。选择与LMG结构相似的5种药物进行交叉反应,计算交叉反应率。结果经紫外光谱及电泳法检测,LMG与载体蛋白偶联成功,偶联比为1∶17。利用该抗原免疫小鼠后获得了效价高于104的抗LMG抗体,交叉反应结果表明该抗体有较好的特异性。结论成功合成了LMG人工抗原,为进一步制备LMG单克隆抗体及研究LMG的新型、快速检测方法奠定了基础。

[Objective]To detect the residues of leucomalachite green（LMG）in aquatic products by immunoassay,and study the synthesis and identification of LMG artificial antigen and its polyclonal antibody.[Methods]The LMG was transformed by mixed acid nitrification reduction method to obtain amino LMG（NH2-LMG）,and then LMG complete antigens were made by improved carbodiimide method. The antiserum was obtained by immunization with this artificial antigen on animals. The valence of antibody was evaluated by ELISA. 5 drugs which construction was similar to LMG were selected to cross-reaction, and the cross-reaction rates were c alculated.[Results] The synthesis of LMG and carrier protein was su cceed through detection of ultraviolet spectrum and electrophoresis, and the coupling ratio was 1∶17. The LMG antibody which valence was more than 104 was obtained by immunization with this antigen on rats. The results of cross-reaction showed that the antibody had better specificity.[Conclusion]The artificial antigen of LMG is compounded successfully,which establishes the foundation for further producing LMG monoclonal antibody and studying a new and rapid detection method of LMG.