氟西汀快速免疫检测用人工抗原的合成与鉴定

Synthesis and Identification of Artificial Antigen for the Rapid Immunological Detection of Fluoxetine

本文针对药物、减肥类保健食品中氟西汀成分及残留问题现状,主要研究了氟西汀人工抗原的合成方法,并对所合成的半抗原、人工抗原进行了结构和特异性鉴定。研究中先将氟西汀与丙烯酸甲酯反应后再经过水解制备出半抗原(H-FXT),再用活泼酯法将半抗原分别与牛血清白蛋白(BSA)和卵清蛋白(OVA)偶联制备了氟西汀人工免疫原(H-FXT-BSA)和氟西汀包被原(H-FXT-OVA),通过免疫原免疫Balb/c小鼠,利用间接ELISA方法对抗血清进行了鉴定评价。研究制备的人工抗原H-FXT-BSA、H-FXT-OVA经紫外光谱扫描计算后偶联比分别达到14.5:1和11.3:1。免疫小鼠后所获得抗血清的效价均达到30000以上,半抑制量浓度IC50值为100 ng/m L。本研究首次合成了鉴定氟西汀人工抗原,并成功获得了氟西汀抗体,本研究可为后期在药物、减肥类保健食品中氟西汀成分的免疫快速检测方法的建立和酶联免疫检测试剂盒等相关产品开发奠定了关键技术研究基础。

Currently,fluoxetine residues in drugs and slimming health foods pose a health problem.Therefore,the synthesis of artificial antigens for fluoxetine was explored,and the structures and specificities of the synthesized hapten and artificial antigen were identified.Fluoxetine reacted with methyl acrylate and underwent hydrolysis to produce the hapten（H-FXT）.Subsequently,the active ester method was used to conjugate H-FXT with bovine serum albumin（BSA） and ovalbumin（OVA） separately,to obtain the fluoxetine immunogen（H-FXT–BSA） and coated fluoxetine antigen（H-FXT-OVA）,respectively.The H-FXT-BSA immunogen was used to immunize Balb/c mice and the generated anti-serum was analyzed by indirect enzyme-linked immunosorbent assay（ELISA）.The artificial antigens,H-FXT-BSA and H-FXT-OVA,were analyzed by ultraviolet（UV） spectroscopy,and calculations showed that the coupling ratios were 14.5：1 and 11.3：1,respectively.The antiserum titers after the immunization of Balb/c mice were all above 30000,and the half maximal inhibitory concentration was approximately 100 ng/m L.The artificial fluoxetine antigens were successfully synthesized for the first time and anti-fluoxetine antibodies were obtained.This study can provide a key technical basis for further studies involving the establishment of a method for the rapid immunological detection of fluoxetine,including the development of an ELISA kit and other related products.