摘要
目的:制备去甲雄三烯醇酮(又称群勃龙,trenbolone,TR)抗体以建立TR特异性检测方法。方法:TR与琥珀酸酐反应生成TR-半琥珀酸酯(TR-HS),采用混合酸酐法和对乙基-N,N-二甲基丙基碳二亚胺(EDC)法将TR-HS与牛血清蛋白(BSA)偶联制备其偶联物;质谱、核磁共振分析偶联结果;免疫家兔并用双向免疫扩散和ELISA检测抗血清效价,判断偶联物的免疫原性。结果:经质谱鉴定显示,所获得的修饰物TR-HS相对分子质量为370;核磁共振分析表明,各吸收峰与理论相符;基质辅助激光解析电离飞行时间质谱图谱分析表明,TR-BSA相对分子质量明显增加、紫外最大吸收峰漂移;TR与BSA混合酸酐法和EDC法偶联比分别为1∶18和1∶12;混合酸酐法偶联物免疫家兔获得的抗血清效价ELISA测得TR特异性抗体效价为1∶320 000;双向免疫扩散测得的抗血清效价为1∶16。结论:成功制备了具有明显免疫原性的TR-BSA偶联物,为TR单克隆抗体的制备和建立TR残留量的快速检测方法奠定了基础。
Objective： To prepare trenbolone（TR） complete antigen and identify the antigenicity of TR.Methods： TR-half-succinate（TR-HS） was produced by TR reacting with succinic anhydride,the mixed anhydride and the [3-mercaptopropionic acid,1-ethyl-3-（3-diethylaminopropyl） carbodiimide,EDC] methods were used to couple TR-HS and BSA.The antigenicity analysis of TR-BSA complex was finished by detecting the titre of anti-serum separated from rabbits which was immunized with TR-BSA.Results： The TR-HS molecular weight was determined as 370 by analysis of mass spectrometry,the absorption peak was in accordance with theoretical results by nuclear magnetic resonance.MALDI-TOF-MS analysis showed that the molecular weight map of TR-BSA a marked increase and the UV maximum absorption peak drift;the coupling ratios of mixed anhydride and EDC method were 1∶18 and 1∶12,respectively.The results of ELISA and double-immunodiffusion test showed that the titre of anti-serum to TR was 1∶320 000,1∶16,respectively.Conclusion： R-BSA coupling was successfully prepared,which established the foundation for the anti-TR monoclonal antibody preparation as well as the method construction of TR Residues rapid test.
出处
《江苏大学学报（医学版）》
CAS
2010年第3期234-238,共5页
Journal of Jiangsu University：Medicine Edition
基金
江苏出入境检验检疫局资助项目(2007KJ32)
