摘要 以合成沙丁胺醇(SAL)完全抗原为基础获得针对SAL的多克隆抗体,建立检测SAL的间接竞争ELISA(ic-ELISA)检测方法,为进一步开发SAL快速检测试剂盒奠定基础。为此,分别用混合酸酐法和活化酯法制备SAL-BSA免疫原和SAL-OVA检测原,免疫新西兰大白兔获得SAL多抗血清,建立检测SAL ic-ELISA检测方法。结果显示,成功合成了SAL-BSA和SAL-OVA,其偶联比分别为17∶1和6.4∶1;免疫后兔血清抗体效价为1∶102400,所建立ic-ELISA的半数抑制浓度(IC50)为24.84μg/L,最低检测限(IC10)为0.78μg/L,线性范围IC20~IC80为3.16μg/L~80.1μg/L;精密度(CV)%<10%,回收率在98%~110%之间。特异性鉴定结果显示,SAL多抗血清除与侧链上含有叔丁基官能团的结构类似物具有较强的交叉反应外,不存在与其他同类分子的交叉反应。以上结果表明,成功建立了检测SAL的ic-ELISA检测方法,为进一步开发SAL快速检测试剂盒奠定了基础。 A polyclonal antibody against SAL was obtained based on the complete antigen of salbutamol(SAL),and an indirect competitive ELISA(ic-ELISA)method for detecting SAL was established to lay the foundation for further development of a rapid detection kit for SAL.The SAL-BSA immunogen and the SAL-OVA detector were prepared by the mixed acid anhydride method and the activated ester method,respectively.New Zealand white rabbits were immunized to obtain the SAL polyclonal antiserum,and a detection method for SAL ic-ELISA was established.The results showed that the SAL-BSA immunogen and the SAL-OVA coated progeny were successfully synthesized,and their coupling ratios were 17∶1 and 6.4∶1 respectively;the titer of rabbit serum antibody after immunization was 1∶102400,and the established ic-ELISA half-inhibition concentration(IC50)is 24.84μg/L,the minimum detection limit(IC10)is 0.78μg/L,and the linear range IC20-IC80 is 3.16μg/L-80.1μg/L;precision(CV)%<10%,recovery rate is between 98%-110%.Specific identification results showed that SAL polyclonal antibody had a strong cross-reaction with structural analogs containing tertbutyl functional groups on the side chain,and there was no cross-reaction with other similar molecules.The above results indicated that the ic-ELISA method for detecting SAL was successfully established,laying a foundation for the further development of a rapid detection kit for SAL.
机构地区 新乡学院生物技术研究中心 郑州大学
出处 《动物医学进展》 北大核心 2021年第5期42-49,共8页 Progress In Veterinary Medicine
基金 2018年国家重点研发计划项目(2018YFC1602902) 河南省科技攻关项目(192102110066) 河南省高等学校重点科研项目(19A230009) 新乡学院博士科研启动经费(1366020120)。
关键词 沙丁胺醇 人工抗原 多克隆抗体 间接竞争ELISA salbutamol artificial antigen polyclonal antibody indirect competition ELISA
