氯唑西林可视化免疫亲和凝胶柱检测方法研究  

Study on the Detection Method of Cloxacillin Visualized Immunoaffinity Gel Column

作　　者：张冬昊 赵桐桐 魏东 邢彦枝 何扩 ZHANG Dong-hao;ZHAO Tong-tong;WEI Dong;XING Yan-zhi;HE Kuo(Key Laboratory of Analysis and Testing of Agricultural Products Food Quality and Safety in Hebei Province,Hebei North University,Zhangjiakou 075000,China;Zhangjiakou Agricultural Product Quality Safety Supervision and Inspection Center,Zhangjiakou 075000,China)

机构地区：[1]河北北方学院河北省农产品食品质量安全分析检测重点实验室,河北张家口075000 [2]张家口市农产品质量安全监督检验中心,河北张家口075000

出　　处：《现代食品科技》2019年第4期273-278,共6页Modern Food Science and Technology

基　　金：河北省科技攻关重点研发项目(16236802D-5);河北省教育厅科技研发项目(QN206242);张家口科学技术与发展项目(1811036C);河北北方学院科学技术与发展项目(22036;12995583)

摘　　要：为了建立氯唑西林(Cloxacillin,CLOX)可视化免疫亲和凝胶柱检测方法。实验采用活化酯法,将辣根过氧化物酶(HRP)与氯唑西林偶联,制得氯唑西林酶标抗原,并通过紫外光谱扫描鉴定表明CLOX酶标抗原合成成功。实验采用恒温振荡法,将溴化氰活化的琼脂糖凝胶分别与氯唑西林抗体、HRP抗体偶联制得氯唑西林抗体胶和HRP抗体胶,作为亲和凝胶柱的检测层与质控层。建立氯唑西林可视化免疫亲和凝胶柱分析方法,确定了该方法在酶标抗原的稀释倍数为4000、抗体胶稀释倍数为10、HRP胶稀释倍数为20、样品稀释液pH值为5.7、加样时间为60 s条件下,达到最佳工作状态。该方法检测限为25μg/L,特异性强,与哌拉西林、阿莫西林、氨苄青霉素、四环素、呋喃它酮、甲砜霉素、链霉素均无交叉反应,动物源食品实际样品中氯唑西林检出限为25μg/kg。该方法便捷、快速、可视化,适合作为高通量筛选氯唑西林的有效检测手段。In order to establish a method for the detection of cloxacillin visualized immunoaffinity gel column,the activated ester method was used to combine horseradish peroxidase (HRP) with cloxacillin to obtain the cloxacillin enzyme standard antigen.Results of the UV spectrum scanning showed that the CLOX enzyme standard antigen was successfully synthesized.The agarose gel activated by hydrogen bromide was combined with cloxacillin antibody and HRP antibody,respectively,which was used as the detection layer and the quality control layer of the affinity gel column,respectively.A visual immunoaffinity gel column analysis method for cloxacillin was established.The optimum condition of this method was the dilution ratio of enzyme-labeled antigen of 4000,dilution ratio of antibody gel of 10,dilution ratio of HRP gel of 20,sample dilution pH of 5.7 and sample addition time of 60 s.The detection limit of this method was 25 μg/L.Its specificity was strong. There was no cross-reaction with piperacillin,amoxicillin,ampicillin,tetracycline,furantanone,thiamphenicol and streptomycin.The detection limit of cloxacillin in animal-derived foods was 25 μg/kg.This method is convenient,rapid and visual.It is suitable for high-throughput screening of cloxacillin.

关 键 词：氯唑西林(cloxacillin CLOX) 多克隆抗体 半抗原 检测方法 

分 类 号：O652[理学—分析化学] TS207.5[理学—化学]