高灵敏黄曲霉毒素B1酶联免疫试剂盒的研制及应用  

Development and application of a highly sensitive ELISA kit for aflatoxin B1

作　　者：孙清[1,2] 李谷丰 邓乾民 刘杰民[1,2] 时国庆[2] 

机构地区：[1]北京科技大学土木与环境工程学院,北京100083 [2]北京科技大学化学与生物工程学院,北京100083 [3]北京普赞生物技术有限公司,北京100085

出　　处：《环境化学》2015年第10期1845-1853,共9页Environmental Chemistry

基　　金：国家高技术研究发展计划(2012AA101609-5);新世纪优秀人才支持计划(NCET-11-0581);中央高校基本科研业务费专项资金(FRFTP-12-007B)资助

摘　　要：为实现对环境及食品样品中黄曲霉毒素B1的高灵敏检测,通过优化一系列试剂盒参数,研制了一步间接竞争ELISA检测试剂盒.优化后的包被缓冲液为90 mmol·L-1、pH 4.6的柠檬酸缓冲液,最佳反应pH值为7.4,抗体包被浓度为0.2μg·m L-1,HRP-BSA-AFB1稀释比为1/4000,标品稀释液为含7%甲醇的PBST溶液.优化后试剂盒IC50值为66 pg·m L-1,检测限为7.6 pg·m L-1,检测线性范围为10—810 pg·m L-1.试剂盒对不同AFB1添加水平(0.5μg·kg-1,1μg·kg-1)的玉米、豆粕和鱼粉样品平均回收率为108.4%—134.8%.对玉米、豆粕和鱼粉样品各20份盲样测试结果表明,试剂盒检测结果与HPLC-MS/MS检测结果吻合.A one-step indirect competitive ELISA kit for AFB1 was successfully developed,for the detection of trace AFB1 in environment and foodstuffs.Optimization was perfonmed for coating buffer,pH of assay buffer,concentrations of anti-AFB1 monoclonal antibody and HRP labeled AFB1-BSA,and sample dilution reagent. The limit of detection and the IC50 for AFB1 was7. 6 pg·mL^-1 and 66 pg·mL^-1 respectively,with good linearity between 10—810 pg·mL^-1 for AFB1. Spike recoveries for maize,soybean meal and fishmeal samples ranged from 108. 4% to 134.8%. Aflatoxin B1 residues in sixty samples were determined by the ELISA kit and high performance liquid chromatography-tandem mass spectrometry（ HPLC-MS / MS） respectively,and the result of ELISA kit was consistent with that of HPLC-MS / MS.

关 键 词：ELISA 黄曲霉毒素B1 残留 试剂盒 

分 类 号：X85[环境科学与工程—环境工程]