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·mL－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·mL－1，检测限为7．6 pg·mL－1，检测线性范围为10—810 pg·mL－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 was 7.6 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.
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