›› 2013, Vol. 40 ›› Issue (3): 242-246.

• 经验交流 • 上一篇    下一篇

动物源性食品中齐帕特罗一步法ELISA试剂盒的研制

董李学1, 葛怀礼1, 郑百芹1, 李爱军1, 张建民1, 蒙君丽1, 苗建民1, 李亮2   

  1. 1. 唐山市畜牧水产品质量监测中心,河北唐山 063001;
    2. 杭州迪恩科技有限公司,浙江杭州 310023
  • 收稿日期:2013-01-28 出版日期:2013-03-20 发布日期:2013-03-19
  • 通讯作者: 葛怀礼(1963-),男,河北人,高级工程师,从事畜牧水产品质量检测工作。E-mail:13831531216@139.com E-mail:13831531216@139.com
  • 作者简介:董李学(1984-),男,河北人,畜牧师,从事畜牧水产品质量检测工作。
  • 基金资助:
    生猪及健康肉安全快速检测关键技术研究与应用(11120206A-1)。

Study on One-step Method ELISA Kit for Detection of Zilpaterol in Animal Derived Food

DONG Li-xue1, GE Huai-li1, ZHENG Bai-qin1, LI Ai-jun1, ZHANG Jian-min1, MENG Jun-li1, MIAO Jian-min1, LI-Liang2   

  1. 1. Tangshan Livestock and Aquatic Products Quality Monitoring Center Laboratory, Tangshan 063001, China;
    2. Hangzhou DNA Sci-Tech Co., Ltd., Hangzhou 310023, China
  • Received:2013-01-28 Online:2013-03-20 Published:2013-03-19

摘要: 本研究依据直接竞争ELISA原理建立了齐帕特罗一步法ELISA试剂盒。以齐帕特罗与载体蛋白偶联物作为免疫原免疫新西兰大白兔制得齐帕特罗多克隆抗体,棋盘包被法确定其最佳抗体包被浓度、酶标抗原工作浓度、包被条件、反应时间、底物显色时间等,并对试剂盒的各项技术指标进行确认。结果表明,成功组装了齐帕特罗一步法ELISA试剂盒,并建立了尿液、饲料、奶粉和奶汁,以及组织等样品的前处理方法,检测限均远低于1 μg/kg。该试剂盒线性检测范围为0.15~10 ng/mL,IC50浮动范围0.43~0.79 ng/mL,样品板内、批内、批间的变异系数均小于15%,平均回收率在70%~110%之间,与其同类药物的交叉反应率均小于0.1%。提示,本试验研制的试剂盒重复性、特异性、稳定性等各项指标均符合技术要求,可用于动物源性食品中齐帕特罗药物残留的检测。

关键词: 动物源性食品; ELISA; 齐帕特罗

Abstract: This study established a competitive enzyme-linked immunosorbent assay (ELISA) method for detecting zilpaterol in animal derived food. Zilpaterol polyclonal antibody was obtained by immunizing New Zealand rabbit with zilpaterol-KLH conjugates. Checkerboard titration method was used to determine the optimum concentration of antibody and enzyme-labelled antigen. Coating condition, reaction time and the color developing time of substrate were also detected. The results showed that an ELISA kit with a linear sensitivity ranged from 0.15~10 ng/mL was successfully established. Half inhibitory concentration (IC50) ranged between 0.43~0.79 ng/mL. The recovery rates for spiked zilpaterol in urine, feed, milk powder, milk and tissue were 65%~90% , with less than 15% coefficient variations (CV). The cross-reaction for other similar medicines was less than 0.1%. The ELISA kit established in current study for detecting zilpaterol in animal derived food and showed the good reproducibility, stability and specificity, hence it could be used to detect the zilpaterol residues in animal derived food.

Key words: animal derived food; enzyme-linked immunosorbent assay; zilpaterol

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