食品科学 ›› 2012, Vol. 33 ›› Issue (8): 216-220.doi: 10.7506/spkx1002-6630-201208047

• 分析检测 • 上一篇    下一篇

环丙沙星时间分辨荧光免疫分析方法的建立与应用

李丽华1,白瑞樱2,孙远明1,沈玉栋1,徐振林1,雷红涛1,张 挺1,王 弘1,杨金易1,*   

  1. 1.广东省食品质量安全重点实验室,华南农业大学食品学院 2.新乡医学院基础医学院生理学与神经生物学教研室
  • 出版日期:2012-04-25 发布日期:2012-03-31
  • 基金资助:
    广东省科技厅2010 粤港招标项目(2010A020104004);广东省- 教育部产学研结合项目(2010A090200084; 2009B090300409);国家公益性行业(农业)科研专项(201003008-08);广东省科技计划项目(农业攻关2009A020101004)

Development and Application of Time-resolved Fluroimmunoassay (TR-FIA) for Ciprofloxacin Determination

LI Li-hua1,BAI Rui-ying2,SUN Yuan-ming1,SHEN Yu-dong1,XU Zhen-lin1,LEI Hong-tao1, ZHANG Ting1,WANG Hong1,YANG Jin-yi1,*   

  1. (1. Guangdong Provincial Key Laboratory of Food Quality and Safety, College of Food Science, South China Agricultural University, Guangzhou 510642, China;2. Department of Physiology and Neurobiology, School of Basic Medical Sciences, Xinxiang Medical University, Xinxiang 453000, China)
  • Online:2012-04-25 Published:2012-03-31

摘要: 建立基于时间分辨荧光免疫分析技术的环丙沙星(CPF)间接竞争检测方法。将环丙沙星-卵清白蛋白(CPF-OVA)包被于固相微孔板,CPF标准品或样品中CPF与CPF包被原(CPF-OVA)共同竞争限量的CPF抗体,用铕标羊抗兔IgG示踪,建立环丙沙星时间分辨荧光免疫分析方法(CPF-TRFIA);考察包被质量浓度、抗体稀释液、反应时间等对方法灵敏度的影响。结果表明:在优化条件下,该方法的检测限为0.5μg/L,半抑制质量浓度(IC50)为4.32μg/L,线性范围(IC20~IC80)为1.14~28.85μg/L;该方法对蜂蜜中CPF平均回收率为80.66%~100.30%,对牛奶中CPF的平均回收率为76.30%~95.22%,两者的平均批内和批间变异系数分别为7.42%和10.92%。CPF-TRFIA间接竞争法测定环丙沙星具有灵敏度高、特异性好、性能稳定的优点,适于高通量筛查。

关键词: 环丙沙星, 时间分辨荧光免疫分析, 抗体, 添加回收

Abstract: A time-resolved fluroimmunoassay (TRFIA) for the determination of ciprofloxacin (CPF) in an indirect competitive immunoassay mode was developed and applied. CPF-OVA was coated by physical adsorption onto a microtitre plate, and CPF standard or sample was competed with CPF-OVA for limited anti-CPF antibody, which was traced using europium-labeled goat anti rabbit IgG. The effects of experimental conditions including CPF-OVA concentration, buffer type, and reaction time on the sensitivity of TRFIA were explored. Under optimal conditions, the method revealed a detection limit of 0.5 μg/L, an IC50 of 4.32μg/L and a linear range (IC20-IC80) of 1.13-28.85 μg/L. The spike recovery rates of CPF in honey and milk at three spike levels were in the range of 80.66%-100.30% and 76.30%-95.22%, respectively, and the intra-batch and inter-batch coefficients of variation were 7.42% and 10.92%, respectively. The method was sensitive, specific, stable and suitable to be used for high-throughput determination of CPF residues.

Key words: ciprofloxacin, time-resolved fluroimmunoassay, antibody, recovery

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