产肠毒素大肠杆菌F41菌毛抗体间接ELISA检测方法的建立

Development of an indirect-ELISA for detecting specific antibody to F41 pilus protein of enterotoxigenic Escherichia coli

以纯化的重组F41菌毛蛋白作为检测抗原,建立了检测产肠毒素大肠杆菌F41菌毛抗体的间接ELISA方法。经优化筛选的最佳反应条件:包被抗原浓度为0.26μg/孔,血清稀释度为1∶800,酶标二抗工作浓度为1∶6 000,30 g/L明胶封闭1 h,底物作用时间为10 min。经试验验证,该方法具有特异性好、敏感性高、重复性好、稳定性强等特点。用建立的间接ELISA方法与PCR方法进行符合性试验,总符合率为97.4%。表明,该方法可以用于产肠毒素大肠杆菌F41菌毛抗体的检测。

An indirect enzyme-linked imrnunosorbent assay（ELISA） was developed based on a purified recombinant F41 pill protein of enterotoxigenic Escherichia coli （ETEC）. The optimum test conditions were as follows：the concentration of coating antigen was 0. 26 μg per well, the dilution of serum and HRP- labeled mice anti-bovine IgG were 1 ： 800 and 1 ： 6 000,respectively,30 g/L gelatin blocked for 1 h,and the reaction time of coloration was 10 rain. The specificity, sensitivity, reproducibility and stability were evaluated. Compared with antigen from PCR,97.4% agreement for positive and negative samples were ob rained. The results indicated that the indirect ELISA could be used for detecting anti-F41 pill antibodies of ETEC.