TTMV间接ELISA检测方法的建立

DEVELOPMENT OF AN INDIRECT ELISA FOR DIAGNOSIS OF TORQUE TENO MINI VIRUS

以TTMV重组蛋白作为包被抗原,通过反应条件优化,建立了检测人TTMV血清抗体的间接ELISA方法,结果显示:抗原最佳包被浓度为1.63μg/mL,37℃孵育1 h后4℃过夜;血清稀释度为1:320;酶标二抗孵育时间为37℃60 min;ELISA酶标板的临界值为0.211。运用该方法对上海市和辽宁省共计280份血清样品进行检测,阳性检出率分别为34.68%和39.74%。检测结果表明,建立的间接ELISA方法特异性与重复性均较好,为进一步完善TTMV临床诊断方法打下了基础。

With recombinant protein of TTMV as the coating antigen, an indirect ELISA for detecting human antibodies against TTMV was established with optimized reaction conditions. The optimum reaction conditions of ELISA were as follows： concentration of coating antigen was 1.63 μg/mL, coating was performed at 37 ℃ for 1 h and left at 4 ℃ over night; the serum samples were diluted to 1：320, and the enzyme labeled secondary antibody was incubated at 37℃ for 60 min. The critical OD value for the ELISA plate was 0. 211. Two hundred eighty serum samples from Shanghai city and Liaoning Province were tested by the method, and the positive rate of TTMV antibodies were 34.68%（Shanghai） and 39.74%（Liaoning）, respectively. The detection results demonstrated that the indirect ELISA method had high specificity and repeatability, which laid a foundation for diagnosis of clinical TTMV infections.