人肠道病毒71型中和抗体检测方法的实验室评价

Laboratory Evaluation of Method for Determination of Neutralizing Antibody against Human Enterovirus 71

目的对人肠道病毒71型(Enterovirus71,EV71)中和抗体检测方法进行实验室评价,为该方法的标准化提供依据。方法 5个实验室按照统一制定的EV71中和抗体检测操作细则(SOP),应用A、B和C基因型的10株EV71病毒株,分别测定10份人免疫球蛋白、20份健康成人血浆和15份EV71动物高效价免疫血清样品中EV71中和效价,检测实验室内、实验室间重复性及不同EV71病毒株对中和抗体检测的影响。结果相同实验室应用各病毒株重复测定中和效价的最大值与最小值(Max-Min)倍数差均在4倍以内,3次重复试验结果间差异无统计学意义(P值均>0.05);不同实验室应用相同病毒株检测结果的Max-Min倍数差在4倍以内;不同实验室应用不同病毒株检测结果为:A型株与其他基因型病毒株中和效价的Max-Min差在192倍以内,B3与C4亚型的病毒株中和效价的Max-Min倍数差在64倍以内,C4亚型不同病毒株之间中和效价的Max-Min倍数差在16倍以内。结论按统一SOP操作后,EV71中和抗体检测方法应用相同毒株在实验室内和实验室间具有较好的重复性,而不同病毒株对中和效价的测定结果有较大影响。

Objective To evaluate the method for determination of neutralizing antibody against human enterovirus 71 （EV71）in laboratory and provide a basis for standardization of the method. Methods The neutralizing antibody titers against EV71 in 10 human immunoglobulin, 20 healthy adult plasma and 15 high titer animal immune serum specimens were determined with 10 EV71 strains of genotypes A, B and C by 5 laboratories according to the SOP for EV71 neutralizing antibody, based on which the intra-and inter-reproducibility of the method as well as effect of various EV71 strains on determination of neutralizing antibody were analyzed. Results The fold differences in maximum and minimum（Max-Min）of determination results of neutralizing antibody titers with various EV71 strains by the same laboratory were less than 4, and the results of 3 repeat tests showed no significant difference （P 0. 05）. The fold differences in determination results with the same EV71 strain by various laboratories were also less than 4. The determination results with various EV71 strains by various laboratories were as follows： the fold differences of the Max-Min of neutralizing antibody titers of EV71 strain genotype A with other genotypes were not more than 192, while those of subtypes B3 with C4 were not more than 64, and those of subtype C4 with other strains were not more than 16. Conclusion By operation according to the unitary SOP, the method for determination of neutralizing antibody against EV71 showed satisfactory intra- and inter-reproducibility. However, various EV71 strains showed significant effect on the determination result.