基于微量中和试验的新冠感染者血清样本中和抗体高通量检测方法评价

Evaluation of high-throughput methods for the detection of neutralizing antibodies in serum samples from individuals infected with 2019-nCoV based on a microneutralization test

目的基于微量中和试验为金标准,评价两类三种高通量中和抗体检测方法的检测能力。方法以江苏省2020年以来发现的64例新冠病例为研究对象,采集早期和随访血清样本共117份开展中和抗体水平分析,用微量中和试验、假病毒中和抗体试验(PBNA)、竞争抑制试验[酶联免疫法(ELISA)和化学发光免疫法(CLIA)]别对血清样本进行测试,评价这些高通量中和抗体检测方法的相关系数及有效性的临界值等。结果新冠感染者假病毒中和抗体试验、酶联免疫法、化学发光免疫法相对于微量中和试验检测的相关系数分别为0.760、0.778和0.725,当微量中和试验的临界值取为1:10时(小于10为阴性),PBNA检测方法的Cutoff值为50时,ROC曲线下面积为0.901;ELISA检测方法的Cutoff值为1∶8时,ROC曲线下面积为0.934;CLIA检测方法的Cutoff值为1.28AU/ml时,ROC曲线下面积为0.838。结论建立的新冠病毒中和抗体高通量检测方法具有科学性和可行性,为新冠疫情常态化防控提供重要的技术手段。

Objective To evaluate the detection ability of two kinds of high-throughput method to determine neutralizing antibodies based on a microneutralization test(MNT).Methods Serum samples were collected from the early phase and follow-up period(117 samples in total)for neutralizing antibody testing.They were tested using MNT,pseudovirus neutralization assay(PBNA),competitive inhibition assay(including enzyme-linked immunosorbent assay(ELISA)and chemiluminescence immunoassay(CLIA))to evaluate the correlation coefficients and threshold values for the effectiveness of these high-throughput neutralizing antibody assays.Results The correlation coefficients for PBNA,ELISA,and CLIA relative to MNT were 0.760,0.778,and 0.725,respectively,for individuals infected with 2019-nCoV.The area under the ROC curve was 0.901 for a cutoff value of 50 for the PBNA assay,0.934 for a cutoff value of 1∶8 for the ELISA assay and 0.838 for a cutoff value of 1.28AU/ml for the CLIA assay when the threshold value for the microneutralization test was taken as 1:10(less than 1:10 is considered negative).Conclusions The high-throughput method for the detection of COVID-19 neutralizing antibodies are scientific and feasible,and provide an important technical tool for the regular prevention and control of the epidemic.