抗-HCV多表位抗体在检测HCV抗原中的应用

目的应用丙型肝炎病毒多表位抗体,探索从血浆或血清样品中检测丙型肝炎病毒抗原(HCAg)的可能性,为献血员筛选及临床早期诊断提供检测方法。方法利用原核系统表达的丙型肝炎病毒(HCV)7个高度保守区基因的多表位复合抗原免疫动物,制备抗-HCV多克隆抗体,采用ELISA双抗体夹心法检测血浆或血清中的HCAg,并与RT-PCR法进行比较。结果制备的抗-HCV多表位复合抗原多克隆抗体,在用ELISA法检测HCAg中表现出较高的特异性及灵敏性,Cutoff值为0·239,批内CV值为5·60%~6·65%,批间CV值为7·80%。与RT-PCR比较,相关系数为0·816,灵敏度为88·89%,特异度为96·30%,一致性为95·24%,调整一致性为90·82%,阳性预测值为80·00%,阴性预测值为98·11%。HCAg检出率与美国ORTHO公司HCV-C抗原检测试剂盒差异无显著意义(P=0·06)。结论用HCV多表位复合抗原制备的多克隆抗体,采用ELISA双抗体夹心法检测HCAg,具有灵敏、特异、快速的优点,有望开发成为HCAg诊断试剂盒。

Application of HCV Multi-epitope Antibody in Test for HCV Antigen

Objective To explore the possibility of test for hepatitis C virus antigen(HCAg) in plasma or serum samples by using HCV multi-epitope antibody and provide a method for screening of blood donors as well as early diagnosis of hepatitis C.Methods Immunize animals with the multi-epitope complex antigen of HCV expressed in E.coli to prepare HCV polyclonal antibody.Determine the HCAg in plasma or serum samples by double-antibody sandwich ELISA using the prepared polyclonal antibody and compare the determination result with that by RT-PCR.Results The prepared HCV polyclonal antibody showed high specificity(96.30%) and sensitivity(88.89%) in test for HCAg by ELISA.The cutoff value was defined as 0.239.The intra- and inter-variation coefficients(CV) were 5.60%-6.65% and 7.80% respectively.The correlation coefficient of ELISA and RT-PCR was 0.816,and their consistency and adjusted consistency were 95.24% and 90.82% respectively.The positive and negative prediction values of ELISA were 80.00% and 98.11% respectively.The detectable rates of HCAg by ELISA using the prepared HCV polyclonal antibody showed no significant difference from that using HCV-C antigen kit manufactured by ORTHO(P=0.06).Conclusion The prepared HCV polyclonal antibody was sensitive,specific and rapid in determination of HCAg by double antibody sandwich ELISA and might be used for the development of HCAg diagnostic kit.