HCV抗原ABS-ELISA检测方法的建立及初步评价  

Development and preliminary evaluation on ABS-ELISA for detecting HCV antigen

作　　者：谢忠平[1,2] 龙润乡[1,2] 杨蓉 李华[1,2] 白惠珠 蒋蕊鞠[1,2] 易红昆 董承红[1,2] 

机构地区：[1]中国医学科学院北京协和医学院医学生物学研究所 [2]云南省重大传染病疫苗研究重点实验室,昆明650118

出　　处：《现代预防医学》2012年第21期5617-5619,共3页Modern Preventive Medicine

基　　金：国家863项目(2007AA02Z480);云南省联合支持国家计划项目(2008GA008)

摘　　要：目的建立新的HCV抗原生物素-亲和素-酶联免疫检测方法(ABS-ELISA)。方法将生物素-亲和素系统与酶联免疫技术相结合,建立用于丙型肝炎病毒抗原检测的方法,分析检测方法的灵敏度、特异度、稳定性及精确度等主要技术指标,并对应用效果进行评价。结果建立的检测方法,对40余种肠道病毒样品均无交叉反应,在第二代HCV-RNA国家标准品的10份阴性标准品中,未检出HCV-Ag阳性样品,在10份HCV-RNA阳性标准品中,检出6份HCV-Ag阳性的样品,表现出良好的特异性;检测试剂CV平均值为6.28%,精密性好;在90份HCV抗体阳性样品中检测出HCV-RNA样品67份(74.44%),检出HCV游离抗原阳性样品30份(33.33%)、HCV总抗原阳性样品68份(75.56%),HCV游离抗原与HCV-RNA两个指标检出率差异有统计学意义(χ2=22.443,P﹤0.01),但HCV总抗原与HCV-RNA检测结果之间差异无统计学意义(χ2=0.037,P﹥0.05);以HCV-RNA检测试剂为金标准,所建立的ABS-HCV抗原检测试剂总抗原检测结果的一致性、灵敏度、特异度、阳性预测值及阴性预测值分别为98.03%、80.60%、98.93%、79.41%、99.00%。结论新建立的HCV抗原ABS-ELISA检测方法特异性好,其灵敏度与荧光定量PCR相近。OBJECTIVE To develop a new detection method for Hepatitis C virus（HCV） antigen.METHODS The biotinavidin system was employed in an enzyme-linked immunosorbent assay,using which HCV standards and samples were checked.The sensitivity,specificity,reproducibility,and accuracy of the assay were analyzed,and also the application potentials of the developed method were evaluated.RESULTS There was no cross-reactivity presented to more than 40 kinds of enterovirus samples.No positive result was found in the measurement of 10 negative controls of the national second-generation HCV-RNA standards,while 6 of 10 HCV antigen-positive standards were detected,showing a nice specificity of the test.The assay precision was good with 6.28% of average CV;In 90 HCV antibody-positive samples,67 for HCV-RNA（74.44%）,30 for free antigen（33.33%）,and 68 for total antigen were detected respectively.The difference of positive rates between HCV free antigen and HCV-RNA tests was statistically significant（χ 2 = 22.443,P 0.01）.However,the results between HCV total antigen and HCV-RNA tests showed no significant differences（χ 2 = 0.037,P ﹥0.05）.With HCV-RNA test served as a gold standard,the consistency,sensitivity,specificity,positive predictive value,and negative predictive value for the detection of HCV total antigen by using the established method were 98.03%,80.60%,98.93%,79.41%,and 99.00%,respectively.CONCLUSION The newly developed ABS-ELISA for detecting HCV antigen shows an excellent specificity and sensitivity comparable to that of a real time quantitative PCR.

关 键 词：丙型肝炎病毒 抗原 酶联免疫吸附 生物素 亲和素 

分 类 号：R373.21[医药卫生—病原生物学]