摘要
目的 建立时间分辨免疫荧光定量分析乙型肝炎病毒PreS1抗原的方法.方法 以基因重组的带有PreS1抗原的乙型肝炎表面抗原作标准品,应用双抗体夹心免疫荧光分析方法,用抗PreS1单克隆抗体和抗-HBs分别作为固相抗体和铕标记抗体,若样本中含有PreS1抗原,则形成抗体-抗原-铕标记抗体复合物,加增强液解离铕离子,采用时间分辨荧光测量技术,对乙型肝炎病毒PreS1抗原进行检测.结果 自制TRFIA试剂检测PreS1抗原,单侧95%参考范围为0～0.26 ng/ml;自制TRFIA试剂与ELISA试剂对乙型肝炎患者血清标本中PreS1抗原的检测,TRFIA方法灵敏度高于ELISA方法,250份乙型肝炎患者血清标本中有3例标本ELISA方法结果为阴性,而用TRFIA方法可检测到PreS1抗原;对于弱阳性标本用ELISA方法检测不到时,TRFIA方法仍可检出.ELISA方法在一阳性标本1：4 096倍稀释时结果为阴性,而TRFIA方法在1：16 384倍稀释时仍为阳性;高、中、低三个浓度,批内和批间精密度测试变异系数（CV,%）均小于10%,平均回收率为103.3%;TRFIA方法检测PreS1抗原与HBsAg及HBeAg无交叉反应;50份我院正常体检者血清标本进行PreS1抗原的检测,结果均正常,特异性100%.结论 TRFIA方法定量检测PreS1抗原,灵敏度高,特异性强,能够准确及时敏感地反映患者体内乙型肝炎病毒的复制情况.
Objective To establish a time-resolved fluoroimmunoassay （TRFIA） for the quantitative detection of hepatitis B virus （HBV） PreS1 antigen. Methods The monoclonal antibody of HBV PreS1 antigen was covered on the microwell plate , incubated with HBV PreS1 antigen in sample and added Eu3^＋ labeled antibody of HBV then enhancement solution to leave the ion of Eu3^＋ and detect the HBV PreS1 antigen in sample by the time-resolved fluoroimmunoassay instrument. Results Single 95% side reference scope was 0 -0. 26 ng/ml . The method of TRFIA was more sensitive than ELISA. Three sample were negeative in ELISA but positive in TRFIA the weak positive specimen, the ELISA can not detect but the THFIA can do. TRFIA can detect PreS1 antigen in a strong positive specimen in it＇s 1：16 384 dilutions ,but the PreS1 antigen can not be detected by the ELISA in it＇s 1：4 096 dilutions. The intra- and inter-assay coefficients of variation were less than 10% in high, medium and low densities respectively. The recovery rate was 103.3%. The HBsAgs and HBeAgs did not have crossing reaction in the examination of PreS1 antigen in TRFIA. The result of 50 normal specimen were all negative, the spectificity was 100%. Conclusion The method of TRFIA raise the sensitivity and specificity for the detection of PreS1. The result was quantitive. It can accurately reflect the replication circumstance of hepatitis B virus in time.
出处
《中华检验医学杂志》
CAS
CSCD
北大核心
2006年第5期407-410,共4页
Chinese Journal of Laboratory Medicine
