荧光定量聚合酶链反应检测乙型肝炎病毒  

Fluorescence quantitative PCR and its application in detection of hepatitis B virus

作　　者：程钢[1] 何蕴韶[1] 周新宇[1] 

机构地区：[1]中山医科大学达安基因诊断中心

出　　处：《中华医学检验杂志》1999年第3期135-138,共4页

基　　金：1998年国家技术创新项目资助

摘　　要：目的用荧光定量聚合酶链反应（ＦＱＰＣＲ）方法准确地定量检测血清中乙型肝炎病毒（ＨＢＶ）的数量和免疫指标，以指导临床。方法设计合成了ＨＢＶＦＱＰＣＲ诊断试剂盒，以一种完全闭管式的ＰＣＲ和荧光探针杂交技术相结合所产生的实时检测定量ＰＣＲ方法，检测了５３２份临床血清标本。结果经ＦＱＰＣＲ检测，１５８份ＨＢＶ的ｓ抗原、ｅ抗原、ｃ抗体（ＨＢｓＡｇ、ＨＢｅＡｇ、ＨＢｃＡｂ）都阳性的标本，其血清标本ＨＢＶＤＮＡ也全部阳性，平均ＨＢＶＤＮＡ拷贝数为１．１×１０８／ｍｌ；９８例ＨＢｓＡｇ、ＨＢｅＡｂ、ＨＢｃＡｂ都阳性的标本，其阳性率为７３％（７１例），平均拷贝数为８．６×１０５／ｍｌ，而常规ＰＣＲ只有３３％的阳性率；６７例ＨＢｓＡｂ、ＨＢｅＡｂ、ＨＢｃＡｂ都阳性标本的平均拷贝数为２．３×１０５／ｍｌ。结论能够避免ＰＣＲ后处理导致的假阳性污染，并实现准确定量。ＦＱＰＣＲ可以检测ＨＢＶ的真实感染和复制情况，对于乙型肝炎的临床诊断，治疗方案的选择和疗效考察有较大的指导意义。Objective To study the relationship of HBV in serum, syndrome of patients, and immune indexes by FQPCR method. Method Fluorescence quantitative PCR (FQPCR), which combines PCR and fluorescence probe hybridization, was used to measure DNA/RNA. Results A clinical diagnostic kit for hepatitis B virus (HBV) was developed. 532 clinical serum samples were tested with this kit, using ELISA as contrast. In 158 HBsAg+/HBeAg+/HBcAb+ samples, FQPCR results were positive, with 1.1108/ml of HBV on average. In 98 HBsAg+/HBeAb+/HBcAb+ samples, the average was 8.6105/ml with a positive rate of 73%; normal PCR only reached 30%. In 67 HBsAb+/HBeAb+/HBcAb+ samples, the amount was 2.3105/ml. Conclusion Intube monitoring eliminates PCR crosscontamination which causes false positive, and realtime detection ensures accurate quantity. So FQPCR can be used to monitor the true state of HBV infection and amplification.

关 键 词：聚合酶链反应 荧光光度法 乙型肝炎病毒 

分 类 号：R512.620.4[医药卫生—内科学]