实时定量聚合酶链反应在肾移植后人巨细胞病毒活动性感染的临床应用研究  

Clinical Application of Real-Time Automated PCR to monitoring Human Cytomeglovirus Active Infection after Renal Allografting

作　　者：成俊[1] 谢森[2] 许贤林[3] 史晓峰[1] 夏穗生[1] 

机构地区：[1]华中科技大学同济医学院器官移植研究所,武汉430030 [2]广州军区武汉总医院泌尿外科 [3]江苏省常州市第一人民医院泌尿外科

出　　处：《华南国防医学杂志》2003年第4期4-7,共4页Military Medical Journal of South China

摘　　要：目的:建立血浆、外周血白细胞HCMV DNA实时定量聚合酶链反应检测方法,评价其监测肾移植后人巨细胞病毒活动性感染的临床应用价值。方法:用重组质粒PQHY-1制作标准曲线;分别用实时定量聚合酶链反应和免疫组织化学方法,对28例肾移植术后病人采血标本动态检测,并将结果进行比较。结果:实时定量PCR最低可检测出20拷贝/孔的HCMV DNA,且可重复性强(批内及批间误差均<5％),在102～107拷贝/孔的浓度范围时与Q值有很好的线性关系(r=-0.978)。与PP65抗原血症检测结果比较,血浆及外周血白细胞定时定量检测的灵敏度分别为88.5％,94.2％;特异度分别为97.8％,95.4％。HCMV活动性感染时血浆和外周血白细胞HCMV DNA平均含量分别为102.4拷ml和103.2拷贝/106PBL。血浆和外周血白细胞中HCMV DNA阳性检出时间较PP65抗原血症阳性检出时间平均要早1.6和3.8天。结论:实时定量聚合酶链反应检测方法是一种灵敏度高,特异度好的定量检测方法,可用于监测早期HCMV活动性感染。Objectives 16 assess the clinical applilation of real-time automated PCR to detecting quantatively human cy-tornegalovirus (HCMV) DNA in plasma and peripheral blood leukocytyes(PLB)and investigate its value to monitoring HCMV active infection after kidney transplantation. Methods The recombinant plasmid PQHY - 1 was used to draw standard curve for HCMV real -time PCR. HCMV DNA and its PP65 antigen in the blood speciments from 28 patients who received renal allogrofts were dynamically measured respectively by real - time automated PCR and immunohistochemistry method. The data were analyzed. Results The real-time PCR had high reprodudbility( the coefficient of variation of both the intra - and inter - assay were below 5% ), the linea relationship between the number of copies well and ct value was obtained from 102 to 107 copies/well (correlation coefficient was0.978), with a minimum detection limit of 20 copies/well. Compared the result of real-time PCR with pp65 antigenemia assay, the sensitivity of the real-time PCR in plasma and in PBL were 88.5% and 94.2%,respectively,and the specificity were 97.8% and 95.4%,respectively.The mean contents of HCMV DNA copy in plasma and in PBL were 102. 4 copies/ml and 103.2 copies/106PBL, respectively during HCMV active infection. There was a time lag of 1.6 or 3.8 days on average between plasma or PBL real - time PCR and the occurrence of positive findings in the antigenemia assay . Conclusion The real - time PCR assay is useful for monitoring HCMV active infection, and the method is highly sensitive, specific and quantifiable.

关 键 词：人巨细胞病毒 实时定量聚合酶链反应 PP 65抗原血症 肾移植 外周血白细胞 

分 类 号：Q783[生物学—分子生物学]