应用半巢式核酸荧光定量技术检测肾移植术后人巨细胞病毒感染

Application of semi nest nucleo fluorescent quantitative technique for the diagnosis of human cytomegalovirus in renal transplant recipients

目的：建立一种新的用于肾移植术后人巨细胞病毒（ＨＣＭＶ）感染的快速定量诊断方法。方法：采用半巢式核酸荧光定量技术（ＡｍｐｌｉＳｅｎｓｏｒ－ＰＣＲ）检测已知含量的ＨＣＭＶ－ＡＤ１６９标准病毒株、无关病毒、无关细菌及临床血标本的ＨＣＭＶ－ＤＮＡ和质粒标准品，以确定该方法的敏感性、特异性和定量的准确性。结果：对已知含量的标准病毒株的定量检测结果与其已知含量基本一致；无关病毒包括单纯疱疹病毒１型（ＨＳＶ－１）、单纯疱疹病毒２型（ＨＳＶ－２）、ＥＢ病毒（ＥＢＶ）、丙型肝炎病毒（ＨＣＶ）及无关细菌包括金黄色葡萄球菌、结核杆菌等均为阴性；文中还检测肾移植受者２０例，结果半巢式核酸荧光定量技术的阳性率高于白细胞中的ＨＣＭＶ抗原（ＨＣＭＶ－Ａｇ）血症的阳性率；ＨＣＭＶ质粒标准品的最高检测敏感度为２５０拷贝／毫升；使用该方法标本处理简单，６小时内得出结果。结论：该方法快速，定量较准确，特异性、敏感性较高。

Objective: To establish a new and fast quantitative method for the diagnosis of human cytomegalovirus(HCMV) infection in renal transplant recipients Method: In order to determine the specificity, sensitivity and accuracy of semi nest nucleo fluorescent quantitative technique (AmpliSensor PCR), standard HCMV AD 169 strain with known quantity, non relevant viruses and bacterium as well as HCMV DNA in clinical blood samples and HCMV standard plasmid were detected in this research Results: The result in detection of HCMV standard strain by AmpliSensor PCR was roughly identical with its actual quantity which was known in advance; non relevant viruses, such as herpes simplex virus 1, herpes simplex virus 2, Epstein Barr vurus, hepatitis C virus and bacteria, such as Staphylococcus aureus and tubercle bacilli were also detected and the results were all negative; In this research, 20 cases of renal transplant recipients were studied as well, the result showed that the positive rate with AmpliSensor PCR was higher than that of HCMV Ag test; the top sensitivity of HCMV standard plasmid was 250 copies/mL; The whole experiment could be finished in 6 hours with simple processing of sample Conclusion: This method is rapid, specific, sensitive and can be accurately quantitated, It can be widely used in clinical practice