Taq Man技术快速定量检测人巨细胞病毒的早期感染

Rapid Quantitative Detection of Human Cytomegalovirus by TaqMan PCR Assay

采用实时TaqMan技术快速定量检测血清中的人巨细胞病毒(HCMV)感染,在HCMV保守性的Ma jorIE基因区段设计了一对引物和一条Taqman探针,在LightCycler及Rotor Gene定量检测仪上检测HCMVDNA.试验证明,两种仪器定量结果具有一致性,以PCR模板浓度来定义,敏感度达到1.0×102拷贝/μL,线性范围为1.0×102～1.0×108拷贝/μL.应用该方法与ELISA法对40例临床婴幼儿患者血清同时进行HCMV检测,结果显示TaqMan法与ELISA法相关性不高,前者更适于HCMV感染早期的定量检测,提示该法可应用于孕妇及新生儿HCMV筛查,并有助于疗效监测和评估.

A real-time TaqMan assay was applied to quickly quantify human cytomegalovirus (HCMV) DNA from human serum. A pair of primes and a TaqMan probe were designed that are specific for the recognition of a conservative Major IE gene in HCMV genome. LightCycler and Rotor-Gene instruments were separately used to quantify HCMV DNA. According to our results of the two instruments, HCMV TaqMan assay has a sensitivity of 1.0×10~2 DNA copy/μL with a dynamic range of detection between 1.0×10~2 and 1.0×10~8 DNA copy/μL. In order to evaluate its performance in clinical diagnosis, TaqMan was used to examine the HCMV DNA levels in sera from 40 sero-positive or negative subjects. The quantification results indicate that the TaqMan assay has low correlation to ELISA and suggest that this approach can apply broadly to early diagnose HCMV-related infections as well as to monitor and evaluate the relevant treatment, especially for pregnant women and newborn children.