摘要
目的应用非对称PCR-荧光偏振（FP）技术建立一种同步检测全血中4种主要疱疹病毒（单纯疱疹病毒1／2型，巨细胞病毒、EB病毒）的新方法。方法以人疱疹病毒属通用引物行非对称PCR，扩增从179例样本中抽提的DNA。PCR扩增产物与特异性单纯疱疹病毒1／2型（HSV-1／-2）、巨细胞病毒（CMV）和EB病毒（EBV）寡核苷酸探针混合物温育杂交，荧光偏振检测技术检测杂交液荧光偏振值，据荧光偏振值判断病毒感染类型，并以DNA序列测定结果为参照。结果与DNA序列测定的阳性检测符合率为100％，但DNA序列测定检测均未检出多重混合感染。非对称PCR-FP方法对HSV-1／-2检测的灵敏度达到1．0×10^3拷贝／ml．对EBV和CMV检测的灵敏度达到2．0×10^3拷贝／ml。结论本法对于4种主要疱疹病毒感染的筛查及预防、预后判断具重要价值。
Objective To establish a novel method to detect four major human herpesvirus simultaneously by fluorescence polarization （FP） assay based on asymmetric PCR. Methods A consensus primer system in human herpesvirus was used in an asymmetric PCR. The probes of herpes simplex virus types 1 and 2 （ HSV-1/-2 ）, cytomegalovirus （ CMV ）, and Epstein-Barr virus （ EBV ） labeled with different fluorophores were hybridized respectively with target amplicons. The infection was determined by the increased FP value. The DNA extracted from 179 samples was subjected to FP and sequence assay to evaluate the feasibility of this method. Results The minimum detection level of FP assay was 10 genome copies for HSV-1/-2,20 genome copies for both EBV and CMV. FP assay could detect confections more effectively than sequence assay. Conclusions A practical method was developed for the simultaneous detection of the four major human herpesviruses by FP assay based on an asymmetric PCR.
出处
《临床检验杂志》
CAS
CSCD
北大核心
2009年第6期405-407,共3页
Chinese Journal of Clinical Laboratory Science
基金
国家高技术研究和发展计划(2008AA02Z444)
关键词
人疱疹病毒
荧光偏振
非对称PCR
同步检测
fluorescence polarization
asymmetric PCR
human herpesvirus
simultaneous detection
