PCR技术检测尿中人巨细胞病毒的方法学研究

Detection of Human Cytomegalovirus in Urine by Polymerase Chain Reaction

应用ＨＣＭＶ早、晚期两对ＤＮＡ引物，建立了聚合酶链反应（ＰＣＲ）检测尿中人巨细胞病毒（ＨＣＭＶ）的方法。肾移植术后ＨＣＭＶ感染的临床尿样检测结果表明，该方法具有快速、特异性强、灵敏度高等优点，与血清ＨＣＭＶ－ＩｇＭ检测结果符合率为８３．３％。使用早、晚期基因组的双引物分别对临床尿样进行ＰＣＲ扩增，减少了由于自然点突变所致的假阴性，且引物与ＨＣＭＶ同属其它病毒及组织ＤＮＡ间无交叉反应；以ＨＣＭＶ组织培养液作模板，灵敏度可达６２５ｆｇ。本方法为诊断肾移植受者ＨＣＭＶ感染提供了新的手段，适于临床推广。

Polymerase chain reaction(PCRamplification was set up with double primer pairs of major immediate-early and late gene in order to detect human cytomegalovirus(HCMV)from urine.This method was appliedto the detection of HCMV in clinical samples of urine from renal transplant recipiens.The result indicatedthat the primers did not cross react with other members of the herpes family of virus and human genomicDNA;that the HCMV AD169 tissue culture mixture used and the dilutions to estimate the sensitivity of PCRrelative to tissue culture,2.5μl of a 10￣(-3) dilution of the culture(625fg HCMV DNA)assayed were detectedby direct gel analysis; that 25 of 30 samples of urine from the renal transplant recipients (serum HCMV IgMpositive)were positive. It is conclude that PCR amplification is a valuable tool for diagnoses of HCMV infec-tion in renal transplant recipients.