抗原捕获聚合酶链式反应（AC－PCR）直接分型检测单纯疱疹病毒

DETECTION AND DIRECT TYPING OF HERPES SIMPLEX VIRUS BY ANTIGEN CAPIURE POLYMERASE CHAIN REACTION (AC-PCR)

用抗单纯疱疹病毒（ＨＳＶ）型共同性ｇＣ和ｇＤ羊克隆抗体（ＭｃＡｂ），包被即Ｅｐｐｅｎｄｏｒｆ管，捕捉ＨＳＶ，同时加入３个引物：一个是ＨＳＶ─１／ＨＳＶ─２型共同性上游引物，另两个分别是ＨＳＶ─１和ＨＳＶ─２型特异性下游引物。借此建立了能直接分型检测ＨＳＶ的抗原捕获聚合酶链式反应（ＡＣ─ＰＣＲ）。ＨＳＶ─１的扩增产物为４７７ｂｐ，ＨＳＶ─２的为３９９ｂｐ两型病毒经ＡＣ─ＰＣＲ扩增后产生分子量不同的ＤＮＡ片段，致使ＡＣ─ＰＣＲ能直接分型检测ＨＳＶ。ＨＳＶ─１和ＨＳＶ─２扩增产物的克隆和序列分析表明，本方法特异性好。用本法检测Ｂａｌｂ／ｃ幼鼠中枢神经系统ＨＳＶ感染的脑标本，进一步证实本方法不仅敏感、特异，而且分型准确。

method for the detection and direct typing of HSV by antigen capture PCR(AC-PCR) technique has been developed. One type-common upstream primer and twotype-specific downstream primerswere prepared to amplify DNA from the HSV type 1 ortype 2 DNA polymerase gene. HSV-1 or HSV-2was capturec by anti-HSV gC and gDMcAbs coating to 0.5ml polypropylene microcentrifuge tubes. Using these three primerssimultaneously in the AC-PCR reaction mixtures, both types of HSV DNA were amplified toproduce products of different sizes. By direct gel analysis, the products of standard HSV-1and HSV-2 strains had the predictive sizes of 477 and 399bp, respectively, and difference inmolecular weight enabled us to type the HSVstrain. The products amplified from samples ofHSV-1 strain 17syn￣+ and HSV-2 Sav werecloned into pUC18 plasmids and sequenced.The result of DNA sepuences indicates that theAC-PCR assay is specific. The sensitivity ofAC-PCR is very high, 10pfu HSV-1 or 1pfu HSV-2 can be detected and typed.