建立了单管逆转录环介导等温扩增法(RT-LAMP)快速检测甲型H1N1流感病毒的方法.针对甲型H1N1流感病毒的M基因和HA基因的保守区,设计了两组特异性引物,分别用于筛选甲型流感病毒及鉴定甲型H1N1流感病毒.对反应体系中的关键因素进行优化,反应结果可直接通过浊度或者SYBR Green Ⅰ荧光进行判定.本方法最低可检测到10拷贝/管的甲型H1N1流感病毒.为验证方法的可行性,对30例临床样本进行了检测,与美国疾病控制与预防中心(CDC)的TaqMan方法进行对比,检测的灵敏度和特异性分别为92%和100%.本方法实现了单管快速检测甲型H1N1流感病毒,为甲型H1N1流感病毒的现场检测提供了新方法.%A single-tube reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay was developed for the detection of influenza A H1N1 virus. First, two sets of primers were designed based on the conserved regions of both the M gene and the HA gene for the screening of influenza A virus and the identification of influenza A H1N1 subtype. Optimization of RT-LAMP reaction at various conditions was then carried out by using the above primers. Positive reactions can be readily observed by a visual inspection based on the turbidity or SYBR Green Ⅰ fluorescence. The detection limit of this method was 10 RNA copies per reaction, higher than that of the Centers for Disease Control and Prevention TaqMan assay. To investigate the feasibility of the RT-LAMP assay,30 clinical specimens were tested, and the sensitivity and specificity were 92％ and 100％, respectively, in comparison with the Centers for Disease Control and Prevention TaqMan assay. Therefore,the RT-LAMP assay proposed here is a perspective tool for the rapid screening of influenza A H1N1 virus at resource-limited areas.
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