目的为适应临床上需同时检测庚型肝炎病毒（ＨＧＶ）与丙型肝炎病毒（ＨＣＶ）感染情况，建立了二重逆转录－聚合酶链反应（ＲＴ－ＰＣＲ）及微孔板反向杂交法。方法根据ＨＣＶ与ＨＧＶ基因高度保守区自行设计引物，建立了用二重ＲＴ－ＰＣＲ同时检测ＨＣＶ与ＨＧＶＲＮＡ方法，并分别用ＨＣＶ与ＨＧＶ特异探针微孔板反向杂交检测ＰＣＲ产物。结果ＰＣＲ产物经测序，ＨＣＶ与Ｔａｋａｍｉｚａｗａ等及Ｃｈｏｏ等报道的核苷酸同源性分别为９３１％～９４１％与９２５％～９３７％，ＨＧＶ与Ｓｉｍｏｎｓ等、Ｌｉｎｎｅｎ等及常锦红等的核苷酸同源性分别为９０７％～９２５％、９２０％～９２１％与９４３％～９４５％。测定敏感度约是单式扩增电泳法的１００倍，２０次重复测定ＨＣＶ与ＨＧＶ的ＣＶ值分别为８９％与９８％。微孔板杂交ＮａＯＨ最佳终浓度为０１～０１５ｍｏｌ／Ｌ，最佳杂交时间为３０～５０分钟。１３７例血清样本检测表明，在肝炎患者与血透患者ＨＣＶ单独阳性为１３９％，ＨＧＶ为５１％，ＨＣＶ与ＨＧＶ同时阳性为５８％。结论本文用微孔板杂交酶呈色技术检测ＨＣＶ与ＨＧＶ二重ＲＴ－ＰＣＲ产物敏感、特异、快速，重复性良好，无溴乙锭污染，经? To establish a dual-reverse-transcription-polymerase chain reaction (RT-PCR) and microplate reverse hybridization assay in order to meet the need of simultaneous detection of hepatitis G virus (HGV) and hepatitis C virus (HCV) . Methods Based on the highly conserved regions of HCV and HGV genes, primers were designed and used to detect HCV and HGV RNA simultaneously by double RT-PCR. PCR products were detected by reverse hybridization between HCV and HGV-specific microplate. Results The PCR products were sequenced. The nucleotide homology of HCV and Takamizawa et al. And Choo et al. Were 931% ~ 941% and 925% ~ 937% respectively. HGV, Simons et al., Linnen et al And Chang Jinhong nucleotide homology were 90  7% ~ 92  5%, 92  0% ~ 92  1% and 94  3%  94  5%. The sensitivity of the assay was about 100-fold higher than that of the single-stranded amplification gel electrophoresis. The CV values ​​of HCV and HGV in 20 replicates were 89% and 98%, respectively. Microplate hybridization NaOH optimum final concentration of 0  1 ~ 0  15mol / L, the best hybridization time of 30 to 50 minutes. 137 cases of serum samples showed that in patients with hepatitis and hemodialysis patients HCV positive alone was 13  9%, HGV was 5  1%, HCV and HGV positive at 5  8%. CONCLUSION: This study was designed to detect the sensitivity of HCV and HGV dual RT-PCR products by microplate hybridization and the specificity, rapidity, reproducibility and ethidium bromide-