Yeh C T, Shyu W C, Sheen I S, Chu C M, Liaw Y F
Liver Research Unit, Chang Gung Memorial Hospital and Medical College, Taipei, Taiwan.
J Virol Methods. 1997 May;65(2):219-26. doi: 10.1016/s0166-0934(97)02187-3.
A method for quantifying hepatitis C virus (HCV) RNA in serum using reverse transcription-polymerase chain reaction (RT-PCR) followed by slot-blot hybridization with a specific, digoxigenin-labeled probe was developed. Using RNA synthesized from cloned HCV cDNA as a standard, serum concentration of HCV RNA above 10 copies/ml can be quantitatively determined. To compare this method with branched DNA (bDNA) assay, 45 serum samples from 26 patients with newly acquired acute hepatitis C (n = 16) or hepatitis C with acute exacerbation (n = 10) were submitted to both assays. HCV RNA in 30 (67%), 12 (27%) and three (6.7%) samples can be quantitatively determined by both, either and none of the two assays, respectively. Using a standardized qualitative HCV RNA detection test (Amplicor HCV test) as a reference, 1 and 0 false positive results were found by bDNA and this assay, respectively. This quantitative assay using RT-PCR and a digoxigenin detection system was comparable to bDNA assay. Since a false positive result was rarely found, this technique can be used as a first line test to screen a large number of samples rapidly and economically.
开发了一种使用逆转录-聚合酶链反应（RT-PCR），随后用特异性地高辛配基标记探针进行狭缝印迹杂交来定量血清中丙型肝炎病毒（HCV）RNA的方法。以从克隆的HCV cDNA合成的RNA作为标准品，可定量测定血清中浓度高于10拷贝/毫升的HCV RNA。为了将该方法与分支DNA（bDNA）测定法进行比较，对来自26例新获得的急性丙型肝炎患者（n = 16）或急性加重期丙型肝炎患者（n = 10）的45份血清样本进行了两种测定。两种测定法分别能对30份（67%）、12份（27%）和3份（6.7%）样本进行定量测定、只能对其中一种进行定量测定以及均无法进行定量测定。以标准化的定性HCV RNA检测试验（Amplicor HCV检测）作为参考，bDNA测定法和本测定法分别发现1例和0例假阳性结果。这种使用RT-PCR和地高辛配基检测系统的定量测定法与bDNA测定法相当。由于很少发现假阳性结果，该技术可用作一线检测方法，以快速且经济地筛查大量样本。
