Pyne Michael T, Konnick Eric Q, Phansalkar Amit, Hillyard David R
ARUP Institute for Clinical and Experimental Pathology, 500 Chipeta Way, Salt Lake City, UT 84108, USA.
J Clin Microbiol. 2009 Sep;47(9):2872-8. doi: 10.1128/JCM.02329-08. Epub 2009 Jul 22.
The accurate and sensitive measurement of hepatitis C virus (HCV) RNA is essential for the clinical management and treatment of infected patients and as a research tool for studying the biology of HCV infection. We evaluated the linearity, reproducibility, precision, limit of detection, and concordance of viral genotype quantitation of the Abbott investigational use only RealTime HCV (RealTime) assay using the Abbott m2000 platform and compared the results to those of the Roche TaqMan Analyte-Specific Reagent (TaqMan) and Bayer Versant HCV bDNA 3.0 assay. Comparison of 216 samples analyzed by RealTime and TaqMan assays produced the following Deming regression equation: RealTime = 0.940 (TaqMan) + 0.175 log(10) HCV RNA IU/ml. The average difference between the assays was 0.143 log(10) RNA IU/ml and was consistent across RealTime's dynamic range of nearly 7 log(10) HCV RNA IU/ml. There was no significant difference between genotypes among these samples. The limit of detection using eight replicates of the World Health Organization HCV standard was determined to be 7.74 HCV RNA IU/ml by probit analysis. Replicate measurements of commercial genotype panels were significantly higher than TaqMan measurements for most samples and showed that the RealTime assay is able to detect all genotypes with no bias. Additionally, we showed that the amplicon generated by the widely used Roche COBAS Amplicor Hepatitis C Virus Test, version 2.0, can act as a template in the RealTime assay, but potential cross-contamination could be mitigated by treatment with uracil-N-glycosylase. In conclusion, the RealTime assay accurately measured HCV viral loads over a broad dynamic range, with no significant genotype bias.
丙型肝炎病毒（HCV）RNA的准确和灵敏检测对于感染患者的临床管理和治疗至关重要，也是研究HCV感染生物学的研究工具。我们使用雅培m2000平台评估了雅培仅供研究使用的实时HCV（RealTime）检测法的线性、可重复性、精密度、检测限以及病毒基因型定量的一致性，并将结果与罗氏TaqMan分析物特异性试剂（TaqMan）和拜耳Versant HCV bDNA 3.0检测法的结果进行了比较。对216份通过RealTime和TaqMan检测法分析的样本进行比较，得出以下Deming回归方程：RealTime = 0.940（TaqMan）+ 0.175 log(10) HCV RNA IU/ml。两种检测法之间的平均差异为0.143 log(10) RNA IU/ml，并且在RealTime近7 log(10) HCV RNA IU/ml的动态范围内保持一致。这些样本中不同基因型之间没有显著差异。通过概率分析，使用八份世界卫生组织HCV标准品重复检测确定检测限为7.74 HCV RNA IU/ml。对于大多数样本，商业基因型检测板的重复测量结果显著高于TaqMan测量结果，表明RealTime检测法能够无偏差地检测所有基因型。此外，我们还表明，广泛使用的罗氏COBAS Amplicor丙型肝炎病毒检测法2.0版产生的扩增子可以作为RealTime检测法的模板，但通过尿嘧啶-N-糖基化酶处理可以减轻潜在的交叉污染。总之，RealTime检测法在广泛的动态范围内准确测量了HCV病毒载量，且无显著的基因型偏差。
