Comanor Lorraine, Elkin Claudia, Leung Kimmy, Krajden Mel, Kronquist Kathryn, Nicolas Keith, Horansky Evelyn, deMedina Maria, Kittichai Promrat, Sablon Erwin, Ziermann Rainer, Sherlock Chris
1801 Waverley Street, Palo Alto, CA 94301, USA.
J Clin Virol. 2003 Sep;28(1):14-26. doi: 10.1016/s1386-6532(02)00234-2.
Hepatitis C virus (HCV) genotyping is a critical part of the diagnostic work-up for chronic hepatitis C. The VERSANT HCV line probe assay (LiPA) marketed by Bayer Corporation requires PCR-derived amplicons for genotyping usually obtained from commercial assays, including Amplicor HCV 2.0 (Amplicor 2.0), Amplicor HCV Monitor 2.0, or SuperQuant. Occasionally, PCR-based methods in conjunction with LiPA fail to give a genotyping result. Although most genotyping failures occur among low viral load specimens, some occur in specimens with relatively high viral loads. The Bayer HCV RNA Qualitative assay (HCV TMA), with a limit of detection of approximately 5-10 IU/ml, is more sensitive than other commercial assays.
An HCV genotyping protocol using HCV TMA linked with LiPA (TMA-LiPA) was developed and tested for ability to genotype samples that had previously failed genotyping by PCR-based methods in conjunction with LiPA.
Clinical specimens were obtained from eight independent laboratories in Canada and the US and tested with TMA-LiPA at the Bayer Reference Testing Laboratory. Specimens included those that failed to produce a genotype result when a PCR-based assay was used in conjunction with LiPA and specimens for which genotyping was not attempted because the viral load was below the validated cut-off determined in the laboratory of origin.
TMA-LiPA successfully genotyped 68 of 75 (90.7%) specimens that had failed genotyping by PCR-based methods used in conjunction with LiPA and 36 of 40 (90.0%) specimens that were rejected for genotyping due to low viral load. Moreover, TMA-LiPA assigned subtype for 79 of 107 (73.8%) specimens. Our TMA-LiPA results reflected the distribution of HCV genotypes found in North America, and were 100% concordant with those of Amplicor 2.0 in conjunction with LiPA for control specimens genotyped by both assays. TMA-LiPA may prove useful both in optimizing LiPA performance and genotyping patient specimens.
丙型肝炎病毒（HCV）基因分型是慢性丙型肝炎诊断检查的关键部分。拜耳公司销售的VERSANT HCV线性探针分析（LiPA）需要通过聚合酶链反应（PCR）获得的扩增子进行基因分型，这些扩增子通常从商业检测中获取，包括Amplicor HCV 2.0（Amplicor 2.0）、Amplicor HCV Monitor 2.0或SuperQuant。偶尔，基于PCR的方法与LiPA结合使用时无法得出基因分型结果。尽管大多数基因分型失败发生在低病毒载量标本中，但也有一些发生在病毒载量相对较高的标本中。拜耳HCV RNA定性检测（HCV TMA）的检测限约为5 - 10 IU/ml，比其他商业检测更灵敏。
开发一种使用HCV TMA与LiPA联用（TMA-LiPA）的HCV基因分型方案，并测试其对先前通过基于PCR的方法与LiPA结合进行基因分型失败的样本进行基因分型的能力。
从加拿大和美国的八个独立实验室获取临床标本，并在拜耳参考检测实验室用TMA-LiPA进行检测。标本包括那些在使用基于PCR的检测与LiPA结合时未能产生基因分型结果的标本，以及那些因病毒载量低于原实验室确定的有效临界值而未进行基因分型尝试的标本。
TMA-LiPA成功地对75个通过基于PCR的方法与LiPA结合进行基因分型失败的标本中的68个（90.7%）以及40个因病毒载量低而被拒绝进行基因分型的标本中的36个（90.0%）进行了基因分型。此外，TMA-LiPA为107个标本中的79个（73.8%）确定了亚型。我们的TMA-LiPA结果反映了北美地区HCV基因型的分布情况，并且与通过两种检测方法对对照标本进行基因分型时Amplicor 2.0与LiPA联用的结果100%一致。TMA-LiPA可能在优化LiPA性能和对患者标本进行基因分型方面都很有用。
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