Hepatitis C virus: propagation, quantification, and storage.
Hepatitis C virus: propagation, quantification, and storage.
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DOI：
10.1002/9780471729259.mc15d01s19
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发表时间：
2010-11
影响因子：
--
通讯作者：
Yi, MinKyung
中科院分区：
文献类型：
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作者：
Yi, MinKyung
丙型肝炎病毒是慢性肝病的主要病因，包括慢性肝炎、肝硬变和肝细胞癌。它以其有限的趋向性而闻名，除了人类和黑猩猩之外，它在其他动物物种中不能很好地复制。由于经典的天然丙型肝炎病毒分离株的体外繁殖是不可能的，我们提供了一种从作为RNA导入允许细胞的cDNA克隆(1a型pH77S和2a型pJFH-1)中拯救感染性病毒的方法。由于这两个分子克隆在繁殖和产生传染性病毒的能力上表现不同，我们描述了这两个病毒株的不同繁殖方法。我们还提供了感染性病毒滴定的方法，这可以通过对感染细胞进行病毒抗原表达的免疫染色(荧光焦点或FFU分析)来完成。
Hepatitis C Virus (HCV) is a leading cause of chronic liver diseases, including chronic hepatitis, cirrhosis, and hepatocellular carcinoma. It is well known for its restricted tropism and does not replicate well in animal species other than humans and chimpanzees. Since classical in vitro propagation of natural HCV isolates is not possible, we provide a protocol for the rescue of infectious virus from cDNA clones (genotype 1a pH77S and genotype 2a pJFH-1) transfected as RNA into permissive cells. Because these two molecular clones behave differently in their ability to propagate and produce infectious virus, we describe different methods for propagation of these two viral strains. We also provide methods for infectious virus titration, which can be accomplished by counting foci of infected cells following immunostaining for viral antigen expression in cells infected with serial dilutions of a virus harvest (fluorescent focus, or FFU, assay).