| 三种丙型肝炎包膜感染性假病毒颗粒的制备及初步应用研究 |
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| 引用本文： | 张柯,谭文杰,邓瑶,李津,吴小兵,阮力. 三种丙型肝炎包膜感染性假病毒颗粒的制备及初步应用研究[J]. 病毒学报, 2008, 24(4): 287-294 |
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| 作者姓名： | 张柯 谭文杰 邓瑶 李津 吴小兵 阮力 |
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| 作者单位： | 张柯 (中国疾病预防控制中心,病毒病预防控制所,北京,100052)； 谭文杰 (中国疾病预防控制中心,病毒病预防控制所,北京,100052)； 邓瑶 (中国疾病预防控制中心,病毒病预防控制所,北京,100052)； 李津 (中国天坛生物制品总公司,北京,100024)； 吴小兵 (中国疾病预防控制中心,病毒病预防控制所,北京,100052)； 阮力 (中国疾病预防控制中心,病毒病预防控制所,北京,100052)； |
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| 基金项目： | 国家高技术研究发展计划(863计划)(项目编号：2007AA022455)国家自然科学基金(项目编号：30571673;30671961) |
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| 摘 要： |
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Development of infectious pseudo-particle harboring three subtypes hepatitis C virus glycoproteins and their application in neutralization assays |
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| Ke Zhang,Wen-jie Tan,Yao Deng,Jing Li,Xiao-bing Wu,Li Ruan. Development of infectious pseudo-particle harboring three subtypes hepatitis C virus glycoproteins and their application in neutralization assays[J]. Chinese journal of virology, 2008, 24(4): 287-294 |
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| Authors: | Ke Zhang Wen-jie Tan Yao Deng Jing Li Xiao-bing Wu Li Ruan |
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| Affiliation: | National Institute for Viral Disease Control and Prevention, China CDC, Beijing 100052, China. |
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| Abstract: | In this study, three expression vectors encoding unmodified glycoproteins E1 and E2 from H77 (1a), Hebei (1b) and JFH1 (2a) strains were constructed to form pVRC-H77-E1E2, pVRC-HeBei-E1E2 and pVRC-JFH1-E1E2 expressing constructs. The protein expression was confirmed by immunofluorescene assay(IFA) and Western blot. The Lentiviral vector has the ability to package the cellular membrane into pseudo-particles. The plasmid expressing HCV E1-E2 glycoproteins in native form was co-transfected into 293FT cells with a lentiviral packaging plasmid (pHR'CMV delta R8.2)and a self-inactivated (SIN) transfer plasmid (pCS-CG) containing a reporter EGFP gene to produce infectious HCV pseudo-particles(pp). Flow cytometry assays showed that the HCVpp could infect Huh7 and Huh7-CD81, and the infectivity in Huh7-CD81 was about 2-3 times higher than that in Huh7 cells. Meanwhile, HCVpp could neither infect non-liver cells, for example, the 293 cells, nor HepG2 cell . Titration of HCVpp by p24 ELISA assay or infection assay showed that this HCVpp may contain 5-25 ng/mL p24 or 10(4)-10(5) TU (transducing unit)/ ml. An in vitro HCV neutralizing assays based on HCVpp (1a, 1b, 2a) were then established using AP33, a monoclone antibody with cross-neutralizing ability to different HCV strains. The neutralizing ability of the antibodies from HCV infected patients was further studied with this HCVpp system. In summary, three kinds of HCVpp (1a, 1b, 2a subtype) were successfully developed; In vitro HCV neutralizing assays based on HCVpp and SIN lentiviral system were established. This system paves a way for characterization of early steps of HCV infection (host tropisms, receptor binding, membrane fusion, et al. ) or screening anti-HCV drugs (such as inhibitor to virus entry). This system can be further applied to assess the human immune responses in HCV patients or evaluate HCV vaccine candidates. |
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