The Open Virology Journal

2012, 6 : 97-102
Published online 2012 November 26. DOI: 10.2174/1874357901206010097
Publisher ID: TOVJ-6-97

RESEARCH ARTICLE
 Development of a Flow Cytometry Live Cell Assay for the Screening of Inhibitors of Hepatitis C Virus (HCV) Replication

Jose A Garcia-Rivera1 , Kai Lin2 , Sam Hopkins3 , Matthew A Gregory4 , Barrie Wilkinson4 and Philippe A Gallay, *,1
1 Department of Immunology & Microbial Science, The Scripps Research Institute, La Jolla, California 92037, USA
2 Novartis Institutes for Biomedical Research, Inc., Cambridge, USA
3 SCYNEXIS, Inc., Research Triangle Park, USA
4 Biotica Technology Ltd, Chesterford Research Park, Cambridge, CB10 1XL, UK

* Address correspondence to this author at the Department of Immunology & Microbial Science, IMM-9, The Scripps Research Institute, 10550 N. Torrey Pines Rd., La Jolla, CA 92037, USA; Tel: (858) 784-8180; Fax: (858) 784-8831; E-mail: gallay@scripps.edu

ABSTRACT

In this study, we established a flow cytometry live cell-based assay that permits the screening of hepatitis C virus (HCV) inhibitors. Specifically, we created a stable cell line, which harbors a subgenomic replicon encoding an NS5A-YFP fusion protein. This system allows direct measurement of YFP fluorescence in live hepatoma cells in which the HCV replicon replicates. We demonstrated that this stable fluorescent system permits the rapid and sensitive quantification of HCV replication inhibition by direct-acting antiviral agents (DAA) including protease and NS5A inhibitors and host-targeting antiviral agents (HTA) including cyclophilin inhibitors. This flow cytometry-based live cell assay is well suited for multiple applications such as the evaluation of HCV replication as well as antiviral drug screening.

Keywords:

Direct-acting antiviral, FACS assay, HCV, hepatoma cells, host-targeting antiviral, inhibitors..