Related Experiment Video
Updated: Aug 11, 2026
13:04
A Protocol for Analyzing Hepatitis C Virus Replication
Published on: June 26, 2014
A novel high throughput screening assay for HCV NS3 helicase activity
M Hicham Alaoui-Ismaili1, C Gervais, S Brunette
1Department of Virology, BioChem Pharma Inc., Québec, Laval, Canada.
Antiviral Research
|June 27, 2000
Summary
A new Flashplate assay accurately measures Hepatitis C virus (HCV) NS3 helicase activity. This reproducible method is ideal for high-throughput screening of potential antiviral drugs targeting HCV.
Area of Science:
- Biochemistry
- Virology
Background:
- Hepatitis C virus (HCV) NS3 helicase is a key viral enzyme.
- Developing efficient assays for measuring helicase activity is crucial for drug discovery.
Purpose of the Study:
- To develop and validate a novel, high-throughput assay for measuring HCV NS3 helicase activity.
- To biochemically characterize the HCV NS3 helicase using the developed assay.
Main Methods:
- Utilized Flashplate technology with a biotinylated and radiolabeled DNA duplex substrate.
- Immobilized the substrate on neutravidin-coated plates.
- Measured helicase activity by detecting the release of the radiolabeled oligonucleotide.
Main Results:
- The assay demonstrated proportionality between DNA substrate, NS3 concentration, and unwinding activity.
- Optimal unwinding occurred within 60 minutes and was dependent on ATP and various ion concentrations (MgCl2, MnCl2, KCl) and pH.
- The assay showed high reproducibility across multiple reactions.
Conclusions:
- The developed Flashplate assay is a fast, convenient, and reproducible method for measuring HCV NS3 helicase activity.
- This assay is suitable for high-throughput screening of potential inhibitors targeting HCV NS3 helicase.
