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A luciferase-based budding assay for Ebola virus.
Sarah E McCarthy1, Jillian M Licata, Ronald N Harty
1Department of Pathobiology, School of Veterinary Medicine, University of Pennsylvania, 3800 Spruce St., Philadelphia, PA 19104, USA.
Journal of Virological Methods
|July 14, 2006
Summary
Ebola virus matrix protein VP40 budding efficiency can be measured using a new high-throughput assay. This assay tracks firefly luciferase incorporated into virus-like particles (VLPs) to identify potential Ebola virus egress inhibitors.
Area of Science:
- Virology
- Molecular Biology
- Biochemistry
Background:
- Ebola virus protein VP40 is crucial for virus egress.
- A functional budding assay was developed to study VP40's role in virus budding.
Purpose of the Study:
- To develop a high-throughput assay for measuring VP40 budding efficiency.
- To assess the incorporation of reporter proteins into virus-like particles (VLPs).
Main Methods:
- Modification of a functional VP40 budding assay for high-throughput screening.
- Quantification of firefly luciferase within VP40-containing VLPs.
- Assessment of VP40 budding efficiency with and without Ebola virus glycoprotein (GP).
Main Results:
- Firefly luciferase was successfully incorporated into budding VP40 VLPs.
- Co-expression of EBOV GP enhanced VLP release containing VP40 and luciferase.
- A budding-deficient VP40 mutant significantly reduced luciferase levels in VLPs.
Conclusions:
- The developed assay provides a direct and quantifiable measure of VP40 budding efficiency.
- This high-throughput assay is a promising tool for identifying inhibitors of Ebola virus budding and egress.