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Method for Measurement of Viral Fusion Kinetics at the Single Particle Level
Published on: September 7, 2009
Disassembly of structurally modified viral nanoparticles: characterization by fluorescence correlation spectroscopy
Jouni Toivola1, Leona Gilbert, Patrik Michel
1Department of Biological and Environmental Science and Nanoscience Center, P.O. Box 35, University of Jyväskylä, 40014 Jyväskylä, Finland.
Comptes Rendus Biologies
|November 30, 2005
Summary
This study used fluorescence correlation spectroscopy to analyze breakdown products of engineered viral particles. Researchers quantified fluorescent subunits in baculovirus and parvovirus-like particles, revealing insights into viral structure.
Area of Science:
- Biophysics
- Structural Biology
- Virology
Background:
- Engineered viral particles, including enveloped baculovirus and non-enveloped virus-like particles (VLPs), are valuable tools in research.
- Understanding the subunit composition and assembly of these particles is crucial for their effective application.
Purpose of the Study:
- To quantify the number of fluorescent subunits within engineered viral particles.
- To investigate the structural breakdown of viral particles using fluorescence correlation spectroscopy (FCS).
Main Methods:
- Analysis of Autographa californica multicapsid nucleopolyhedrovirus (AcMNPV) displaying green fluorescent protein (GFP)-fused gp64 envelope protein.
- Treatment of AcMNPV with SDS and DTT to induce breakdown.
- Analysis of fluorescent virus-like particles (fVLPs) from canine and human B19 parvoviruses.
- Treatment of fVLPs with urea and SDS, respectively.
- Utilizing fluorescence correlation spectroscopy (FCS) to analyze diffusion times and particle numbers.
Main Results:
- An average of 4.5 molecules of gp64-GFP were estimated per modified baculovirus.
- Hydrodynamic radii of parvovirus-like particles reduced significantly after treatment, indicating structural disassembly.
- Approximately 10 and 9 fluorescent units were associated with canine parvovirus and B19 VLPs, respectively.
Conclusions:
- Fluorescence correlation spectroscopy is effective for estimating the number of fluorescent subunits in engineered viral particles.
- The study provides quantitative data on subunit incorporation in baculovirus and parvovirus-like particles.
- This methodology offers insights into the structural organization of viral components.
