Published in:
Open Access 01-12-2014 | Technical advance
Automated image-based assay for evaluation of HIV neutralization and cell-to-cell fusion inhibition
Published in: BMC Infectious Diseases | Issue 1/2014
Login to get accessAbstract
Background
Standardized techniques to detect HIV-neutralizing antibody responses are of great importance in the search for an HIV vaccine.
Methods
Here, we present a high-throughput, high-content automated plaque reduction (APR) assay based on automated microscopy and image analysis that allows evaluation of neutralization and inhibition of cell-cell fusion within the same assay. Neutralization of virus particles is measured as a reduction in the number of fluorescent plaques, and inhibition of cell-cell fusion as a reduction in plaque area.
Results
We found neutralization strength to be a significant factor in the ability of virus to form syncytia. Further, we introduce the inhibitory concentration of plaque area reduction (ICpar) as an additional measure of antiviral activity, i.e. fusion inhibition.
Conclusions
We present an automated image based high-throughput, high-content HIV plaque reduction assay. This allows, for the first time, simultaneous evaluation of neutralization and inhibition of cell-cell fusion within the same assay, by quantifying the reduction in number of plaques and mean plaque area, respectively. Inhibition of cell-to-cell fusion requires higher quantities of inhibitory reagent than inhibition of virus neutralization.