Automated image-based assay for evaluation of HIV neutralization and cell-to-cell fusion inhibition

Abstract

BackgroundStandardized techniques to detect HIV-neutralizing antibody responses are of great importance in the search for an HIV vaccine.MethodsHere, 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.ResultsWe 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.ConclusionsWe 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.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2334-14-472) contains supplementary material, which is available to authorized users.

Scaling up delivery of HIV services in Africa through harnessing trends across global emerging innovations

“…This is based on AI image analysis for the diagnosis of HIV from test images. Several reports and reviews have highlighted the potential of AI in HIV services especially for instant diagnosis; however, few apps have been clinically tested and deployed for clinical use (56).…”

Where in the Cell Are You? Probing HIV-1 Host Interactions through Advanced Imaging Techniques

“…Notably, an automated image-based assay to evaluate antibody neutralization of HIV and inhibition of cell–cell fusion has been implemented allowing screening for neutralizing antibodies which may act as inhibitors of viral entry [113]. Particularly, in line with attempts to develop an effective HIV-1 vaccine, part of the evaluation process is the assessment of the neutralizing capacity of antibody responses.…”

“…U87.CD4 or GHOST(3) cells expressing HIV receptors have been exposed to HIV-1 and imaged for plaque formation and plaque area using automatic image acquisition in an automated plaque reduction assay [113]. In this case, antibody neutralization of free virus is indicated via a reduction in plaque number and a reduction in plaque area corresponds to an inhibition of cell–cell fusion (syncytia) which is characteristic of infected cells [113]. Computational analysis quantifies the number of plaques and plaque area, giving high-throughput analysis of therapeutic efficacy.…”

Uncertainty in cell confluency measurements

“…Automated cell image analysis is employed for several research purposes: to measure single cell and colony shape, density, size, location, co-localization, texture, intensity and several other features in cell culture [8,9]; to measure the cell morphometry in bone growth in vivo studies [10]; to analyze cell migration in living cells in culture [11]; to analyze the biomarker distribution in tumor cells [12]; to detect the presence of tumor cells in tissue samples [13]: to measure HIV particles neutralization performed by neutralizing antibodies [14].…”