Development of high-content imaging assays for lethal viral pathogens

Area of Science:

• Virology
• Drug Discovery
• High-Content Imaging

Background:

• Filoviruses, including Ebola (EBOV) and Marburg (MARV), are highly pathogenic RNA viruses causing severe hemorrhagic fever with no current human therapeutics.
• The urgent need for effective treatments necessitates the development of advanced screening methods for antiviral drug discovery.

Purpose of the Study:

• To describe challenges and solutions in optimizing high-content imaging bioassays for EBOV and MARV.
• To evaluate biological and imaging variables for robust assay development.
• To demonstrate the utility of single-cell data analysis for identifying antiviral compounds.

Main Methods:

• Optimization of biological variables (e.g., plating density, multiplicity of infection) and imaging parameters (e.g., fields scanned, fluorescence intensity).
• Application of high-content imaging technology for primary drug screening against filoviruses.
• Statistical analysis of single-cell data to address heterogeneity in viral antigen staining.

Main Results:

• Successful development of optimized image-based bioassays for EBOV and MARV.
• Identification of key variables impacting assay performance and reproducibility.
• Demonstration of improved data analysis using single-cell metrics for enhanced screening.

Conclusions:

• Image-based methods are powerful tools for screening antiviral compounds against highly pathogenic filoviruses.
• Optimized high-content assays facilitate the discovery of novel therapeutics for EBOV and MARV infections.
• Single-cell data analysis enhances the accuracy and efficiency of antiviral drug screening.