A flexible high content imaging assay for profiling macrophage efferocytosis

Area of Science:

• Immunology
• Cell Biology
• Biotechnology

Background:

• Macrophages are key immune cells involved in innate immunity and tissue homeostasis.
• Efferocytosis, the clearance of apoptotic cells by macrophages, is crucial for immune regulation and debris removal.
• Quantifying efferocytosis in human macrophages is essential for understanding immune responses.

Purpose of the Study:

• To develop and validate a robust 384-well microplate-based imaging assay for quantifying efferocytosis.
• To enable specific measurement of efferocytosis in human primary monocyte-derived macrophages.
• To provide a flexible assay format for both fixed-end-point and live-cell analysis.

Main Methods:

• Utilized apoptotic target cells and human primary monocyte-derived macrophages in a 384-well microplate format.
• Employed high-content imaging on the Cell Voyager 7000S for image acquisition.
• Integrated multi-parametric image analysis using Perkin Elmer Columbus and GeneData Screener.

Main Results:

• Successfully established a robust imaging assay for specific quantification of efferocytosis.
• The assay allows for multiple morphological and intensity-based readouts in fixed-end-point format.
• The live-cell format offers potential for future kinetic profiling of efferocytosis.

Conclusions:

• The described microplate imaging assay provides a powerful tool for studying macrophage efferocytosis.
• This methodology facilitates accurate and efficient quantification of efferocytosis in human macrophages.
• The assay's flexibility supports diverse research applications in immunology and cell biology.