A flexible high content imaging assay for profiling macrophage efferocytosis.

Macrophages are a diverse population of cells originating from the myeloid lineage, which form an important component of the innate immune system, helping to regulate immune response through secretion of pro/anti-inflammatory cytokines. However they also have an important homeostatic role - helping to remove cellular debris and apoptotic cells from the body (a phagocytic process known as efferocytosis). Here we describe a robust 384 well microplate based imaging assay, using apoptotic target cells for the specific quantification of efferocytosis in human primary monocyte derived macrophages. The methodology described allows for the assay to run in either fixed end-point or live-cell format (the former offering multiple morphological and intensity-based readouts, whilst the latter opens the possibility for future expansion of the methodology to encompass kinetic profiling). Within the methodology described we couple high content image acquisition (on the Cell Voyager 7000S) with multi-parametric image analysis - using Perkin Elmer Columbus combined with GeneData Screener.