Background Macrophages are phagocytic cells of the innate immune system involved in degradation of cellular debris and modulation of immune activity through cytokine and chemokine release. iCell® Macrophages 2.0 are functionally naïve macrophages derived from human induced pluripotent stem cells (iPSC) using a directed differentiation protocol that follows the natural macrophage developmental pathway. This enables an effective, dynamic response to stimulation for a biologically relevant model of human macrophage function.

Methods iCell Macrophages 2.0 can be used directly out of thaw or plated and maintained for up to 14 days in culture. To measure cytokine release, cells were plated for 3 days, then stimulated for 24 hours with LPS, IFNγ, IL-4, IL-13, TGFβ, or IL-10. Supernatants were collected and cytokines quantified using the Luminex multiplex system. Seahorse XF Pro Analyzer was used to measure the metabolic kinetics of stimulated and unstimulated iCell Macrophages 2.0. To assay phagocytosis, iCell Macrophages 2.0 were plated for 3 days, then incubated with pHrodo-labeled bioparticles and monitored for uptake over 5 days. Lastly, an ADCP assay was run on IFNγ-stimulated macrophages to determine the dose response to rituximab.

Results iCell Macrophages 2.0 express macrophage surface markers such as CD68, CD11c, and CD11b. These full function macrophages can be polarized toward a pro- or anti-inflammatory state, with cytokine release comparable to human primary macrophages for IL-6, TNFα, IL-10, and CCL18. This response can be measured within hours in both mRNA and secreted protein and can be detected at stimulant concentrations below 100 pg/ml. Phagocytosis is readily measurable in both short- and long-term cultures. iCell Macrophages 2.0 have a functional ADCP response to antibody-labeled target cells with a larger dynamic range than primary monocyte-derived macrophages.

Conclusions Human iPSC-derived macrophages provide a constant, reproducible source of biologically relevant material for use in immunology and oncology. iCell Macrophages 2.0 are in a naïve functional state, able to respond to pro- or anti-inflammatory stimuli, such as IFNγ or TGFβ. Functionally, cytokine release and phagocytosis assays can be performed anytime from thaw up to 14 days in culture with consistent, reliable lot-to-lot performance. The functional and metabolic profile of iCell Macrophages 2.0 demonstrates their suitability as a continuing source of human macrophages.