Assessing human iPSC-derived microglia identity and function by immunostaining, phagocytosis, calcium activity, and inflammation assay

STAR Protoc. 2022 Dec 16;3(4):101866. doi: 10.1016/j.xpro.2022.101866. Epub 2022 Nov 18.


To understand how potential gene manipulations affect in vitro microglia, we provide a set of short protocols to evaluate microglia identity and function. We detail steps for immunostaining to determine microglia identity. We describe three functional assays for microglia: phagocytosis, calcium response following ATP stimulation, and cytokine expression upon inflammatory stimuli. We apply these protocols to human induced-pluripotent-stem-cell (hiPSC)-derived microglia, but they can be also applied to other in vitro microglial models including primary mouse microglia. For complete details on the use and execution of this protocol, please refer to Bartalska et al. (2022).1.

Keywords: Antibody; Cell Biology; Cell culture; Cell-based Assays; Immunology; Microscopy; Neuroscience.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Calcium / metabolism
  • Humans
  • Induced Pluripotent Stem Cells* / metabolism
  • Inflammation / metabolism
  • Mice
  • Microglia* / metabolism
  • Phagocytosis


  • Calcium