Assessing human iPSC-derived microglia identity and function by immunostaining, phagocytosis, calcium activity, and
Verena Hübschmann1, Medina Korkut-Demirbaş1, Sandra Siegert1
1Institute of Science and Technology Austria (ISTA), Am Campus 1, 3400 Klosterneuburg, Austria.
Abstract:
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.
Insights
This study offers protocols to assess microglia identity and function in vitro. These methods evaluate microglia characteristics and responses, aiding research into gene manipulation effects.
Area of Science:
- Neuroscience
- Cell Biology
- Immunology
Background:
- Microglia are crucial immune cells of the central nervous system.
- Understanding in vitro microglia function is vital for neurological disease research.
- Gene manipulation studies require reliable methods to assess microglia identity and function.
Purpose of the Study:
- To provide standardized protocols for evaluating in vitro microglia.
- To enable assessment of microglia identity and key functional responses.
- To support research on the impact of gene manipulations on microglia.
Main Methods:
- Immunostaining protocols to confirm microglia identity.
- Functional assays including phagocytosis.
- Assessment of calcium responses and cytokine expression upon stimulation.
Main Results:
- Detailed protocols for microglia characterization are presented.
- Methods are validated using human induced-pluripotent-stem-cell (hiPSC)-derived microglia.
- Protocols are adaptable for other in vitro microglia models.
Conclusions:
- The provided protocols offer a robust framework for in vitro microglia research.
- These methods facilitate the study of gene manipulation effects on microglia.
- The assays are applicable across various in vitro microglia systems.
