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Published on: September 15, 2014
Protocol for differentiation and efficient AAV-mediated gene delivery to hiPSC-derived microglia for functional
Christina N Heiss1, Anastasia Mihai2, Stefanie Fruhwürth1
1Department of Psychiatry and Neurochemistry, Institute of Neuroscience and Physiology, University of Gothenburg, Gothenburg, Sweden.
STAR Protocols
|March 31, 2026
Summary
This study details a protocol for generating human induced pluripotent stem cell-derived microglia (hiMG) and delivering genes using adeno-associated virus (AAV). The method enables studying microglial function and calcium activity, advancing research in neuroinflammation and neurological diseases.
Area of Science:
- Neuroscience
- Stem Cell Biology
- Genetics
Background:
- Human induced pluripotent stem cell-derived microglia (hiMG) are crucial for studying microglial biology in health and disease.
- Existing methods for microglial research have limitations in efficiency and applicability.
Purpose of the Study:
- To present a comprehensive protocol for hiMG differentiation and gene delivery using adeno-associated virus (AAV).
- To establish a method for analyzing microglial function, specifically calcium activity, in a disease context.
Main Methods:
- Differentiation of hiPSCs into microglia.
- Gene delivery to hiMG using AAV vectors.
- Validation of hiMG characteristics via flow cytometry and fluorescence imaging.
- Recording and analysis of calcium activity using GCaMP8s.
Main Results:
- Successfully differentiated hiMG with high efficiency.
- Demonstrated efficient gene transduction in hiMG using AAV.
- Validated hiMG identity and functionality through multiple assays.
- Successfully recorded and analyzed calcium transients in hiMG.
Conclusions:
- The developed protocol provides a robust platform for studying human microglia.
- This method facilitates genetic manipulation and functional analysis of hiMG, advancing research in neurological disorders.
- The protocol enables detailed investigation of microglial calcium signaling pathways.
