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Protocol for generating a human iPSC-derived tri-culture model to study interactions between neurons, astrocytes, and
Alexandra M Lish1, Paige C Galle1, Gwendolyn A Orme1
1Ann Romney Center for Neurologic Diseases, Department of Neurology, Brigham and Women's Hospital and Harvard Medical School, Boston, MA, USA.
STAR Protocols
|October 24, 2025
Summary
Researchers developed a novel tri-culture system using human induced pluripotent stem cells (hiPSCs) for studying brain cell interactions. This cryopreservation-compatible method enables reproducible co-culture of neurons, astrocytes, and microglia for disease modeling and drug discovery.
Area of Science:
- Neuroscience
- Stem Cell Biology
- Biotechnology
Background:
- Human induced pluripotent stem cells (hiPSCs) are valuable tools for disease modeling and drug screening.
- Standardized methods for co-culturing multiple neural cell types from hiPSCs are limited, hindering complex brain research.
Purpose of the Study:
- To present a standardized, cryopreservation-compatible tri-culture system for human neurons, astrocytes, and microglia derived from hiPSCs.
- To establish a reproducible platform for investigating dynamic interactions between distinct human brain cell types.
Main Methods:
- Induced pluripotent stem cells (iPSCs) were transduced with cell type-specific factors.
- Intermediate cryopreserved stocks of differentiated cell populations were generated.
- Neurons, astrocytes, and microglia were differentiated and assembled into a tri-culture system.
Main Results:
- A reproducible tri-culture protocol was established using hiPSC-derived neurons, astrocytes, and microglia.
- The system supports the study of cell-cell interactions in a physiologically relevant context.
- Cryopreservation compatibility allows for flexible generation and storage of cell stocks.
Conclusions:
- The developed tri-culture system offers a robust platform for studying human brain cell biology.
- This method facilitates advanced disease modeling and drug screening applications.
- The protocol enhances the utility of hiPSC models for neurological research.
