Protocol for differentiation and efficient AAV-mediated gene delivery to hiPSC-derived microglia for functional studies

STAR Protoc. 2026 Mar 30;7(2):104455. doi: 10.1016/j.xpro.2026.104455. Online ahead of print.

ABSTRACT

Human induced pluripotent stem cell (hiPSC)-derived microglia (hiMG) constitute a powerful platform to study human microglial biology in health and disease. Here, we present a protocol for microglial differentiation and efficient gene delivery to these cells using adeno-associated virus (AAV). We describe steps for differentiation, viral transduction, and validation using flow cytometry and fluorescence imaging. Finally, to demonstrate the utility of this approach, we outline procedures to record and analyze calcium activity in hiMG using the genetically encoded sensor GCaMP8s. For complete details on the use and execution of this protocol, please refer to Fruhwürth et al.¹ and Heiss et al.².

PMID:41915476 | DOI:10.1016/j.xpro.2026.104455