Abstract
Numerous approaches have been developed to isolate microglia from the brain, but procedures using enzymatic dissociation at 37°C can introduce drastic transcriptomic changes and confound results from gene expression assays. Here, we present an optimized protocol for microglia isolation using mechanical homogenization. We use Dounce homogenization to homogenize mouse brain tissue into single-cell suspension. We then isolate microglia through Percoll gradient and flow cytometry. Isolated microglia exhibit a gene expression pattern without the changes induced by heated enzymatic digestion. For complete details on the use and execution of this protocol, please refer to Clayton et al. (2021).
Original language | English (US) |
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Article number | 101670 |
Journal | STAR Protocols |
Volume | 3 |
Issue number | 4 |
DOIs | |
State | Published - Dec 16 2022 |
Keywords
- Cell isolation
- Cell separation/fractionation
- Classification Description: Cell biology
- Flow cytometry/Mass cytometry
- Gene expression
- Molecular biology
- Neuroscience
- RNAseq
ASJC Scopus subject areas
- General Neuroscience
- General Biochemistry, Genetics and Molecular Biology
- General Immunology and Microbiology