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Updated: Mar 7, 2026
Using Fluorescence Activated Cell Sorting to Examine Cell-Type-Specific Gene Expression in Rat Brain Tissue
Published on: May 28, 2015
ACSA2 is not astrocyte-specific: implications for cell sorting strategies in the rodent brain
Emerson Daniele1, Gabriel Khelifi2,3,4, Daniel Beretta5
1Institute of Medical Science, University of Toronto, Toronto, ON, Canada.
Introduction:
Astrocyte-specific cell surface antigen-2 (ACSA2) has been established as the gold-standard marker for isolating astrocytes via magnetic-activated cell sorting (MACS) or fluorescence-activated cell sorting (FACS) for downstream transcriptomic studies. In a prior study of the astrocyte response to cortical stroke, we used ACSA2-based cell sorting prior to single cell RNA sequencing (scRNAseq). We found a substantial population of ACSA2+ cells exhibiting robust microglial gene expression signatures, suggesting contamination of purified astrocyte preparations.
Methods:
An intravenous antibody labeling strategy coupled with flow cytometry was used to determine whether contamination originated from circulating immune cells or microglia.
Results:
Contaminating cells were identified as CNS-resident microglia that express CD45, CD11b, and ACSA2.
Discussion:
These findings caution against the usage of ACSA2 for astrocyte purification without exclusion markers to achieve high-purity astrocyte populations for downstream multi-omics analyses.
