Methodological comparison of FACS and MACS isolation of enriched microglia and astrocytes from mouse brain

J Immunol Methods. 2020 Nov:486:112834. doi: 10.1016/j.jim.2020.112834. Epub 2020 Aug 15.


Microglia and astrocytes, the two innate cells in CNS, are thought to protect and remodel of synapses for proper maintenance and plasticity of neuronal circuits. The two types of cells are the major responders by producing and releasing inflammatory mediators. Isolation of microglia and astrocytes from CNS tissue provides a powerful tool to study basic cell biology and examine the effects of in vivo treatments on microglia and astrocytes immunophenotype and function. The widely used approach of enrichment microglia and astrocytes from CNS was MACS (Magnetic activated cell sorting) and FACS (Fluorescence activated cell sorting). Here we described an optimized protocol of enzymatic dissociation generating single cell suspensions from brain tissue. Then the ability of the two methods to isolate microglia and astrocytes from brain dissociated cells was compared. Both MACS and FACS processing could obtain microglia and astrocytes with high viability (>85%). Microglia sorted by MACS comprises a slight myeloid cells contamination but with a little bit higher efficiency than that sorted by FACS. MACS processing was faster than FACS for either single or multiple samples. ACSA2 can be used to isolate astrocytes from both postnatal and adult brain, and is more suitable for purify astrocytes from newborn. FACS could get purer microglia which is helpful for deep sequencing and other related research. ACSA2 is a good marker of astrocytes.

Keywords: Astrocyte; FACS; Microglia.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Astrocytes / metabolism
  • Astrocytes / physiology*
  • Biomarkers / metabolism
  • Brain / cytology*
  • CD11b Antigen / metabolism
  • Cell Separation / methods*
  • Cell Survival
  • Cells, Cultured
  • Flow Cytometry*
  • Gene Expression Regulation
  • Immunomagnetic Separation*
  • Leukocyte Common Antigens / metabolism
  • Mice, Inbred C57BL
  • Mice, Transgenic
  • Microglia / metabolism
  • Microglia / physiology*
  • Phenotype


  • Biomarkers
  • CD11b Antigen
  • Leukocyte Common Antigens
  • PTPRC protein, human