A Guide to Glial Cell Markers

Central nervous system (CNS) glia are cells that support neurons and protect them from injury. Four primary types of glia exist: oligodendrocytes, astrocytes, microglia, and ependymal cells. In the human brain, glia are as numerous as neurons. Most of these glia are oligodendrocytes (45–75%) or astrocytes (19–40%); relatively few are microgila (<10%) or ependymal cells. This guide summarizes the markers most commonly used to identify each of these cell types. 

Oligodendrocyte Markers

The most prominent glial cell in the CNS, oligodendrocytes, serve a vital function: sheathing neuronal axons with a fatty substance called myelin. The primary purpose of myelin is to speed the transmission of electrical signals along axons. Degradation of myelin can have dramatic behavioral consequences, as demonstrated by multiple sclerosis, which is characterized by loss of coordination, vision loss, pain, and fatigue, among other physical, mental, and psychiatric symptoms.

Oligodendrocyte differentiation begins with oligodendrocyte progenitor cells (OPCs). These bipolar cells provide a renewable pool of oligodendrocytes throughout development and adulthood. Early OPC transcriptional regulators such as SOX10, NKX2.2, OLIG1, and OLIG2 can serve as markers of this cell type. More commonly, though, PDGFRα is used to identify OPCs. CSPG4 (more commonly known as NG2) as well as gangliosides recognized by the A2B5 antibody can support the identification of OPCs, but these markers also label neurons and other glial progenitors.

Image: The figure above highlights different glial cell types and commonly associated protein markers. The original, unmodified image is obtained from Wikimedia Commons (artwork by Holly Fischer).

As OPCs differentiate into oligodendrocytes, they continue to express OLIG1, OLIG2, SOX10, and NKX2-2 and begin to express myelin-associated proteins such as proteolipid protein 1 (PLP1). Upregulation of MYRF and Zfp488 instructs oligodendrocytes to produce additional proteins that participate in myelin formation, including myelin basic protein (MBP), MAG, myelin oligodendrocyte protein (MOG), and CNP. A switch from sulfatide to galactocerebroside (also known as GalC) marks the initiation of myelination; GalC can be detected using the O1 antibody.

Explore more oligodendrocyte markers in our guide to oligodendrocyte markers. 

Astrocyte Markers

Astrocytes are star-shaped glial cells that originate from the same precursor cell as OPCs. They perform many homeostatic functions, ranging from regulating the blood brain barrier and cerebral blood flow to supplying neurons with key neurotransmitter building blocks and synaptic support.

Astrocyte differentiation is transcriptionally regulated by SOX9 and nuclear factor 1 (NF1) family member NF1A. SOX9 in particular is highly specific to astrocytes.

GFAP, ALDH1L1, and S100β are popular markers of astrocytes. Of the three, ALDH1L1 is the most astrocyte-specific.

Additional astrocyte markers with notable functional significance include the structural protein vimentin (VIM), which helps form the astrocyte morphology; the biosynthetic enzyme glycogen synthase (GS, encoded by GYS1), which plays a role in synaptic plasticity; the water channel AQP4, which maintains water homeostasis in the CNS; the sodium-dependent glutamate transporters EAAT1 and EAAT2 (GLAST and GLT1 in rodents), which facilitate metabolite shuttling between astrocytes and neurons; and the gap junction protein connexin 43 (Cx43, encoded by GJA1), which forms part of the intercellular networks astrocytes use to regulate synaptic plasticity and support neuronal metabolism, among other functions. Of note, AQP4 is also expressed by ependymal cells, underlying the participation of both cell types in regulating water homeostasis.

For a more detailed overview of astrocyte markers, read our guide to astrocyte markers. 

Microglia Markers

Microglia are the resident tissue macrophage of the CNS. As such, they play important roles in maintaining tissue homeostasis (via synaptic pruning, for example) and defense. Morphologically, they can be identified by their small cell bodies and highly ramified processes.

As macrophages, microglia can be identified using nonspecific macrophage markers such as CD11b (ITGAM), CD45, CX3CR1, and Iba1 (AIF1). These markers enable identification and/or isolation of microglia via flow cytometry (CD45^lo/CD11b), immunohistochemistry (Iba1), or the fluorescent reporter transgenic mice that are widely used for in vivo microscopic studies (namely, CX3CR1-GFP mice). PU.1 (SPI1) and MAFB are nonspecific transcriptional regulators of macrophage identity, and CD68 is a lysosomal glycoprotein shared by all macrophages (as phagocytes).

Several largely microglia-specific markers have been identified in the past decade. These markers include TMEM119, P2RY12, HEXB, and SALL1. TMEM119 is the most specific to microglia of the four markers, labeling mature microglia as early as postnatal day 14 in the mouse.

Microglia alter their marker profile upon activation. Markers typically expressed at high levels under steady-state conditions—TMEM119, P2RY12, HEXB, and SALL1—are downregulated, whereas many nonspecific macrophage markers are upregulated, including CD45, Iba1, and CD68. Morphologically, activated microglia take on a more amoeboid shape, retracting and thickening their once thin processes while enlarging their cell body. Activated microglia can be difficult to distinguish from other brain-resident macrophages as well as infiltrating macrophages. TMEM119 represents one of the few markers that allow these cell types to be distinguished from one another.

In some contexts, microglia undergo more specific activation programs. One such phenotype is called disease-associated microglia (DAM), which has been observed in Alzheimer’s disease and other neurodegenerative contexts. DAMs downregulate homeostatic microglial markers (P2RY12, etc.) and upregulate TREM2 and APOE. Astrocytes can also express APOE, limiting the usefulness of this marker for discrimination between microglia and astrocytes when used alone.

For more discussion of microglial markers, check out our guide to microglial markers. 

Ependymal Cell Markers

Ependymal cells are a rare glial cell type that populates the brain’s ventricle walls. These cells are morphologically distinguished by ventricle-facing cilia, which help to direct cerebrospinal fluid flow and the distribution of nutrients and neurotransmitters to neurons. As a transcriptional regulator of motile cilia generation, FOXJ1 marks ependymal cells, although it also marks a subset of neural stem cells. Acetylated ɑ-tubulin (TUBA1A), as a cilia protein, also marks ependymal cells.

Similarities between ependymal cells and neural stem cells, such as their shared location in a neurogenic niche and expression of several markers (SOX2, Nestin, and CD133) have prompted debate over the neurogenic potential of ependymal cells. However, recent lineage tracing via ɑ-SMA (ACTA2) suggests that the neurogenic potential of ependymal cells detected by others was a consequence of neural stem cell contamination.

Additional markers of ependymal cells, identified by RNA-sequencing, include RARRES2, CCDC153, TMEM212, TM4SF1, and MIA.

Table of Glial Cell Markers

The table below lists characteristic glial cell proteins as described by review literature. The list includes a variety of marker types, including transcription factors, membrane proteins, secreted factors, signaling proteins, and structural proteins. Accompanying each marker are links to relevant antibodies and ELISA kits that can be used to detect glia in vitro and in vivo. The associated products are offered by a variety of manufacturers and can serve as a useful reference for glial cell characterization.

Note: Some markers are lipids* or antibodies**. Information on Protein Type, Localization, and Size (kDa) obtained from UniProt.org (for human genes only). 

References

1. Redwine JM, Evans CF. Markers of central nervous system glia and neurons in vivo during normal and pathological conditions. Curr Top Microbiol Immunol. 2002;265:119-140. doi:10.1007/978-3-662-09525-6_6

2. Goldman SA, Kuypers NJ. How to make an oligodendrocyte. Development. 2015;142(23):3983-3995. doi:10.1242/dev.126409

3. Shah PT, Stratton JA, Stykel MG, et al. Single-Cell Transcriptomics and Fate Mapping of Ependymal Cells Reveals an Absence of Neural Stem Cell Function. Cell. 2018;173(4):1045-1057.e9. doi:10.1016/j.cell.2018.03.063

4. Low D, Ginhoux F. Recent advances in the understanding of microglial development and homeostasis. Cell Immunol. 2018;330:68-78. doi:10.1016/j.cellimm.2018.01.004

5. Jurga AM, Paleczna M, Kuter KZ. Overview of General and Discriminating Markers of Differential Microglia Phenotypes. Front Cell Neurosci. 2020;14:198. Published 2020 Aug 6. doi:10.3389/fncel.2020.00198

6. Jurga AM, Paleczna M, Kadluczka J, Kuter KZ. Beyond the GFAP-Astrocyte Protein Markers in the Brain. Biomolecules. 2021;11(9):1361. Published 2021 Sep 14. doi:10.3390/biom11091361