Isolation and Phenotyping of Adult Mouse Microglial Cells
Open accessAbstract
Microglia are the resident macrophages of the central nervous system parenchyma and fulfill crucial roles in brain development, homeostasis, and inflammation. The isolation of a pure microglia population from brain tissue enables the examination of microglial phenotypes without the interference of other cell populations. Microglial extractions from the neonatal brain have been described in various protocols, yet the more established and complex adult mouse brain poses a greater challenge. Here we describe a refined protocol including enzymatic and mechanical dissociation of adult mouse brain tissue and removal of myelin by Percoll density gradient. Microglial cells were subsequently extracted by an immunomagnetic approach. This isolation procedure enables the use of functionally viable cells for various applications such as cell culture, flow cytometry, functional assays including bacteria- or bead-based phagocytosis, stimulation assays, and transcriptome profiling techniques such as qRT-PCR and microarray/RNA sequencing.
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Many widely used protocols for microglial isolation continue to include enzymatic digestion, likely because some experimental conditions require greater yield.
“Some experimental conditions may dictate the need for greater yield, especially for older or less-sensitive approaches (including many older bulk RNA-sequencing (RNA-seq) studies or protein-based analyses), which is likely why many widely used and widely cited protocols for microglial isolation continue to include enzymatic digestion4,63,64.”
From Dissection of artifactual and confounding glial signatures by single-cell sequencing of mouse and human brain (2022) · cites this paper as [63]
The Introduction describes a microglial-isolation protocol that uses enzymatic dissociation, and Notes (Note 3) reports that digestion improved microglial yield. Our copy does not establish that many widely used protocols include digestion, or that a need for greater yield explains their use of it, particularly for older RNA-seq or protein-based analyses. (LLM, conf=0.83)
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Cites
A Unique Microglia Type Associated with Restricting Development of Alzheimer’s Disease
Keren-Shaul, H., Spinrad, A., Weiner, A., Matcovitch-Natan, O., Dvir-Szternfeld, R., Ulland, T. K., David, E., Baruch, K., Lara-Astaiso, D., Toth, B., et al. (2017) · Cell · doi:10.1016/j.cell.2017.05.018 · Cited by 5,493 · see citing works
Fate Mapping Analysis Reveals That Adult Microglia Derive from Primitive Macrophages
Ginhoux, F., Greter, M., Leboeuf, M., Nandi, S., See, P., Gokhan, S., Mehler, M. F., Conway, S. J., Ng, L. G., Stanley, E. R., et al. (2010) · Science · doi:10.1126/science.1194637 · Cited by 5,175 · see citing works
Synaptic Pruning by Microglia Is Necessary for Normal Brain Development
Paolicelli, R. C., Bolasco, G., Pagani, F., Maggi, L., Scianni, M., Panzanelli, P., Giustetto, M., Ferreira, T. A., Guiducci, E., Dumas, L., et al. (2011) · Science · doi:10.1126/science.1202529 · Cited by 4,244 · see citing works
Microglia Sculpt Postnatal Neural Circuits in an Activity and Complement-Dependent Manner
Schafer, D. P., Lehrman, E. K., Kautzman, A. G., Koyama, R., Mardinly, A. R., Yamasaki, R., Ransohoff, R. M., Greenberg, M. E., Barres, B. A., & Stevens, B. (2012) · Neuron · doi:10.1016/j.neuron.2012.03.026 · Cited by 4,089 · see citing works
Kettenmann, H., Hanisch, U.-K., Noda, M., & Verkhratsky, A. (2011) · Physiological Reviews · doi:10.1152/physrev.00011.2010 · Cited by 3,547 · see citing works
Microglia Function in the Central Nervous System During Health and Neurodegeneration
Colonna, M., & Butovsky, O. (2017) · Annual Review of Immunology · doi:10.1146/annurev-immunol-051116-052358 · Cited by 2,894 · see citing works
Identification of a unique TGF-β–dependent molecular and functional signature in microglia
Butovsky, O., Jedrychowski, M. P., Moore, C. S., Cialic, R., Lanser, A. J., Gabriely, G., Koeglsperger, T., Dake, B., Wu, P. M., Doykan, C. E., et al. (2014) · Nature Neuroscience · doi:10.1038/nn.3599 · Cited by 2,543 · see citing works
Heterogeneity in the distribution and morphology of microglia in the normal adult mouse brain
Lawson, L. J., Perry, V. H., Dri, P., & Gordon, S. (1990) · Neuroscience · doi:10.1016/0306-4522(90)90229-w · Cited by 2,045 · see citing works
Microglial physiology: unique stimuli, specialized responses
Ransohoff, R. M., & Perry, V. H. (2009) · Annual Review of Immunology · doi:10.1146/annurev.immunol.021908.132528 · Cited by 1,858 · see citing works
New tools for studying microglia in the mouse and human CNS
Bennett, M. L., Bennett, F. C., Liddelow, S. A., Ajami, B., Zamanian, J. L., Fernhoff, N. B., Mulinyawe, S. B., Bohlen, C. J., Adil, A., Tucker, A., et al. (2016) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.1525528113 · Cited by 1,804 · see citing works
Wake, H., Moorhouse, A. J., Jinno, S., Kohsaka, S., & Nabekura, J. (2009) · The Journal of Neuroscience · doi:10.1523/jneurosci.4363-08.2009 · Cited by 1,619 · see citing works
An environment-dependent transcriptional network specifies human microglia identity
Gosselin, D., Skola, D., Coufal, N. G., Holtman, I. R., Schlachetzki, J. C. M., Sajti, E., Jaeger, B. N., O’Connor, C., Fitzpatrick, C., Pasillas, M. P., et al. (2017) · Science · doi:10.1126/science.aal3222 · Cited by 1,220 · see citing works
iPSC-derived human microglia-like cells to study neurological diseases
Abud, E. M., Ramirez, R. N., Martinez, E. S., Healy, L. M., Nguyen, C. H. H., Newman, S. A., Yeromin, A. V., Scarfone, V. M., Marsh, S. E., Fimbres, C., et al. (2017) · Neuron · doi:10.1016/j.neuron.2017.03.042 · Cited by 1,154 · see citing works
Microglial brain region-dependent diversity and selective regional sensitivities to aging
Grabert, K., Michoel, T., Karavolos, M. H., Clohisey, S., Baillie, J. K., Stevens, M. P., Freeman, T. C., Summers, K. M., & McColl, B. W. (2016) · Nature Neuroscience · doi:10.1038/nn.4222 · Cited by 1,151 · see citing works
Self-citationDebris clearance by microglia: an essential link between degeneration and regeneration
Neumann, H., Kotter, M. R., & Franklin, R. J. M. (2008) · Brain · doi:10.1093/brain/awn109 · Cited by 1,111 · see citing works
The Role of Microglia in the Healthy Brain: Figure 1.
Tremblay, M.-È., Stevens, B., Sierra, A., Wake, H., Bessis, A., & Nimmerjahn, A. (2011) · The Journal of Neuroscience · doi:10.1523/jneurosci.4158-11.2011 · Cited by 992 · see citing works
Microglia as neuroprotective, immunocompetent cells of the CNS
Streit, W. J. (2002) · Glia · doi:10.1002/glia.10154 · Cited by 919 · see citing works
Bohlen, C. J., Bennett, F. C., Tucker, A. F., Collins, H. Y., Mulinyawe, S. B., & Barres, B. A. (2017) · Neuron · doi:10.1016/j.neuron.2017.04.043 · Cited by 676 · see citing works
Efficient derivation of microglia-like cells from human pluripotent stem cells
Muffat, J., Li, Y., Yuan, B., Mitalipova, M., Omer, A., Corcoran, S., Bakiasi, G., Tsai, L.-H., Aubourg, P., Ransohoff, R. M., et al. (2016) · Nature Medicine · doi:10.1038/nm.4189 · Cited by 651 · see citing works
Ford, A. L., Goodsall, A. L., Hickey, W. F., & Sedgwick, J. D. (1995) · The Journal of Immunology · doi:10.4049/jimmunol.154.9.4309 · Cited by 631 · see citing works
Szalay, G., Martinecz, B., Lénárt, N., Környei, Z., Orsolits, B., Judák, L., Császár, E., Fekete, R., West, B. L., Katona, G., et al. (2016) · Nature Communications · doi:10.1038/ncomms11499 · Cited by 573 · see citing works
TMEM119 marks a subset of microglia in the human brain
Satoh, J., Kino, Y., Asahina, N., Takitani, M., Miyoshi, J., Ishida, T., & Saito, Y. (2016) · Neuropathology · doi:10.1111/neup.12235 · Cited by 434 · see citing works
Microglial displacement of inhibitory synapses provides neuroprotection in the adult brain
Chen, Z., Jalabi, W., Hu, W., Park, H.-J., Gale, J. T., Kidd, G. J., Bernatowicz, R., Gossman, Z. C., Chen, J. T., Dutta, R., et al. (2014) · Nature Communications · doi:10.1038/ncomms5486 · Cited by 319 · see citing works
Microglia function during brain development: New insights from animal models
Bilimoria, P. M., & Stevens, B. (2015) · Brain Research · doi:10.1016/j.brainres.2014.11.032 · Cited by 204 · see citing works
Frank, M. G., Wieseler‐Frank, J., Watkins, L. R., & Maier, S. F. (2005) · Journal of Neuroscience Methods · doi:10.1016/j.jneumeth.2005.06.026 · Cited by 164 · see citing works
Targeting innate immunity for neurodegenerative disorders of the central nervous system
Andreasson, K. I., Bachstetter, A. D., Colonna, M., Ginhoux, F., Holmes, C., Lamb, B., Landreth, G., Lee, D. C., Low, D., Lynch, M. A., et al. (2016) · Journal of Neurochemistry · doi:10.1111/jnc.13667 · Cited by 126 · see citing works
Unique transcriptome signature of mouse microglia
Beutner, C., Linnartz‐Gerlach, B., Schmidt, S. V., Beyer, M., Mallmann, M. R., Staratschek‐Jox, A., Schultze, J. L., & Neumann, H. (2013) · Glia · doi:10.1002/glia.22524 · Cited by 100 · see citing works
Isolation of highly enriched primary human microglia for functional studies
Rustenhoven, J., Park, T. I.-H., Schweder, P., Scotter, J., Correia, J., Smith, A. M., Gibbons, H. M., Oldfield, R. L., Bergin, P. S., Mee, E. W., et al. (2016) · Scientific Reports · doi:10.1038/srep19371 · Cited by 99 · see citing works
Marek, R. A., Caruso, M. L., Rostami, A. M., Grinspan, J. B., & Sarma, J. D. (2008) · Journal of Neuroscience Methods · doi:10.1016/j.jneumeth.2008.08.016 · Cited by 83 · see citing works
An optimized protocol for the acute isolation of human microglia from autopsy brain samples
Olah, M., Raj, D. D. A., Brouwer, N., Haas, A. H. D., Eggen, B. J. L., den Dunnen, W. F. A., Biber, K. P. H., & Boddeke, H. W. G. M. (2011) · Glia · doi:10.1002/glia.21251 · Cited by 80 · see citing works
Harms, A. S., & Tansey, M. G. (2013) · Methods in molecular biology · doi:10.1007/978-1-62703-520-0_5 · Cited by 26 · see citing works
Daniele, S. G., Edwards, A. A., & Maguire‐Zeiss, K. A. (2014) · Journal of Visualized Experiments · doi:10.3791/51005 · Cited by 19 · see citing works
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Marsh, S. E., Walker, A. J., Kamath, T., Dissing-Olesen, L., Hammond, T. R., de Soysa, T. Y., Young, A. M. H., Murphy, S., Abdulraouf, A., Nadaf, N., et al. (2022) · Nature Neuroscience · doi:10.1038/s41593-022-01022-8 · Cited by 432 · see citing works
Colombo, G., Cubero, R. J. A., Kanari, L., Venturino, A., Schulz, R., Scolamiero, M., Agerberg, J., Mathys, H., Tsai, L.-H., Chachólski, W., et al. (2022) · Nature Neuroscience · doi:10.1038/s41593-022-01167-6 · Cited by 100 · see citing works
- Authors
- Grabert, K., & McColl, B. W.
- Affiliations
- Roslin Institute (GB)
- Corresponding author
- Grabert, Kathleen
- Venue
- Methods in Molecular Biology
- Publisher
- Springer New York
- Published
- 2018
- Volume
- 1784
- Pages
- 77-86
- ISSN
- 1064-3745 (ISSN-L)
- 1940-6029
- OpenAlex
- W2804367279
- Cited by
- 48
- References
- 32
- Citations by year
- 2026: 2 · 2025: 2 · 2024: 7 · 2023: 13 · 2022: 9 · 2021: 11 · 2020: 3 · 2018: 1
- FWCI
- 8.46
- Type
- book chapter
- Language
- en
- Open access
- hybrid
- doi.org
- Licence
- Locations
- Methods in molecular biology (publishedVersion, cc-by)
- PubMed (publishedVersion)
- Edinburgh Research Explorer (University of Edinburgh) (submittedVersion, cc-by)
- Funders
- Directorate for Biological Sciences
- Medical Research Council (MR/L003384/1, MR/K001744/1)
- Biotechnology and Biological Sciences Research Council (BBS/E/D/10002071, BB/J4243/1, BB /J004332/1)
- Topics
- Life Sciences › Neuroscience › Neurology › Neuroinflammation and Neurodegeneration Mechanisms
- Life Sciences › Neuroscience › Developmental Neuroscience › Neurogenesis and neuroplasticity mechanisms
- Health Sciences › Medicine › Pathology and Forensic Medicine › Multiple Sclerosis Research Studies
- Keywords
- microglia, adult mouse brain, Percoll density gradient, magnetic cell separation, phagocytosis
- MeSH terms
- Animals, Brain, Cell Separation, Flow Cytometry, Myelin Sheath, Phagocytosis, Povidone, Silicon Dioxide, Microglia, Cell Culture Techniques, Mice
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