Isolation and Culture of Adult Human Microglia Within Mixed Glial Cultures for Functional Experimentation and High-Content Analysis
Abstract
Microglia are thought to be involved in diseases of the adult human brain as well as normal aging processes. While neonatal and rodent microglia are often used in studies investigating microglial function, there are important differences between rodent microglia and their adult human counterparts. Human brain tissue provides a unique and valuable tool for microglial cell and molecular biology. Routine protocols can now enable use of this culture method in many laboratories. Detailed protocols and advice for culture of human brain microglia are provided here. We demonstrate the protocol for culturing human adult microglia within a mixed glial culture and use a phagocytosis assay as an example of the functional studies possible with these cells as well as a high-content analysis method of quantification.
Integrity signals
Registry
Cites
Statistics
Claims the paper makes
Claims other papers make about this one
Each is checked against this paper’s own text: does it say what the citing paper says it does?
Primary human microglia are poorly characterized, differ greatly from mature mouse microglia, are not easily accessible, and current isolation methods likely alter their defining characteristics.
“primary human microglia remain poorly characterized as they differ greatly from mature mouse microglia, are not easily accessible and current methods employed for isolation likely alter their defining characteristics6.”
From Efficient derivation of microglia-like cells from human pluripotent stem cells (2016) · cites this paper as [6]
Cites
Microglia: Active sensor and versatile effector cells in the normal and pathologic brain
Hanisch, U.-K., & Kettenmann, H. (2007) · Nature Neuroscience · doi:10.1038/nn1997 · Cited by 3,676 · see citing works
Microglial senescence: does the brain's immune system have an expiration date?
Streit, W. J. (2006) · Trends in Neurosciences · doi:10.1016/j.tins.2006.07.001 · Cited by 397 · see citing works
The adult human brain in preclinical drug development
Dragunow, M. (2008) · Nature Reviews Drug Discovery · doi:10.1038/nrd2617 · Cited by 120 · see citing works
Self-citationThe impact of glial activation in the aging brain.
Lynch, A. M., Murphy, K. J., Deighan, B. F., O’Reilly, J.-A., Gun’ko, Y. K., Cowley, T. R., González-Reyes, R. E., & Lynch, M. A. (2010) · Europe PMC (PubMed Central) · Cited by 103 · see citing works
High-content analysis in neuroscience
Dragunow, M. (2008) · Nature reviews. Neuroscience · doi:10.1038/nrn2492 · Cited by 87 · see citing works
Self-citationAdult human brain cell culture for neuroscience research.
Gibbons, H., & Dragunow, M. (2010) · International Journal of Biochemistry and Cell Biology · doi:10.1016/j.biocel.2009.12.002 · Cited by 60 · see citing works
Self-citationCellular composition of human glial cultures from adult biopsy brain tissue
Gibbons, H. M., Hughes, S. M., van Roon‐Mom, W., Greenwood, J. M., Narayan, P. J., Teoh, H. H., Bergin, P., Mee, E. W., Wood, P. C., Faull, R. L. M., et al. (2007) · Journal of Neuroscience Methods · doi:10.1016/j.jneumeth.2007.07.005 · Cited by 55 · see citing works
Self-citationValproic acid enhances microglial phagocytosis of amyloid-β1–42
Smith, A. M., Gibbons, H. M., & Dragunow, M. (2010) · Neuroscience · doi:10.1016/j.neuroscience.2010.04.041 · Cited by 53 · see citing works
Self-citationValproic acid induces microglial dysfunction, not apoptosis, in human glial cultures
Gibbons, H. M., Smith, A. M., Teoh, H. H., Bergin, P. M., Mee, E. W., Faull, R. L. M., & Dragunow, M. (2011) · Neurobiology of Disease · doi:10.1016/j.nbd.2010.08.024 · Cited by 49 · see citing works
Self-citationDavis, M. M. (2012) · Science Translational Medicine · doi:10.1126/scitranslmed.3003385 · Cited by 35 · see citing works
High throughput quantification of cells with complex morphology in mixed cultures
Narayan, P. J., Gibbons, H. M., Mee, E. W., Faull, R. L. M., & Dragunow, M. (2007) · Journal of Neuroscience Methods · doi:10.1016/j.jneumeth.2007.04.016 · Cited by 27 · see citing works
Self-citation
Flagged first, then by citations
Cited by
Sorted by citations
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
Smith, A. M., & Dragunow, M. (2014) · Trends in Neurosciences · doi:10.1016/j.tins.2013.12.001 · Cited by 256 · see citing works
Self-citationMIF-CD74 signaling impedes microglial M1 polarization and facilitates brain tumorigenesis
Ghoochani, A., Schwarz, M. A., Yakubov, E., Engelhorn, T., Doerfler, A., Buchfelder, M., Bucala, R., Savaskan, N. E., & Eyüpoglu, I. Y. (2016) · Oncogene · doi:10.1038/onc.2016.160 · Cited by 171 · see citing works
Brain microglia serve as a persistent HIV reservoir despite durable antiretroviral therapy
Tang, Y., Chaillon, A., Gianella, S., Wong, L. M., Li, D., Simermeyer, T. L., Porrachia, M., Ignacio, C. C., Woodworth, B., Zhong, D., et al. (2023) · Journal of Clinical Investigation · doi:10.1172/jci167417 · Cited by 165 · see citing works
Wang, X., Wang, C., Wang, J., Zhao, S., Zhang, K., Wang, J., Zhang, W., Wu, C., & Yang, J. (2014) · Neuropharmacology · doi:10.1016/j.neuropharm.2014.01.022 · Cited by 122 · 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
Self-citationChoi, J., Stradmann‐Bellinghausen, B., Yakubov, E., Savaskan, N. E., & Régnier‐Vigouroux, A. (2015) · Cancer Biology & Therapy · doi:10.1080/15384047.2015.1056406 · Cited by 90 · see citing works
Xu, W., Wu, Y., Hu, Z., Sun, L., Dou, G., Zhang, Z., Wang, H., Guo, C., & Wang, Y. (2019) · Molecular Therapy — Nucleic Acids · doi:10.1016/j.omtn.2019.04.029 · Cited by 65 · see citing works
Park, T., Feisst, V., Brooks, A. E. S., Rustenhoven, J., Monzó, H. J., Feng, S. X., Mee, E. W., Bergin, P., Oldfield, R. L., Graham, E. S., et al. (2016) · Scientific Reports · doi:10.1038/srep26587 · Cited by 47 · see citing works
Self-citationXu, M. F., Zhang, L., Liu, G., Jiang, N., Zhou, W., & Zhang, Y. (2018) · Journal of Alzheimer s Disease · doi:10.3233/jad-180722 · Cited by 40 · see citing works
Smith, A. M., Graham, E. S., Feng, S. X., Oldfield, R. L., Bergin, P. M., Mee, E. W., Faull, R. L. M., Curtis, M. A., & Dragunow, M. (2013) · PLoS ONE · doi:10.1371/journal.pone.0080463 · Cited by 38 · see citing works
Self-citationRoutine culture and study of adult human brain cells from neurosurgical specimens
Park, T., Smyth, L. C. D., Aalderink, M., Woolf, Z., Rustenhoven, J., Lee, K., Jansson, D., Smith, A. M., Feng, S. X., Correia, J. A., et al. (2022) · Nature Protocols · doi:10.1038/s41596-021-00637-8 · Cited by 38 · see citing works
Self-citationAlzheimer’s Disease Research Using Human Microglia
Lue, L., Beach, T. G., & Walker, D. G. (2019) · Cells · doi:10.3390/cells8080838 · Cited by 26 · see citing works
Physiology of Cultured Human Microglia Maintained in a Defined Culture Medium
Tewari, M., Khan, M., Verma, M., Coppens, J. R., Kemp, J. M., Bucholz, R. D., Mercier, P., & Egan, T. M. (2021) · ImmunoHorizons · doi:10.4049/immunohorizons.2000101 · Cited by 16 · see citing works
Smith, A. M., Park, T., Aalderink, M., Oldfield, R. L., Bergin, P., Mee, E. W., Faull, R. L. M., & Dragunow, M. (2022) · Frontiers in Cellular Neuroscience · doi:10.3389/fncel.2022.1047928 · Cited by 9 · see citing works
Self-citationBlenkle, A., Geladaris, A., & Weber, M. S. (2026) · Experimental & Molecular Medicine · doi:10.1038/s12276-026-01647-w · Cited by 6 · see citing works
Darzenta, N., Davis, E., Cochran, A. M., Murray, M., Moore, O., & Naskou, M. C. (2025) · Stem Cells and Development · doi:10.1089/scd.2025.0042 · Cited by 2 · see citing works
FONTES, G. C., Hayashide, L. D. S., Messor, D. F., Marques, M., Leocadio, V. E., Amorim, P., Minho, A. S., Paiva-Pereira, E., Savio, L. E. B., Fernandes, P. D., et al. (2026) · Neurochemical Research · doi:10.1007/s11064-025-04647-w · Cited by 3 · see citing works
- Authors
- Smith, A. M., Gibbons, H. M., Lill, C., Faull, R. L. M., & Dragunow, M.
- Affiliations
- University of Auckland (NZ)
- Venue
- Methods in Molecular Biology
- Publisher
- Humana Press
- Published
- 2013
- Volume
- 1041
- Pages
- 41-51
- ISSN
- 1064-3745 (ISSN-L)
- 1940-6029
- OpenAlex
- W2397793632
- Cited by
- 18
- References
- 11
- Citations by year
- 2026: 2 · 2025: 1 · 2023: 1 · 2022: 2 · 2021: 1 · 2019: 2 · 2018: 1 · 2016: 4 · 2015: 1 · 2014: 2 · 2013: 1
- FWCI
- 6.18
- Type
- book chapter
- Language
- en
- Open access
- closed
- Licence
- Locations
- Methods in molecular biology (publishedVersion)
- PubMed (publishedVersion)
- Funders
- National Research Centre
- Health Research Council of New Zealand
- Topics
- Life Sciences › Neuroscience › Neurology › Neuroinflammation and Neurodegeneration Mechanisms
- Life Sciences › Neuroscience › Cellular and Molecular Neuroscience › Neuroscience and Neuropharmacology Research
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Inflammasome and immune disorders
- Keywords
- human microglia, microglia isolation, mixed glial cultures, adult human brain, phagocytosis assay, high-content analysis, microglial function, microglial aging
- MeSH terms
- Brain, Cell Separation, Cells, Cultured, Humans, Neuroglia, Microglia
About these checks
Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.
Statistics and arithmetic are recomputed, not judged.
Confidence bands are shown only for fields calibrated on our annotated test set. None is calibrated yet, so no band is shown.
What counts as a claim ›How we measure confidence ›How we check this ›
Are you an author, or do you see a problem with this page? Request a review
