A simple magnetic separation method for high-yield isolation of pure primary microglia

Gordon, Richard, Hogan, Colleen E., Neal, Matthew L., Anantharam, Vellareddy, Kanthasamy, Anumantha G. and Kanthasamy, Arthi (2011) A simple magnetic separation method for high-yield isolation of pure primary microglia. Journal of Neuroscience Methods, 194 2: 287-296. doi:10.1016/j.jneumeth.2010.11.001

Author Gordon, Richard
Hogan, Colleen E.
Neal, Matthew L.
Anantharam, Vellareddy
Kanthasamy, Anumantha G.
Kanthasamy, Arthi
Title A simple magnetic separation method for high-yield isolation of pure primary microglia
Journal name Journal of Neuroscience Methods   Check publisher's open access policy
ISSN 0165-0270
Publication date 2011-01-15
Year available 2011
Sub-type Article (original research)
DOI 10.1016/j.jneumeth.2010.11.001
Open Access Status Not yet assessed
Volume 194
Issue 2
Start page 287
End page 296
Total pages 10
Place of publication Amsterdam, Netherlands
Publisher Elsevier
Language eng
Formatted abstract
Microglial cells play a dynamic role in the brain beyond their established function of immune surveillance. Activated microglia play key roles in neural development, neuroinflammation, neural repair and neurotoxicity. They are particularly important in several neurodegenerative diseases in which sustained microglial activation contributes to the progression of neurodegenerative processes. Consequently, understanding microglial function in CNS health and disease has become an area of active research in recent years. However, a significant obstacle to progress in this field has been the inherent difficulties in obtaining large amounts of primary microglial cells to routinely perform mechanistic studies and characterize signaling pathways regulating the dynamics of microglial activation. Herein, we describe a novel column-free magnetic separation protocol for high-yield isolation of primary microglia from mouse postnatal mixed glial cultures. The procedure is based on optimized culture conditions that enable high microglial cell densities in confluent mixed glial cultures followed by highly efficient recovery of pure microglia by magnetic separation. The novel column-free magnetic separation system utilizes tetrameric antibody complexes (TAC) with dual specificity for CD11b-PE labeled microglia and dextran magnetic nanoparticles. An FcR blocker (anti-CD16/32) is added to enhance the purity of the microglial separation by preventing non-specific labeling of other cell types. This procedure yields on average >3×106 microglial cells per mouse pup, with a remarkable purity of 97% and recovery of around 87% of microglia from the mixed glial population. Importantly, the microglia obtained by this method are fully functional and respond like cells obtained by conventional isolation techniques.
Keyword Cell culture
Magnetic separation
Microglial activation
Q-Index Code C1
Q-Index Status Provisional Code
Grant ID NS 065167
Institutional Status Non-UQ
Additional Notes Published online 11 November 2010

Document type: Journal Article
Sub-type: Article (original research)
Collection: School of Biomedical Sciences Publications
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Citation counts: TR Web of Science Citation Count  Cited 34 times in Thomson Reuters Web of Science Article | Citations
Scopus Citation Count Cited 33 times in Scopus Article | Citations
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Created: Thu, 01 May 2014, 09:39:49 EST by Richard Gordon on behalf of School of Biomedical Sciences