A refined rat primary neonatal microglial culture method that reduces time, cost and animal use

Marieta Georgieva, Alasdair Leeson-Payne, Maria Dumitrascuta, Ann Rajnicek, Marzia Malcangio, Wenlong Huang

Research output: Contribution to journalArticle

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Abstract

Background: Primary microglial cultures have been used extensively to facilitate the development of therapeutic strategies for a variety of CNS disorders including neurodegeneration and neuropathic pain. However, existing techniques for culturing these cells are slow and costly. 
New Method: Here, we report a refined protocol based on our previously published methods described by Clark et al., which reduces in the time, reagents and the number of animals used for each culture whilst yielding high number and excellent quality microglial cells.
Results: Our refined protocol offers an isolation of > 96% microglia from a mixed glial culture after only four days of incubation. It results in a high yield of microglia, in excess of one million cells per cortex with predominantly resting morphology and a low level of cell activation.
Comparison with Existing Method(s): Compared to conventional procedures our refined protocol requires only one third of the time to prepare high quality microglial cultures, cuts the cost more than four-fold, and significantly reduces the number of animals used per culture.
Conclusion: Our consistent, reliable, and time/cost effective microglial culture protocol is crucial for efficient in vitro screening of potential therapeutics. By dramatically reducing the culture time from 2 weeks to just 4 days and increasing the laboratory research output it has implications for the Reduction, Refinement and Replacement policies endorsed by many government funding agencies and animal research regulatory bodies.
Original languageEnglish
Pages (from-to)92-102
Number of pages11
JournalJournal of Neuroscience Methods
Volume304
Early online date27 Apr 2018
DOIs
Publication statusPublished - Jul 2018

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Costs and Cost Analysis
Microglia
Government Agencies
Neuralgia
Neuroglia
Therapeutics
Research
In Vitro Techniques

Keywords

  • cell culture
  • microglia
  • tissue culture dish
  • neonatal rats
  • cortex
  • in vitro

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A refined rat primary neonatal microglial culture method that reduces time, cost and animal use. / Georgieva, Marieta; Leeson-Payne, Alasdair; Dumitrascuta, Maria; Rajnicek, Ann; Malcangio, Marzia; Huang, Wenlong.

In: Journal of Neuroscience Methods, Vol. 304, 07.2018, p. 92-102.

Research output: Contribution to journalArticle

Georgieva, Marieta ; Leeson-Payne, Alasdair ; Dumitrascuta, Maria ; Rajnicek, Ann ; Malcangio, Marzia ; Huang, Wenlong. / A refined rat primary neonatal microglial culture method that reduces time, cost and animal use. In: Journal of Neuroscience Methods. 2018 ; Vol. 304. pp. 92-102.
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AU - Malcangio, Marzia

AU - Huang, Wenlong

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N2 - Background: Primary microglial cultures have been used extensively to facilitate the development of therapeutic strategies for a variety of CNS disorders including neurodegeneration and neuropathic pain. However, existing techniques for culturing these cells are slow and costly. New Method: Here, we report a refined protocol based on our previously published methods described by Clark et al., which reduces in the time, reagents and the number of animals used for each culture whilst yielding high number and excellent quality microglial cells.Results: Our refined protocol offers an isolation of > 96% microglia from a mixed glial culture after only four days of incubation. It results in a high yield of microglia, in excess of one million cells per cortex with predominantly resting morphology and a low level of cell activation.Comparison with Existing Method(s): Compared to conventional procedures our refined protocol requires only one third of the time to prepare high quality microglial cultures, cuts the cost more than four-fold, and significantly reduces the number of animals used per culture.Conclusion: Our consistent, reliable, and time/cost effective microglial culture protocol is crucial for efficient in vitro screening of potential therapeutics. By dramatically reducing the culture time from 2 weeks to just 4 days and increasing the laboratory research output it has implications for the Reduction, Refinement and Replacement policies endorsed by many government funding agencies and animal research regulatory bodies.

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