Related Experiment Video
Updated: Aug 29, 2025
Isolation of Cortical Microglia with Preserved Immunophenotype and Functionality From Murine Neonates
Published on: January 30, 2014
A Step-by-step Protocol for Obtaining Mature Microglia from Mice
1Department of Pharmacology, Research Institute for Basic Medical Science, School of Medicine, CHA University, CHA BIO COMPLEX, 335 Pangyo, Bundang-gu, Seongnam-si, Gyeonggi-do 13488, Republic of Korea.
Abstract:
In mice, microglial precursors in the yolk sac migrate to the brain parenchyma through the head neuroepithelial layer between embryonic days 8.5 (E8.5)-E16.5 and acquire their unique identity with a ramified form. Based on the microglial developmental process, we dissected the neuroepithelial layer (NEL) of E13.5 mice, which is composed of microglial progenitor and neuroepithelial cells. The NEL was bankable and expandable. In addition, microglial precursors were matured according to NEL culture duration. The matured microglia (MG; CD11b-positive cells) were easily isolated from the cultured NEL using a magnetic-activated cell sorting system and named NEL-MG. In conclusion, we obtained higher yields of adult-like microglia (mature microglia: NEL-MG) compared to previous in vitro surrogates such as neonatal microglia and microglial cell lines. Graphical abstract.
Insights
Researchers developed a novel method to culture adult-like microglia (NEL-MG) from mouse neuroepithelial layers. This technique provides higher yields than existing in vitro models for studying brain immune cells.
Area of Science:
- Developmental neuroscience
- Immunology
- Stem cell biology
Background:
- Microglia, the brain's resident immune cells, originate from yolk sac precursors.
- These precursors migrate to the brain and differentiate within the neuroepithelial layer (NEL) during embryonic development.
- Existing in vitro models for microglia research have limitations in yield and maturity.
Purpose of the Study:
- To establish a novel in vitro system for generating mature microglia.
- To utilize the neuroepithelial layer (NEL) as a source for microglial precursors.
- To compare the yield and characteristics of derived microglia with existing models.
Main Methods:
- Dissection of the embryonic mouse neuroepithelial layer (NEL) at E13.5.
- Culture and expansion of NEL cells to promote microglial precursor maturation.
- Isolation of mature microglia (NEL-MG) using magnetic-activated cell sorting (MACS) for CD11b-positive cells.
Main Results:
- The NEL was found to be bankable and expandable, supporting cell culture.
- Microglial precursors within the NEL matured with extended culture duration.
- Higher yields of adult-like microglia (NEL-MG) were obtained compared to neonatal microglia and cell lines.
Conclusions:
- The neuroepithelial layer (NEL) serves as a viable source for generating mature microglia in vitro.
- NEL-derived microglia (NEL-MG) represent a promising in vitro model for studying microglial function.
- This method offers an improved yield of adult-like microglia for research applications.
