Obtaining a Mixed Glial Cell Culture from a Neonatal Mouse Brain

Abstract

Source: Sarkar, S. et al., Rapid and Refined CD11b Magnetic Isolation of Primary Microglia with Enhanced Purity and Versatility. J. Vis. Exp. (2017)

The video demonstrated the method of isolating and culturing cells from a neonatal mouse brain. It involved enzymatic and mechanical dissociation of the brain, followed by culturing in growth media lacking neuronal growth factor to obtain a mixed glial population.

Protocol

All procedures involving animal samples have been reviewed and approved by the appropriate animal ethical review committee.

1. Growing of Mixed Glial Cultures

1. Decapitate 1- 2-day-old pups quickly with 5.5 inch operating scissors and place the heads immediately in a 50 mL tube on ice. Note that this decapitation is the mode of euthanasia.
2. In a laminar airflow hood, make a small incision in the skull and meninges using 4.5 inch straight micro-dissecting scissors. Begin cutting from the caudal end to the rostral end (nose). Get underneath the skin by using the opening formed by the decapitation.
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Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
DMEM/F12 (1:1) (1x)	Life Technologies	11330057
Sodium Pyruvate	Life Technologies	11360070
MEM Non-essential amino acids (100x)	Life Technologies	11140050
L-Glutamine (100x)	Life Technologies	25030081
EDTA	Fisher Scientific	AM9260G
Fetal Bovine Serum	Sigma	13H469
0.25% Trypsin-EDTA	Gibco by Life Technologies	25200