Isolating and Culturing Oligodendrocyte Precursor Cells from Mouse Pup Brain Cortex

Abstract

Source: Sinyuk, M., et al. Dissection and Isolation of Murine Glia from Multiple Central Nervous System Regions. J. Vis. Exp. (2020)

This video demonstrates a technique for isolating and culturing oligodendrocyte precursor cells (OPCs) from the mouse pup brain cortex.

Protocol

All procedures involving sample collection have been performed in accordance with the institute's IRB guidelines.

1. Tissue dissociation

NOTE: All the following procedures are carried out in a sterile tissue culture designated biosafety cabinet using aseptic technique and sterile materials.

1. Add 1 mL of 0.05% trypsin containing 0.53 mM EDTA to each 15 mL conical tube of 9 mL DMEM and tissue to begin tissue lysis.
   NOTE: DMEM contains a high amount of calcium chloride, which can act as an inhibitor of trypsin. If dissociation is n....

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
0.05% Trypsin and 0.53 mM EDTA	Gibco	25300054	Tissue dissociation
1X PBS pH 7.4	Gibco	10010031	Standard reagent
50 mL, 25 cm square cell culture flask	Greiner Bio-One	690 175	Cell culture (T25) flask
Antibiotic-Antimycotic 100X	Gibco	15240-096	Media component
B-27 Supplement 50X	Gibco	17504-044	Media component
Bovine serum albumin	Sigma	A9647-50G	Antibody diluent
DMEM (1X), high glucose with Na pyruvate	Gibco	11995040	Media component
Dnase I	Sigma	10104159001	Tissue dissociation
Fetal bovine serum heat inactivated	Gibco	A3840001	Media component
Hanks' Balanced Salt Solution (w/o Ca or Mg)	ThermoFisher	14170120	Tissue dissociation
L-glutamine, 200mM	Gibco	20530081	Media component
Neurobasal	Gibco	21103-049	Media component
Normal goat serum	Sigma	G9023	Blocking solution component
Poly-D-Lysine 12 mm #1 German Glass Coverslip	Corning Biocoatt	354086	Cell adherent