Preparation of Collagen-Coated Compartmented Culture Dishes for Neuronal Cell Culture

Abstract

Source: Zepecki, J. P., et al. Real-Time monitoring of human glioma cell migration on dorsal root ganglion axon-oligodendrocyte co-cultures. J. Vis. Exp. (2019)

This video demonstrates constructing a three-compartment cell culture system using collagen and non-reactive grease. This system is used to observe interactions between glioma cells and axons in real-time.

Protocol

1. Compartmented culture of Rat Dorsal Root Ganglia (DRGs), Oligodendrocytes (OPCs), and hGCs (human glioma cells)

1. Preparation of compartmented culture dishes

NOTE: Perform the following steps the days before the planned harvest of the DRGs.

1. Assemble compartmented culture dishes.
2. Dilute collagen stock solution to 500 µg/mL in sterile distilled H₂O; mix thoroughly.
3. With a sterile transfer pipette, fill a 35 mm culture dish with 2 mL of collagen solution; remove the solution, leaving a thin collagen film behind, and place it into the next 35....

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Cell Culture Dish	Corning	430165	35mm X 10mm
Hypodermic Needle, 18G	BD	511097
Ammonium Hydroxide Solution	Fisher Scientific	A669-500	Concentrated
Cell Culture Dish	Corning	430165
Campenot Chamber	Tyler Research	CAMP-10
Hemostatic Forceps	Roboz	RS-7035
Cultrex Rat Collagen I	Trevigen	3440-100-01
Pin Rake	Tyler Research	CAMP-PR
Neurobasal Medium	Thermo Fisher	21103049
silicone grease
metal file
Tray
gauze
CO2 incubator
glass pipette