Source: Weinert, M. et al., Isolation, Culture and Long-Term Maintenance of Primary Mesencephalic Dopaminergic Neurons From Embryonic Rodent Brains. J. Vis. Exp. (2015)
This video outlines a method for cultivating dopaminergic neurons from mouse embryonic brain tissue. The process includes treating the midbrain with a proteolytic enzyme, detaching the cells, and culturing them on polymer-coated coverslips in a nutrient-rich medium with antibiotics for their growth and long-term viability.
All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.
1. Dissociation of Ventral Midbrain Cells
2. Plating Ventral Midbrain Cells
3. Culture Growth and Maintenance
The authors have nothing to disclose.
Dulbecco's modified Eagle medium nutrient mixture F-12 | Invitrogen | 11330 | |
Hanks' Balanced Salt Solution (HBSS) (1X), liquid | Invitrogen | 24020-117 | |
Fetal bovine serum, heatinactivated (FBS) | Invitrogen | 16140 | |
N2 Supplement (100X), liquid | Invitrogen | 17502-048 | |
D-(+)-Glucose solution (45% (wt/ vol) in water | Sigma | G8769 | |
Bovine serum albumin BSA | Sigma | A9430 | |
Laminin from Engelbreth-HolmSwarm murine sarcoma basement membrane | Sigma | L2020 | |
Penicillin/streptomycin | Invitrogen | 15070 | |
Trypsin (0.05% (wt/vol) | Invitrogen | 25300 | |
Bovine serum albumin (BSA) cell culture tested | Sigma | A9418 | |
Phosphate-buffered saline (PBS) | Sigma | P3813 | |
Poly-L-ornithine, 0.01% solution | Sigma | P4957 | |
Trypan blue solution (0.4% (wt/vol)) | Biowhittaker | 17-942E |