Difference between revisions of "Isolation of Mouse Embryonic Fibroblasts (MEFs)"

Revision as of 16:32, 4 September 2012

From Zolov et al (under review)

Generation of Mouse Primary Fibroblast Cells

Fibroblasts were prepared from tails and legs of P0 pups. Tissues from individual animals were kept separate, disaggregated with a surgical blade and digested with 500 U/ml type II collagenase (LS004204, Worthington, US) in 5 ml RPMI media (Cat. # 1640, Invitrogen, US) supplemented with 10% FBS, 2 mM L-glutamine and penicillin/streptomycin for 12 hours at 37°C. After incubation, cells were harvested at 200 g for 5 min and plated in 100 mm dishes, in fresh RPMI medium supplemented with 10% FBS, 2 mM L-glutamine and penicillin/streptomycin. Experiments were performed on cells that were passaged 1-3 times.