View source for Isolation of Mouse Embryonic Fibroblasts (MEFs)

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.