View source for Isolation of Mouse Embryonic Fibroblasts (MEFs)

From Zolov et al (under review)

==Materials==
* Collagenase, type II (LS004204, Worthington, US)
* Clean surgical blades
* DMEM/PSG/10% FBS

==Generation of Mouse Primary Fibroblast Cells== 

# MEFs were prepared from tails and legs of P0 pups. 
# Disect  out all four limbs plus the tail.  Place one small piece into PCR tube/plate for genotyping
# Using a clean surgical blade, chop into tiny pieces.  
# Place into 5 mL of collagenase in DMEM/PSG/10% Serum (500U/mL) in a 15 mL falcon tube
# Incubate in incubator for 12 hours at 37°C. 
# After incubation, cells were harvested at 200 g for 5 min
# Wash cells once with warm media, then resuspend in DMEM.  
# Plate into 2-3 100 mm dishes and change media afer 1d
# Split when confluent