Isolation of Mouse Embryonic Fibroblasts (MEFs)
From Bridges Lab Protocols
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
