Fibroblasts Isolation (from mouse ear)

This is a usefull technique to screen phenotype of transgenic mice

• cut piece of mouse ear (app. 3x3mm)
• put into 70% ETOH for 2min
• transfer into DMEM
• remove from DMEM cut into small pieces using scalpel
• transfer into 13ml round bottomed tube and add 1-2ml of 0.25% trypsin, vortex briefly
• incubate for 1hr at 37 °C (water bath) vortexing briefly every 10min
• inactivate trypsin by adding 3ml of DMEM/10%FCS
• spin 1000rpm/5min
• aspirate sup, resuspend in 2ml of fresh DMEM/10%FCS, transfer into well of 6well plate
• change medium for fresh day after

Fibroblasts will grow after 1-2 days and cover well bottom after 3-5 days.