Passaging C2C12 cells

materials

• 1x 15ml FALCON tube
• PBS w/o Ca2+ and Mg2+
• TE in PBS (4 ml TE + 36 ml PBS)
• HS for C2C12 cells
• pipettes
• 75 cm2 flask

procedure

• remove 5 ml of the old media in the 15 ml FALCON tube
• throw the rest of the old media away
• wash the cells with 5 ml PBS
• remove the PBS
• add 2 ml of the TE in the dish
• let this incubate for ~4min at 37ºC
• stop the TE with the old media and resuspend the cells in this
• pipette the cell suspension in the FALCON tube back
• centrifuge the suspension for 1000 xg for 3 min at 4 ºC
• remove the old media
• resuspend the cells in an appropriate volume of HS medium (f.e.: 10ml)
• fill in the new 75 cm2 flask 14 ml of new media
• pipette an appropriate volume of the cell suspension in he new flask and resuspend it in the new flask (f.e.: if you want to dilute your cell 1: 20 put in the new flask 0.5 ml of the resuspended cells in the FALCON tube)
• close the flask and put it in the 37 ºC