Passaging C2C12 cells

Material

• 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

1. remove 5 ml of the old media in the 15 ml FALCON tube
2. throw the rest of the old media away
3. wash the cells with 5 ml PBS
4. remove the PBS
5. add 2 ml of the TE in the dish
6. let this incubate for ~4min at 37ºC
7. stop the TE with the old media and resuspend the cells in this
8. pipette the cell suspension in the FALCON tube back
9. centrifuge the suspension for 1000 xg for 3 min at 4 ºC
10. remove the old media
11. resuspend the cells in an appropriate volume of HS medium (f.e.: 10ml)
12. fill in the new 75 cm2 flask 14 ml of new media
13. 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)
14. close the flask and put it in the 37 ºC