Difference between revisions of "Passaging C2C12 cells"
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| Line 9: | Line 9: | ||
* remove 5 ml of the old media in the 15 ml FALCON tube | * remove 5 ml of the old media in the 15 ml FALCON tube | ||
* throw the rest of the old media away | * 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 | - let this incubate for ~4min at 37ºC | ||
- stop the TE with the old media and resuspend the cells in this | - stop the TE with the old media and resuspend the cells in this | ||
Revision as of 08:34, 28 April 2014
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