Freezing of C2C12 cells: Difference between revisions
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Materials: | ==Materials:== | ||
* 15 ml FALCON tube | * 15 ml FALCON tube | ||
* PBS w/o Ca2+ and Mg2+ | * PBS w/o Ca2+ and Mg2+ | ||
| Line 10: | Line 10: | ||
* freezing box | * freezing box | ||
==Procedure:== | |||
- wash the cells with 5 ml PBS | - wash the cells with 5 ml PBS | ||
Revision as of 07:35, 24 October 2014
Materials:
- 15 ml FALCON tube
- PBS w/o Ca2+ and Mg2+
- TE in PBS
- Freezing media (10% DMSO + 40% serum + 50% DMEM)
- Pipettes and tips
- Glas pipetts
- cryo tube
- ice
- freezing box
Procedure:
- wash the cells with 5 ml PBS - remove the PBS and add 2 ml TE - let it incubate for 3 – 5 min - stop the TE with 5 ml HS – media and resuspend the cells - pipette the cell suspension back in the FALCON tube - centrifuge the cells (1000 xg, 3 min at 4 ºC) - remove the old media - resuspend the cells 1 ml freezing media - quickly pipette the liquid in the cryo tube - put the cells in the freezing box and store this for one day at –80 ºC - place the cells one day later into the liquid nitrogen