Freezing of C2C12 cells: Difference between revisions
Jump to navigation
Jump to search
No edit summary |
No edit summary |
||
| Line 11: | Line 11: | ||
== Procedure == | == 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 | |||
Revision as of 08:38, 27 March 2015
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