Freezing of A1 cells: Difference between revisions
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(Created page with "''in one 175 cm2 flask'' Materials: - APBS (100 ml PBS + 25 ml diwater) - TE in APBS (36 ml APBS + 4 ml TE) - 1x 15 ml FALCON tubes - Pipettes - 1 cryo tube - freezing media ...") |
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| Line 2: | Line 2: | ||
Materials: | Materials: | ||
* APBS (100 ml PBS + 25 ml diwater) | |||
* TE in APBS (36 ml APBS + 4 ml TE) | |||
* 1x 15 ml FALCON tubes | |||
* Pipettes | |||
* 1 cryo tube | |||
* freezing media | |||
Procedure: | Procedure: | ||
# aspirate the old media from the cells | |||
# wash the cells 1 times with 5 ml APBS | |||
# remove the APBS | |||
# add 5 ml TE | |||
# wait till the cells are detached | |||
# stop the TE with HS – media | |||
# centrifuge at 1000 rpm, 3 min at 4ºC | |||
# aspirate the supernatant | |||
# dissolve the pellet into 1 ml freezing media | |||
# transfer the cells into the cryo tube and label the tube with the cell name, passage number and date | |||
# store the tube for one day at –80ºC | |||
# store the vial in liquid N2 | |||
Revision as of 08:40, 12 January 2015
in one 175 cm2 flask
Materials:
- APBS (100 ml PBS + 25 ml diwater)
- TE in APBS (36 ml APBS + 4 ml TE)
- 1x 15 ml FALCON tubes
- Pipettes
- 1 cryo tube
- freezing media
Procedure:
- aspirate the old media from the cells
- wash the cells 1 times with 5 ml APBS
- remove the APBS
- add 5 ml TE
- wait till the cells are detached
- stop the TE with HS – media
- centrifuge at 1000 rpm, 3 min at 4ºC
- aspirate the supernatant
- dissolve the pellet into 1 ml freezing media
- transfer the cells into the cryo tube and label the tube with the cell name, passage number and date
- store the tube for one day at –80ºC
- store the vial in liquid N2