Difference between revisions of "Passaging A1 cell"
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| + | '''Materials:''' | ||
| + | * APBS (100 ml PBS + 25 ml ddwater) | ||
| + | * TE in APBS (36 ml APBS + 4 ml TE) | ||
| + | * Gelatin (warmed to 37ºC) | ||
| + | * HS for A1 cells | ||
| + | * 1x 15 ml FALCON tubes | ||
| + | * Pipettes | ||
| + | * 3x 175 cm2 flasks | ||
| + | Procedure: | ||
| + | - put 10 ml Gelatin into the new 175 cm2 flasks ( transfer the gelatine from the first to the second and from the second into the third → remove the remaining liquid) | ||
| + | - 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 5 ml HS – media | ||
| + | - centrifuge at 1000 rpm, 3 min at 4ºC | ||
| + | - aspirate the supernatant | ||
| + | - dissolve the pellet into 3 ml fresh media | ||
| + | - fill in the new flasks 25 ml HS for A1 cells | ||
| + | - add into each flask 1 ml of the cell suspention | ||
| + | |||
| + | The cells grow at 25ºC | ||
Revision as of 08:06, 26 September 2014
Materials:
- APBS (100 ml PBS + 25 ml ddwater)
- TE in APBS (36 ml APBS + 4 ml TE)
- Gelatin (warmed to 37ºC)
- HS for A1 cells
- 1x 15 ml FALCON tubes
- Pipettes
- 3x 175 cm2 flasks
Procedure: - put 10 ml Gelatin into the new 175 cm2 flasks ( transfer the gelatine from the first to the second and from the second into the third → remove the remaining liquid) - 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 5 ml HS – media - centrifuge at 1000 rpm, 3 min at 4ºC - aspirate the supernatant - dissolve the pellet into 3 ml fresh media - fill in the new flasks 25 ml HS for A1 cells - add into each flask 1 ml of the cell suspention
The cells grow at 25ºC