Preparation of High Serum for A1 cells: Difference between revisions
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==== For 400 ml ==== | |||
FCS 40 ml | FCS 40 ml | ||
| Line 8: | Line 8: | ||
Insulin (stock: 1mg/ ml) 4 ml | Insulin (stock: 1mg/ ml) 4 ml | ||
* grow at 25ºC in 2% CO2 | |||
* if you want to grow without CO2, then substitute L-15 for the MEM | |||
* passage once per week 1:3 | |||
* wash with APBS (100 ml PBS + 25 ml ddH2O) | |||
* trypsinize with TE | |||
* stop with media | |||
* centrifuge cells at 100 rpm 3 minutes | |||
to make myotubes: | ==== to make myotubes: ==== | ||
* wash cells 1x with APBS | |||
* put cells in 0.5% FCSmedia | |||
* wait 3 days | |||
Revision as of 10:15, 13 February 2015
For 400 ml
FCS 40 ml Eagle’s MEM 252 ml DdH2O 100 ml Pen/ Strep 4 ml Glutamin 4 ml Insulin (stock: 1mg/ ml) 4 ml
- grow at 25ºC in 2% CO2
- if you want to grow without CO2, then substitute L-15 for the MEM
- passage once per week 1:3
- wash with APBS (100 ml PBS + 25 ml ddH2O)
- trypsinize with TE
- stop with media
- centrifuge cells at 100 rpm 3 minutes
to make myotubes:
- wash cells 1x with APBS
- put cells in 0.5% FCSmedia
- wait 3 days