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 Splitting ASM cells
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Summary
To keep cells viable they need to be divided (split) over multiple plates after they have become confluent. This provides cells with space to keep dividing.
Materials & Methods
Materials
Method
- Warm solution required in water (PBS, DMEM S+)
- Switch on laminar flow, clean it and spray everything required (except for cells) with EtOH (70%) prior to putting into the laminar flow
- Switch the vacuum pump on
- Check cells under microscope
- Remove medium with vacuum pump
- Rinse cells twice with 5 mL PBS and remove buffer with vacuum pump
- Defrost trypsin
- Add 1 mL trypsin
- Incubate 37 °C, 5 min.
- Prepare Greiner tube with 39 mL DMEM S+
- Prepare 3 Petri dishes with Name, Donor and Passage
- After incubation add 5 - 10 mL DMEM S+ to cells (from prepared 39 mL)
- Rinse the dish with the medium in order to get all cells removed from plate and in the liquid
- Remove cell suspension and add to the remaining medium in the prepared Greiner tube (39 mL + 1 mL trypsin = 40 mL total volume)
- Divide cells over the 4 petri dishes (recycle previous one) 10 mL each
- Incubate @ 37 °C
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Splitting ASM cells
