Precautions:
Make sure that all equipment is functioning as expected first, so that you do not get caught unaware in the middle of a protocol.
Media is to be made in the hood. Wash EVERYTHING that goes into the hood. As much as the water is changed in the hot water bath and is blue from an antimicrobial, it is still nasty. We don't want what ever could be in there growing in our culture media, or with our cells.
Materials:
| Component | Where? |
|---|---|
| DMEM | Cold room |
| Fetal Bovine Serum (FBS) | Freezer |
| Penicillin/Streptomycin + (Pen/Strep + L-Glu) | Freezer |
| Non- Essential Amino Acid | +4 Fridge |
Protocol:
- Get a filter column from the cabinet outside the Tissue Culture Room
- Attach the vacuum tube to the nozzle
- Add 400mL of DMEM to the filter column
- Add 50mL of FBS
- Add 5mL of 100x Penicillin-Streptomycin-Glutamine
- Add 5mL of 100x Non Essential Amino Acids
- Let the liquid run through
- Add the final 35mL of DMEM to wash through any of the ingredients above through the filter.
