There are a few different ways of making slice culture medium for keeping cells happy. I believe there’s currently even a debate out about whether or not it’s good to have heat-inactivated horse serum in the medium since the amino acids are good but there are other hormones in there that might be bad for the cells.
This recipe uses horse serum, though keep in mind the above and the second disadvantage of variability between lots.
Add together the following:
- 190ml water
- 2.1g MEM (Modified Essential Medium)
- 50ml heat-inactivated horse serum (either bought as heat-inactivated or HI’d in house)
- 1.25ml 200mM L-glutamine (also sold as 100x L-glutamine)
- 0.25ml CaCl2
- 0.5ml MgSO4
- 0.58g d-glucose
- 0.11g sodium bicarbonate
- 1.79g Hepes salt
- 0.012ml 25% ascorbic acid (not too yellow as that’s a sign that it’s turning)
- 0.25ml 1mg/ml insulin
Check the pH and adjust it with NaOH so it’s at 7.28. Check the osmolarity and add between 20 and 25ml of water (dd if you have it) so the osmolarity is at 320.
Ascorbic acid, l-glutamine, and insulin should be stored in aliquots in the -80. Horse serum should be stored in 50ml tubes in the -20. MEM powder should be kept in the fridge.
Finally, sterile-filter the slice culture medium and aliquot it into 50ml tubes to be kept in the fridge.
You can keep it in one large bottle if you go through it quickly but keep in mind getting into one container repeatedly means contamination is a greater risk. That, and it will contaminate your entire made-up stock.
