Media
CO-1
(the media is clear, but complex to make up)
| Ingredient | Amount |
| CaCl2•H2O | .025 gm |
| NaCl | .216 gm |
| KCl | .075 gm |
| KH2PO4 | .016 gm |
| MgSO4•7H2O | .059 gm |
| Fructose | 1.52 gm |
| Glucose | 0.1 gm |
| NaPyruvate | .002 gm |
| NaLactate | 0.37 ml |
| NaHCO3 | 0.3 gm |
| HEPES | 0.240 gm |
| BSA | 0.3 gm |
| Water | 100 ml |
| Ingredient | MW |
mM |
amount / 100 ml |
| CaCl2 | 147.02 |
2.0 |
29.4 mg |
| KCl | 74.56 |
3.1 |
23.1 mg |
| MgCl2•6H2O | 203.3 |
0.4 |
8.1 mg |
| NaH2PO4•4H2O | 137.99 |
0.3 |
4.1 mg |
| Lactic acid (60%) | 112.1 |
21.6 |
368 µl |
| NaCl | 58.44 |
100.00 |
584 mg |
| Na Pyruvate | 110.0 |
1.0 |
11 mg |
| NaHCO3 |
84.01 |
25.0 |
210 mg |
| HEPES (free acid) | 238.3 |
10.0 |
238 mg |
| BSA (Fraction V) | 600 mg |
||
| Add approximately 80 ml of double distilled water to a beaker. With stiring, add components 1-9 in order. pH the media to 7.54 at 20-22°C using 5N NaOH. Add water to bring to 100 ml. This can be stored up to two weeks as a stock at 5°C if filtered and kept sterile. On day of use, add BSA. The pH of the final medium is 7.4 at 39°C. If planning to use at another incubation temperature, the change in the medium pH with temperature is -0.008 pH units / °C. | |||
| Ingredient | Amount (ml) |
| CO-1 or BGM-3 | 47.5 ml |
| Skim milk (CO-2) | 47.5 ml |
| Egg yolk | 2 ml |
| Glycerol | 3 ml |
| Ingredient | Amount (ml) |
| Sanalac | 10.29 g |
| Glucose | 0.57 g |
| Water | 100 ml |
| Osmolarity = 310 mOsm | |