| Prepare a 1 m solution of HEPES in ddH2O and adjust the pH to 7 with 3 m KOH. Then prepare 30 mL of the following solution in ddH2O (the final concentrations are for a 50-mL volume). Adjust the volume to 45 mL and verify that pH ∼ 7.
|
| KCl |
125 mm |
| K Gluconate |
2.5 mm |
| HEPES (1 m, pH 7)
|
10 mm |
| To the previous solution, add the following (the final concentrations are for a 50-mL volume). Readjust the pH to 7.2–7.4
with KOH and correct the osmolality to 294 mOsm (rat) or 311 mOsm (mouse) with sucrose.
|
| MgATP |
4 mm |
| NaGTP |
0.3 mm |
| Na-Phosphocreatine |
10 mm |
| Filter the solution using a nylon filter (pore size 0.2 µm; e.g., Nalgene #176) and make 1-mL aliquots. Store at −20°C for
up to 6 mo. Verify the osmolality when thawed.
|
