Description
pH calibration buffers are prepared to obtain two or more defined pH values for calibration of pH electrodes and pH indicator dyes.
MitoPedia methods:
Respirometry,
Fluorometry
MitoPedia topics:
Media for respirometry
Modified hanks buffer
Substance | Formula | Concentration [mM] |
---|---|---|
Calcium chloride dihydrate | CaCl2 * 2 H2O | 0.42 |
Imidazole | C3H4N2 | 2.0 |
Potassium chloride | KCl | 2.4 |
Magnesium chloride hexahydrate | MgCl2*6 H2O | 0.49 |
Sodium chloride | NaCl | 137 |
- pH adjusted to 7.1 at 30 ยฐC.
- pH adjusted with KOH or HCl to create a buffer series: (pH values at 25 ยฐC)
- A (pH 3.3),
- B (pH 6.1)
- C (pH 6.6)
- D (pH 6.9)
- E (pH 7.3)
- F (pH 7.5)
- G (pH 11.1) pH 10.1 at 37ยฐ did not represent the basic endpoint for carboxy-SNARF-1.
- Calibration
- Basic endpoint (for SNARF): fluorophore completely deprotonated.
- Acidic endpoint (for SNARF): fluorophore completly protonated.
- Calibration
Media with low buffering capacity
- Media with low pH buffering capacity are required for measurement of proton flux in intact cells. Commercially available variants of RPMI1640 that contain neither hydrogen carbonate nor HEPES may constitute a starting point, see e.g. Sigma Product number 6504 (not tested by Oroboros Instruments, due to the inclusion of phenol red this medium may (or may not) be problematic for fluorescence).