Base Medium Yeast Extract (BD 212750) 10.0 g Charcoal (Sigma C-5510) 2.0 g ACES Buffer 10.0 g DI Water 980.0 ml Adjust pH to 6.90 +/- 0.05 using KOH (pH is critical). Autoclave at 121oC. Cool to 50oC. Aseptically add Solutions A and B (see below). Recheck pH and adjust if necessary. Allow sample to cool to room temperature before reading pH. Keep charcoal in suspension when dispensing. For solid medium, add 17 grams of Agar to base medium above. **Note: Medium should be wrapped in foil to protect from light. **Note: Be sure to allow solid products adequate time to dry (at least two days). Agar will become soft and unusable if it does not dry properly. Solution A L-Cysteine HCl 0.4g DI Water 10.0 ml Filter Sterilize. Solution B Fe-pyrophosphate 0.25g DI Water 10.0 ml Filter Sterilize. Note: Keep soluble Ferric pyrophosphate dry and in the dark. Do not use if the chemical loses its green color and becomes brown or yellow. Do not heat over 60oC to dissolve. The mixture dissolves readily in a 50oC water bath.
