Supplement medium after autoclaving with 5.0 g/l D-glucose and 2.0 g/l NaHCO3 added from sterile stock solutions prepared under 100% N2 or 80% N2 and 20% CO2 gas atmosphere, respectively. Adjust pH to 7.0 - 7.2, if necessary.
Compound | Amount | Unit | |||
---|---|---|---|---|---|
|
dehydrated Wilkins-Chalgren medium
(Oxoid CM0643) |
33.0 | g | ||
|
Sodium resazurin
(0.1% w/v) |
0.5 | ml | ||
|
L-Cysteine HCl | 0.3 | g | ||
|
D-Glucose | 5.0 | g | ||
|
NaHCO3 | 2.0 | g | ||
|
Distilled water | 1000.0 | ml | ||
1 Dissolve 33.0 g/l dehydrated Wilkins-Chalgren medium (Oxoid CM0643) in distilled water and add 0.5 ml/l Na-resazurin solution (0.1% w/v), bring to the boil and cool to room temperature while sparging with 100% N2 gas. Add L-cysteine-HCL x H2O (0.3 g/l), dispense the medium under same gas atmosphere into anoxic Hungate-type tubes or serum vials and autoclave. After autoclaving check pH and adjust to pH 6.8 - 7.0, if necessary. | |||||
2 Note: In case resazurin is not colourless addition of 10 - 20 mg sodium dithionite per liter (e.g. from 5% (w/v) solution, freshly prepared under N2 and filter-sterilized) can be used to completely reduce the medium. |