- Glucose Oxidase powder
- Catalase
- 20x SSC buffer
- 50% Glucose Solution
- 15 g Glucose + 30 mL water, heat up to 60C and/or Nutate overnight to dissolve.
- ( Recipe for 10 mL, rescale appropriately, up to 40mL in a 50mL tube. Buffer lasts 24-48 hours at RT)
- In a 1.5 uL tube measure out 2.5 mg Glucose Oxidase (yellow powder, stored at -20C) and dissolve it in 500 uL 2x SSC by vortexing
- 500 uL should be enough to dissolve up to 40 mgs
- final concentration should be 0.25 mg/mL (0.5 mg/mL is saturating)
- Add 20 uL dissolved Catalase solution (20 mg/mL stock) (yellow brown liquid, sediment free). Mix gently. Do Not vortex.
- In a 50 mL flask, add 1 mL 20x SSC
- Add 2 mL of 50% Glucose Solution (stored at RT, filter sterilize)
- Add all 520 uL of enzyme mix
- Gently fill to 10 mL with ddH2O.
- (avoid mixing in extra air, the point of this buffer is to remove disolved oxygen from the buffer).
- overlay solution with ~1 cm (2-3 mL) of mineral oil.
- (this prevents oxygen from the air dissolving back into the buffer)