Altman:Protocols/Preparing Reagents/Making GOC

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Making Glucose Oxidase Catalase

FINAL CONCENTRATION of 50x GOC stock:\\ 10.8 mg/mL glucose oxidase\\ 1.8 mg/mL catalas in 1x AB-DTT and 50% Glycerol \\

Glucose Oxidase, Aspergillus niger Calbiochem, 346385

Catalase, from Bovine Liver Sigma, 19.9 mg/mL

Glycerol, Anhydrous

Made 2xAB-DTT (10x AB stock and the 100x DTT stock in half the usual AB volume) Assay Buffer (AB) 1 mL AB 10x AB stock (Duncan Freezer) 200 μL 100x DTT stock (Duncan Freezer) 20 μL Filtered water 780 μL

1. Make 43 mg/mL glucose oxidase (GO) in 2xAB. Combine 17 mg of GO with 400 μL 2xAB to make ~400 μL of GO.

2. Make 7.2 mg/mL catalase. Combine 145 μL of catalase with 255 μL of 2xAB to make 400 μL of catalase.

3. Add glycerol to sample 1:1 volume/volume. Added 400 μL of glycerol to each. NOTE: be careful to not introduce bubbles, especially in the GO. This produces 100x stocks of GO and catalase.

4. Combine the GO stock and catalase stock 1:1 volume/volume to yield a 50x stock of GOC. Combined 700 μL of each Aliquoted into 50 μL stocks which are stored at 20°C (in Ron’s freezer. Do not freeze. Made 27 aliquots.