Difference between revisions of "Glycogen Determination from Tissues"

Revision as of 15:01, 6 May 2010

Materials and Buffers

• Screw Capped Vials
• 30% KOH, prepared fresh
• 1M Sodium Sulfate
• Ethanol
• 50 mM Sodium Acetate, pH 4.8
• Amyloglucosidease

Protocol

1. Weight out 30-90 mg tissue into screw cap vial and record weights.
2. Add 300 uL 30% KOH and place on 95C heat block for 30 min with occasional mixing.
3. Cool and then add 100 uL Sodium Sulfate and 800 uL Ethanol.
4. Boil for 5 min.
5. Centrifuge at 13 000 RPM for 5 min.
6. Resuspend pellet in 200 uL water then add 400 uL ethanol. Boil 5 min, spin 5 min and Repeat wash steps twice more.
7. Dry pellet
8. Resuspend pellet in 200 uL of 50 mM Sodium Acetate, pH 4.8
9. Add 0.3 mg/mL amyloglucosidase and place at 37C for 3h
10. Quantify glucose

Reference:

PMID 15282316