2.8. Glycogen Quantification

Hepatic and muscle glycogen was isolated as described [12]. Tissue homogenates (10%) were made in 0.03 N HCl and spread evenly on pieces of filter paper (Whatman 3 MM chromatography paper, 2.0 × 2.0 cm) in duplicate. The papers were dropped immediately into a beaker containing 66% EtOH stirred gently by a rotating magnet screened from the papers by a wire mesh. The papers were subsequently washed three times for 40 min in 66% EtOH. They were then briefly rinsed with acetone and dried under a stream of warm air. The dried filter papers were cut into four pieces and placed in a tube containing 0.4 mL of 0.2 M acetate buffer, pH 4.8; 0.2 mg of amylo-α-1,4-a-1,6-glucosidase; and H2O to a final volume of 2 mL. The vials were incubated for 90 min at 37 °C with gentle shaking. Appropriate controls were prepared by incubating aliquots of homogenate in acetate buffer minus amyloglucosidase. Glucose content in the incubated samples was determined by the glucose oxidase method.