Determination of glycogen in small tissue samples.

S Lo, JC Russell, AW Taylor - Journal of applied physiology, 1970 - journals.physiology.org
S Lo, JC Russell, AW Taylor
Journal of applied physiology, 1970journals.physiology.org
MATERIALS AND METHODS Reagents. 1) Thirty percent potassium hydroxide solution
saturated with sodium sulfate. Potassium hydroxide pellets (300 g; Merck no. 73201, reagent
grade) were dissolved in distilled water to 1 liter and saturated with sodium sulfate (Fisher
no. S-421, anhydrous, certified ACS). 2) Ninety-five percent ethanol. 3) Five percent phenol.
Phenol crystals (250 g; Mallinckrodt no. 9928, reagent grade) were dissolved in distilled
water to 5 liters. 4) Sulfuric acid 96-98%(Fisher no. A-300, reagent grade). 5) Standard …
MATERIALS AND METHODS
Reagents. 1) Thirty percent potassium hydroxide solution saturated with sodium sulfate. Potassium hydroxide pellets (300 g; Merck no. 73201, reagent grade) were dissolved in distilled water to 1 liter and saturated with sodium sulfate (Fisher no. S-421, anhydrous, certified ACS). 2) Ninety-five percent ethanol. 3) Five percent phenol. Phenol crystals (250 g; Mallinckrodt no. 9928, reagent grade) were dissolved in distilled water to 5 liters. 4) Sulfuric acid 96-98%(Fisher no. A-300, reagent grade). 5) Standard glycogen solutions. Glycogen powder (25 mg; Fisher no. G-47, reagent chemical) was dissolved in distilled water to 5 ml. This gave a glycogen concentration of 5 mg/ml. Less concentrated standard glycogen solutions were prepared by volumetric dilution of this stock solution.
Procedure. I) Human muscle samples were taken, with a biopsy needle, isolating a piece of muscle weighing approximately 35-50 mg. The muscle samples were immediately transferred to a weighing pan, and all visible fat, connective tissue, and blood were removed with a probe, forceps, and gauze. The samples were then weighed on a Roller-Smith precision torsion balance and transferred with forceps to the bottom of a capped test tube. Immediately the sample and test tube were frozen in Dry Ice and alcohol. The pan, with traces of blood still remaining, was again weighed, and the weight of blood subtracted from the previous weight to give the weight of the sample in the test tube. Samples were maintained deep-frozen until ready for assay. 2) The tubes were kept on ice after removal from frozen storage. One-half milliliter of 30? & KOH saturated with Na2SOd was added to the samples, making sure that the tissue was completely immersed in the solution. 3) The above tubes with the screw cap on were put in a boiling water bath for 20-30 min until a homogeneous solution was obtained.
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