Glycogen Branching Assay to Measure the Degree of Glycogen Branching

Abstract

Source: Wilson, W. A., Spectrophotometric Methods for the Study of Eukaryotic Glycogen Metabolism. J. Vis. Exp. (2021).

In this video, we describe a qualitative spectrophotometric assay to determine the extent of branching in a glycogen sample with an uncharacterized structure. Comparing the absorbance maxima of the uncharacterized sample with that of polysaccharides with known branched structures indicates the presence of branching in the uncharacterized sample.

Protocol

1. Qualitative assessment of glycogen branching Prepare saturated calcium chloride solution by adding 74.5 g of anhydrous calcium chloride to ~40 mL of water and stirring. Add a little more water and continue to stir. Make the volume up to 100 mL with water and continue stirring until the CaCl2 is fully dissolved. Prepare a working stock of iodine/CaCl2 color reagent by mixing 50 µL of KI/iodine stock solution with 13 mL of saturated CaCl2 solution in a 15 mL tu…

Materials

Amylopectin (amylose free) from waxy corn	Fisher Scientific	A0456
Amylose	Biosynth Carbosynth	YA10257
Glycogen, Type II from oyster	MilliporeSigma	G8751
Methacrylate cuvettes, 1.5 mL	Fisher Scientific	14-955-128	Methacrylate is required since some procedures are conducted at 340 nm or below