Comparing different techniques for obtaining molecular size distributions of glycogen

Glycogen is a hyperbranched glucose polymer, serving as a major energy reservoir in animals. Characterization of the size distributions of complex branched polysaccharides helps in the understanding of their structural and functional properties. This study compared the effectiveness of various techniques in analyzing glycogen size distributions, namely size-separation-based techniques (size exclusion chromatography (SEC) and asymmetric-flow field-flow fractionation (AF⁴)) and imaging-based methods (transmission electron microscopy (TEM) and atomic force microscopy (AFM)). SEC and AF⁴ gave similar glycogen size distributions. TEM was suboptimal for obtaining size distributions due to the difficulty of preventing aggregation artifacts during sample preparation. AFM appeared to avoid the problem, but the size distributions so obtained had substantially lower resolution and were much more laborious to acquire. The microscopy-based techniques, especially TEM, do however provide richer morphological information on individual particles than do SEC and AF⁴. These results suggest that combining size-separation and microscopy (imaging) techniques could provide a holistic view of glycogen size distribution and morphology, which may also be applicable to the characterization of other complex branched polymers.