A Comparative Study of Three Methods for the Ultrastructural Demonstration of Glycogen in Thin Sections

In order to evaluate 3 staining methods for demonstration of glycogen in thin sections, 2 tissues containing an abundance of this carbohydrate in β-particle form were studied. Tissues were aldehyde-fixed, postfixed in osmium tetroxide, embedded in Araldite and sectioned in the usual manner without special precautions. The first method for staining thin sections employed a sequential combination of periodic acid, thiosemicarbazide and osmium tetroxide vapour, while in the second procedure a silver protein solution was substituted for the osmium tetroxide vapour. The third technique utilized periodic acid, sodium chlorite and uranyl acetate, also in sequential combination. Each method yielded glycogen particles of greater electron density than were seen in sections stained by the usual uranyl acetate-lead citrate procedure. Under high magnification, considerable method-dependent variation in the appearance of the glycogen granules was noted. Particulate substructure, only faintly visible in routinely stained sections, was easily resolved with the periodic acid-thiosemicarbazide-silver protein technique. Conversely, periodic acid-thiosemicarbazide-osmium tetroxide completely obscured this substructure. With periodic acid-sodium chlorite-uranyl acetate, glycogen particles appeared larger, more confluent, and of a less regular outline than with the other methods. Sections were also stained by incubation in periodic acid prior to treatment with lead citrate. The alteration in appearance of the glycogen granules produced by this modification was so great that high-resolution analysis of particle size and substructure could not be undertaken. The usefulness of the procedures investigated here resides in their ability to stain glycogen in thin sections in an intense and selective manner.