Demonstration of intracellular glycogen, peroxidases and viruses by light and electron microscopy

Abstract

The objective of the present study vvas primarily to determine vvhether modifications to the existing osmium tetroxide/potassium ferrocyanide method could be used as a general ultrastructural glycogen stain and whether this glycogen, vvhen so contrasted, differed in appearance in different tumours, such as Evving's sarcoma, Leiomyosarcoma, Rhabdomyosarcoma and Hepatocellular carcinoma. Normal skeletal muscle and liver were used to obtain a standard for the appearance of glycogen, and then, with diagnostic criteria in mind, the four glycogen-rich tumours, mentioned above vvere examined to determine the appearance, distribution and amount of glycogen present in them. Modifications to the osmium tetroxide/potassium ferrocyanide (OPF) method consisted of raising the concentration of the ferrocyanide, and using no en bloc or thin section staining by any uranyl salt solutions, because solutions of uranyl salts leaches glycogen from tissue. This procedure resulted in extremely electron-dense intracellular glycogen being retained, vvhich aided diagnosis of the these tumours. At high magnification. (>X30000) there vvas no morphological difference betvveen the glycogen particles in the various tumours, so these particles as such could not be used as a diagnostic criterion. Existing methods for the demonstration of myeloperoxidase and platelet peroxidase vvere modified to obtain more precise localization of the diaminobenzidine (DAB) reaction product. Different anti-coagulants, one of which was heparin and a modified fixation procedure vvas follovved.