AN EVALUATION OF METHODS COMMONLY USED FOR THE FIXATION AND STAINING OF GLYCOGEN

Abstract

Blocks of liver from eight one-month-old Sprague Dawley rats were fixed in the following fixatives: 10% neutral formalin, 10% formalin, formol-saline, alcohol formalin, acetic alcohol formalin, Bouin's, Carnoy's and Rossman's for the following times: 24 hours,48 hours, seven days, two weeks, one month and three months. Sections were cut from these specimens and stained for glycogen by the periodic acid Schiff reaction and the Best carmine method. It is concluded from these experiments that if the tissues are fixed in acetic alcohol formalin, then consistently good demonstration of glycogen will be obtained with the periodic acid Schiff stain, in livers stored in this fixative up to three months. Nearly as good results can also be obtained with the Best carmine stain, if the tissues are similarly fixed in Bouin's. Aqueous based fixatives cannot be relied upon after 24 hours to 48 hours to demonstrate the maximum amount of glycogen in tissues. The phenomena of polarization is discussed and it was shown that acetic alcohol formalin fixed tissues gave the most marked polarization, whilst sections taken from specimens fixed in the aqueous based fixatives show considerably less polarization, and in some cases none at all. Alcohol based fixatives give a coarser and larger granule of glycogen which stains with more vividness, and is easier to recognize following the periodic acid Schiff reaction than the Best carmine. An unusual distribution of glycogen was observed with the two stains that were used. With the periodic acid Schiff reaction, some sections showed a very marked heavy peripheral accumulation of glycogen within the cells, whilst with the Best carmine method some sections showed a glycogen free peripheral zone. They were consistently reproduced but no suitable explanation can be offered for these observations.