Limitation of cane sugar solution as a tissue fixative for glycogen preservation in mouse liver

Sugar-based natural fixatives have been proposed as substitutes for formalin in histopathological practice because of their promising results in preserving tissue structures. However, they have not yet been applied. This study aimed to determine the optimal concentration of cane sugar solution (CSS) for tissue fixation, histochemical staining, and glycogen storage in mouse liver tissues.

A total of 24 mouse liver tissues from six mice were divided into four groups and fixed with 10% neutral buffered formalin (NBF), 30%, 50% and 70% CSS, respectively, for 24 h at room temperature. After tissue processing and sectioning, the samples were stained with hematoxylin and eosin (H&E), methyl green pyronin Y (MGPY) and peroxidase acid-Schiff (PAS). The tissue sections were evaluated under a light microscope by two blinded pathologists. Data were analyzed using one-way analysis of variance (ANOVA) and a post-hoc test to assess the differences among the experimental groups.

The gross morphology of all the samples showed minimal shrinkage without color changes. Microscopic examination revealed that the 50% and 70% CSS groups showed comparable efficacy in cytoplasmic and nuclear staining, H&E staining intensity, deoxyribonucleic acid (DNA) and ribonucleic acid (RNA) staining, and collagen staining with the 10% NBF group. The 30 percent CSS group showed significantly poorer nuclear and DNA staining than the 10% NBF group. All CSS groups failed to preserve glycogen in mouse liver tissues (p < 0.001) compared with the 10% NBF.

Our study concluded that 50% and 70% CSS are potentially replaceable, eco-friendly fixatives for 10% NBF in small tissue samples for tissue fixation and histochemical staining. However, 10% NBF was still superior to CSS for glycogen preservation in mouse liver tissue.