There are various methods for staining mucus material, such as AB-PAS staining, mucus HID-AB staining, and standard Alisin Blue staining. Most of the above methods utilize the principle that cationic dyes such as Alisin Blue (Alcian) can bind to acidic groups, i.e., Alisin Blue forms insoluble complexes with anionic groups such as carboxyl groups and sulfate contained in the tissue.

The staining principle of the modified Hale colloidal iron mucopolysaccharide staining solution is that the colloidal iron particles can be tightly bound to the acidic mucus material to form an insoluble complex under acidic conditions, and then the acidic mucopolysaccharides are rendered blue by using the Prussian blue reaction. The coloration of this method is deeper than that of Alisin blue method, and it is not easy to be eluted. It is commonly used for the diagnosis of renal smoky cell carcinoma and acidic mucopolysaccharides in rat epithelial cells.