Histochemical conditions influencing metachromatic staining. A comparative study by means of a model system of polyacrylamide films.

The influence of different histochemical conditions on some metachromatic staining reactions has been studied using polyacrylamide films containing pure glycostaminoglycans. The films were incubated in fixatives without staining, and in glycerol, diethylene glycol and other glycols, formamide, N,N-dimethylformamide, dimethyl sulphoxide and ethanol (of several concentrations) after staining and their absorption (metachromatic) spectra recorded. In the case of heparin and heparan sulphate the metachromasy was disturbed when the films were immersed before staining in some fixative solutions containing formaldehyde and acid. After equilibration of stained films in organic solvents, changes in the absorption peaks were found to depend on the type and concentration of solvent, the type of glycosaminoglycan and the type of dye. Films containing glycosaminoglycan plus protein were used to investigate the blocking of the metachromatic reaction as the result of ionic interactions with proteins. The parameters that influence this phenomenon (e.g type of protein, glycosaminoglycan and dye, pH of staining) are discussed and a three-dimensional picture is introduced which can explain some of the results obtained in these experiments.