Optical anisotropy of alcian blue-stained acid glycosaminoglycans

Optical anisotropy as dispersion of birefringence (DB) (birefringence studied for light of different wavelengths) and linear dichroism (LD) (selective absorption of polarized light) in stained substrates reflects their macromolecular orientation states. Birefringence interference colors of alcian blue (AB)-stained glycosaminoglycans (GAG) and glycoconjugates observed with polarization microscopy have been found to vary, although their staining characteristics under unpolarized light are practically the same. We investigated the optical anisotropy of GAG-AB and some glycoconjugate-AB complexes used as standards, to provide a basis for interpreting results for AB-stained materials in situ. Filamentous preparations of hyaluronic acid (HA), chondroitinsulfates, proteoglycans, and a mucus sulfoglycoconjugate were studied. Anomalous DB (birefringence sign changing with the wavelength of the incident light) was generally observed, but LD was seen only in the AB-HA complex. LD simultaneous to anomalous DB characteristics on the AB-HA complex were assumed to be caused by a maximally oriented helical conformation of the HA. For the other AB-GAG studied, the optical anisotropic characteristics were suggestive of some degree of folding of their chains into a tertiary structure. The profiles of the anomalous DB curves for the AB-stained sulfoglycoconjugate differed from those of the other materials, probably due to different organization of its dye-binding sites.