Contribution of confocal laser scanning microscopy to glycochemistry of mouse and rat submandibular glands by single and double lectin staining

The localization of individual glycosidic residues in the mouse and rat submandibular glands was examined using confocal laser scanning microscopy (CLSM). For these organs we tested some procedures of fixation and embedding to better understand the distribution of some lectin-probes inside well preserved secretory cells and observed that fixation and inclusion steps did not influence appreciably the location and intensity of the reactive sites. The fixation mixture of 4% paraformaldehyde, 1% glutaraldehyde and 0.2% picric acid produced the most satisfactory results. In specimens labeled with PNA-, Con A-, LTA-FITC and WGA-, DBA-TRITC lectins, the convoluted granular tubules (CGT) proved to be composed of secretory granules with high-density lectin labeling. The complex organization of secretory glycocomponents within the granule matrix was further resolved by double labeling and dual scanning experiments. Some lectins exhibited colocalization while others displayed differential localization providing information about the occurrence of O- and N-linked glycoconjugates. The CLSM technique applied to fluorochrome-conjugated lectins also revealed a more marked dimorphism in the rat rather than in the mouse submandibular gland. In particular, the male rat submandibular gland was found to consist of CGT heterogeneous cell populations, while the mouse submandibular gland did not show glycochemical differences between cells. Female rats exhibited a lectin profile very different from that of female mouse.