SUBSTRUCTURES OF THE ACINAR BASEMENT-MEMBRANE OF RAT SUBMANDIBULAR-GLAND AS SHOWN BY ALCIAN BLUE STAINING AND CRYO-FIXATION FOLLOWED BY FREEZE-SUBSTITUTION

The ultrastructure of the epithelial basement membrane was studied in the acinar cells of adult rat submandibular glands after: (i) immersio n fixation in 0.1 M sodium cacodylate buffer (CB, pH 7.2) containing 2 .5% glutaraldehyde (GA) and alcian blue (0.5%) in conjunction with mic rowave irradiation, (ii) perfusion fixation with 2.5% GA in CB, (iii) rapid freezing followed by freeze-substitution (RF-FS) with 1% GA in a cetone, and (iv) RF-FS with 2% OsO4 in acetone. The specimens were pos t-fixed with 1% OsO4 in CB after methods (i) and (ii) but not (iii) an d (iv). Fixed specimens were embedded in epoxy resin and the ultrathin sections were cut, stained with both lead and uranium, and observed u nder a transmission electron microscope. Various substructural compone nts could be seen in the acinar basement membrane. In the specimens pr ocessed by method (iv), a clear meshwork structure could be found just beneath the basal plasma membrane. This meshwork could not be seen in the specimens processed by method (iii) but thin filaments of similar to 100 nm in length extending from the epithelial base toward the con nective tissue space were evident. By methods (i) and (ii), an electro n dense 30-75 nm layer could be seen subjacent to basal cell membranes . By method (ii), particularly, thick threads connecting this layer to collagen fibres in the connective tissue were stained with alcian blu e. Lamina lucida was not identified.