Isolation and characterization of rat tubular basement membrane

Tubules were isolated by the passage of coarsely homogenized kidney parenchyma through a series of stainless steel sieves of varying pore sizes. The isolated tubules, which were free of glomeruli or any contaminating tissue, based on light microscopic examination, served as starting material for the isolation of tubular basement membrane (TBM). The tubules were sonically disrupted, and the subsequent isolation procedure was conceptually similar to that of the isolation of glomerular basement membrane (GBM). The chemical composition of TBM was compared to that of GBM and collagen. 3-hydroxyproline, a marker for basement membrane protein, was present in higher amount in TBM (10 residues per 1,000 amino acid residues) than in GBM (7 residues per 1,000). Glycine and hydroxylysine content was also higher in TBM (255 and 27 residues per 1,000, respectively, in contrast to 204 and 18 residues in GBM). The total carbohydrate content was 7.6 mg per 100 mg of dry weight (neutral sugars, 6.3; GlcN, 0.9; GalN, 0.1; and sialic acid, 0.3). By electron microscopic examination, the isolated TBM was similar to GBM, except for greater variability in diameter and serrated appearance of one surface of the tubular basement membrane fragments. The method is suitable for the isolation of TBM in sufficient quantity for subsequent chemical analyses and immunologic studies.