Fig. 2.

Isolation and characterization of human gastric mucins.A, each specimen was separated into four types of mucin samples based on their tissue location and solubility in GuHCl. B, mucins isolated from these samples using isopycnic cesium chloride density gradient centrifugation (representative graph). Fractions from the density gradient were analyzed for carbohydrate (black circle) and DNA (absorbance at 260 nm, white diamond) contents. The absorbance at 260 nm that represents the DNA is present as a sharp peak in fraction 4, whereas the absorbance at 260 nm present as a slope above fraction 15 represents nonmucin proteins. The vertical dashed lines indicate the mucin-containing fractions that were pooled for further experiments. C, Western blot from an SDS-AgPAGE (0.5% agarose with a 0–6% polyacrylamide gradient) of soluble mucins from surface soluble (SS), insoluble mucins from surface (SI), soluble mucins from the glands/deeper tissue (SD), insoluble mucins from the glands/deeper tissue (ID), stained with antibodies against MUC5AC and Leb. The line where the mucin bands were halted represents the start of the polyacrylamide gradient, that is, the mucins remained in the part of the gel that contained only agarose. AgPAGE, agarose PAGE; GuHCl, guanidinium chloride; Leb, Lewis b.