Two‐dimensional electrophoresis of human parotid salivary proteins from normal and connective tissue disorder subjects using immobilised pH gradients

Abstract

Two‐dimensional electrophoretic analysis of human salivary proteins using immobilised pH gradients in the first dimension, thin‐layer gradient horizontal sodium dodecyl sulfate‐polyacrylamide gel electrophoresis in the second, and modified staining procedures has resulted in a substantial improvement in their resolution. Unlike carrier ampholyte‐based techniques, immobilised pH gradients prevent the loss of proteins of pI>8; accordingly, basic components, including basic proline‐rich proteins, can now be resolved. A two‐dimensional map showing the locations and identities of most of the major proteins has been constructed. Narrow‐range pH gradients can be constructed to give increased resolution of proteins of particular interest. By means of a pH 3.5–5.0 gradient, the abnormal salivary proteins associated with connective tissue disorders were found to be a highly heterogeneous group of pI ∼ 3.75–4.75 and M_r ∼ 32 000; although low levels occurred in some normal individuals, there was less heterogeneity (pI∼3.75–4.25). The technique should form a base for future structural, functional, and clinical studies on human salivary proteins.