A rapid method for separation and detection of human salivary amylase isoenzymes by isoelectric focusing in polyacrylamide gel

Abstract

Human salivary proteins were separated by isoelectric focusing on polyacrylamide gel in flat beds at 1000 V for 40 min. Amylase activity was detected after immersing the gel in 0.4 M tris-HCI buffer pH 7.4 to equilibrate the pH gradient. The enzyme activity was detected after diffusion into an overlayer of agarose gel containing an insoluble dye-starch polymer (Phadebas). Both whole human saliva and parotid saliva from 15 different persons contained four amylase components, except in three cases where only three bands were detected. The bands were all focused within a rather narrow pH range (pH 5.4-7.2) and the results were very reproducible.