THEORETICAL AND EXPERIMENTAL ASPECTS OF ENZYME DETERMINATION IN A CYTOCHEMICAL MODEL SYSTEM OF POLYACRYLAMIDE FILMS CONTAINING ALKALINE PHOSPHATASE

Abstract

A theoretical treatment of the influence of slow substrate diffusion on the kinetics of an enzyme reaction in a cytochemical system is given. Quantitative aspects of the effect of several parameters on the diffusion of substrate were studied experimentally on a model consisting of polyacrylamide films into which alkaline phosphatase was incorporated. Spectra and staining intensity of the films were measured in a special film colorimeter. Enzyme activity in the films was determined cytochemically (with the azo dye coupling method) and biochemically. An effectiveness factor for which diffusion is not rate-limiting proved to be attainable with both methods.