A Critical Technique for Enzyme Cytochemistry

Abstract

THE accurate localization of an enzyme at the cellular level by a cytochemical technique requires a full investigation of artefacts due to diffusion of enzyme, reagents and reaction products. The classical superimposed-section technique does not have a sufficient resolving power to evaluate diffusion artefacts over the small distances inside a cell. A technique has been developed which enables the enzyme in small areas inside a cell in a tissue section to be inactivated. In this way diffusion during a cytochemical reaction can be investigated. The method makes use of the enzyme-inactivating action of the long wave-length ultra-violet light obtained from a glass-jacketed 250-watt B.T.H. mercury lamp. This is focused on the section mounted in water between two coverslips by an 8-mm. apochromatic objective. A mask in front of the lamp acts as the light source, and in this way slits or patterns of small holes can be focused on the section.