QUANTITATIVE HISTOCHEMICAL ANALYSIS OF GLYCOLYTIC INTERMEDIATES AND COFACTORS WITH AN OIL WELL TECHNIQUE

Abstract

Quantitative histochemical measurements of glycolytic intermediates and cofactors in samples of the order of 0.1 µg dry weight have been made possible by the use of enzymatic fluorometric methods in combination with the technique of enzymatic cycling. In order to prevent evaporation of the necessarily small volumes of fluid and to avoid exposure to CO₂ in the air, incubations are carried out under oil in wells drilled in Teflon. The general technique for conducting analyses in oil wells is described together with specific procedures for measuring glycogen, glucose, glucose 6-phosphate, uridine diphosphoglucose, fructose diphosphate, lactate, adenosine triphosphate, phosphocreatine and inorganic phosphate. The potential of the technique is illustrated by measurements of these compounds in each of nine discrete layers of the retina.