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A quantitative biochemical and histochemical study of the lead method for localization of adenosine triphosphate-by-drolyzing enzymes
Abstract and Figures
The purpose was to study the nature of the adenosine triphosphatase (ATPase) localized to plasma membranes by the lead method and the significance of the lead-catalyzed hydrolysis of adenosine triphosphate (ATP) for the staining of fixed kidney. A modified Wachstein and Meisel's medium was used. The staining of sections of microsomal sediments from rat kidney was compared with the in vitro activity of (Na+ + K+)–, Mg++– and Ca++– ATPase in the presence of lead. The histochemical relevance of these observations was tested on sections of rat kidney. Localization of (Na+ + K+)–ATPase was not possible at concentrations of lead below the inactivating level (0.5 mM). Mg++–ATPase and Ca++– ATPase were only partially inhibited by 3.6 mM lead and were localized to plasma membranes in glutaraldehyde-fixed kidney. The staining followed a course of activation by Ca++ and Mg++ characteristic for enzymatic hydrolysis and different from the activation of nonenzymatic hydrolysis of ATP. Staining of fixed tissue was maximal at 2-3 mM lead and decreased at higher concentrations. In Mg++–deficient media membrane staining was weak or absent at 5.2-6.8 mM lead, where the rate of nonenzymatic hydrolysis of ATP was high. Thus, in the medium used the lead-catalyzed hydrolysis of ATP contributed little to the staining of fixed kidney.
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