✦ LIBER ✦

A fluorometric method for the determination of functional (Na,K)-ATPase and cardiac glycoside receptors

✍ Scribed by P.A.George Fortes

Book ID: 102986281
Publisher: Elsevier Science
Year: 1986
Tongue: English
Weight: 774 KB
Volume: 158
Category: Article
ISSN: 0003-2697
DOI: 10.1016/0003-2697(86)90575-0

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✦ Synopsis

A method to measure high-affinity binding sites for fluorescent ligands is described. The method is applied to the determination of (Na,K)-ATPase using the fluorescent ouabain derivative anthroylouabain (P. A. G. Fortes (1977) Biochemistry 16, 531-540). The method consists of measurements of the fluorescence intensities of a saturating concentration of anthroylouabain in the presence and absence of (Na,K)-ATPase and ouabain. These data and the fluorescence enhancement factor upon anthroylouabain binding are used to calculate the concentration of binding sites. The measurements can be done in a few minutes and 10 to 100 pmoles of ouabain sites is sufficient for accurate determinations. Because phosphorylation of (Na,K)-ATPase is necessary to bind anthroylouabain, only functional enzyme is detected by this method.

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