A kinetic assay for Na-K-ATPase activity in isolated renal proximal tubules.
A kinetic assay system which provides reliable measurements of Na-K-ATPase activity on 0.2 to 0.5-mm segments of renal proximal convoluted tubules isolated from collagenase-digested renal cortical slices is described. The use of collagenase digestion provides higher values for Na-K-ATPase, possibly by making the enzyme more accessible to the reaction system. The advantages of a kinetic vs an endpoint assay include the ability to use the same tubule as its own reference for the determination of total, ouabain-sensitive, and ouabain-insensitive ATPase activity. In addition, it allows dose-response studies on the effect of inhibitors on ATPase activity in the same tubule segment.
D000818:Animals; D004305:Dose-Response Relationship, Drug; D066298:In Vitro Techniques; D007687:Kidney Tubules, Proximal / Q000201:enzymology*; D007700:Kinetics; D010042:Ouabain / Q000494:pharmacology; D011817:Rabbits; D000254:Sodium-Potassium-Exchanging ATPase / Q000037:antagonists & inhibitors / Q000302:isolation & purification*
D010042:Ouabain; D000254:Sodium-Potassium-Exchanging ATPase