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(Na+,K+) -ATPase: a new assay of Na+-ATPase reveals covert anti-pump antibodies

Schenk, D.B.; Grosse, R.; Ellisman, M.H.; Bradshaw, V.; Leffert, H.L.

Analytical Biochemistry 125(1): 189-196

1982

ISSN/ISBN: 0003-2697
PMID: 6128943
Document Number: 190886
A new assay is described for rat (Na+, K+)-ATPase prepared for renal medullary or crude liver membranes. With ATP at 1 .mu.M, initial rates of ouabain-sensitive decreases in substrate concentrations are followed by measuring diminished ATP-driven luciferin-luciferase light production. Under these conditions, using highly purified enzyme preparations, Na+ and K+ ions stimulate and inhibit initial ATP hydrolysis rates, respectively. Therefore, it is likely that the assay measures Na+-ATPase partial reactions of the pump. A monospecific polyclonal rabbit anti-rat pump antiserum blocks Na+-dependent ATPase measured with the luciferase-linked ATPase assay, whereas conventional assays of purified pump activity at 3.0 mM ATP fail to reveal immunochemical blockade.

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