A simple assay for adenylate cyclase in intact cells by affinity elution chromatography.
A method using the principle of affinity elution chromatography is described for the assay of adenylate cyclase in intact human platelets. By incubating platelet-rich plasma in the presence of radioactively labelled adenine, the ATP pool of the cells was prelabelled. Formation of labelled cyclic AMP from ATP was determined by extracting the platelets with HC1O4. After removal of the latter as KC1O4, the extract containing cyclic AMP and other adenine nucleotides was adsorbed in a NN-diethyl-N-2-hydroxypropylamino (QAE)-cellulose column. The column was washed, and subsequently cyclic AMP was specifically eluted with a cyclic AMP-dependent protein kinase and the radioactivity of the eluate was determined.
D000262:Adenylyl Cyclases / Q000097:blood*; D001792:Blood Platelets / Q000201:enzymology*; D002846:Chromatography, Affinity / Q000379:methods; D000242:Cyclic AMP / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D011458:Prostaglandins E / Q000494:pharmacology; D011494:Protein Kinases; D013806:Theophylline / Q000494:pharmacology
D011458:Prostaglandins E; D013806:Theophylline; D000242:Cyclic AMP; D011494:Protein Kinases; D000262:Adenylyl Cyclases