An easy cAMP extraction method facilitating adenylyl cyclase assays.
We present a facile procedure for measuring adenylyl cyclase activity which circumvents the two-step chromatographic purification of 32P-labeled cAMP. cAMP produced by stimulated cell membrane preparations is easily purified by organic extraction and thus available for quantification using tritium-labeled tracer cAMP and commercially available cAMP-binding protein. The quantification of cAMP is unaffected by the extraction procedure and sample handling. Data obtained by this method were identical to those obtained by the chromatographic method. This procedure could be shown to be suitable for measuring receptor-mediated adenylyl cyclase modulation.
Voss
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Ernst Boehringer Institut für Arzneimittelforschung, Department of Protein Chemistry, Fa. Bender + Company, Vienna, Austria.
Wallner
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D000262:Adenylyl Cyclases / Q000032:analysis*; D000527:Alprostadil / Q000494:pharmacology; D002352:Carrier Proteins / Q000378:metabolism; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D002478:Cells, Cultured; D002623:Chemistry Techniques, Analytical / Q000379:methods; D002725:Chloroform; D000242:Cyclic AMP / Q000032:analysis* / Q000096:biosynthesis; D002373:Cyclic AMP Receptor Protein; D004622:Embryo, Mammalian; D006801:Humans; D007668:Kidney / Q000166:cytology; D007700:Kinetics; D000432:Methanol; D011953:Receptors, Cyclic AMP / Q000378:metabolism; D013268:Stimulation, Chemical; D014316:Tritium
D002352:Carrier Proteins; D002373:Cyclic AMP Receptor Protein; D011953:Receptors, Cyclic AMP; D014316:Tritium; D002725:Chloroform; D000242:Cyclic AMP; D000262:Adenylyl Cyclases; D000527:Alprostadil; D000432:Methanol