A highly sensitive adenylate cyclase assay

• Published in: Analytical biochemistry Vol. 58; no. 2; pp. 541 - 548
• Main Authors: Salomon, Yoram, Londos, Constantine, Rodbell, Martin
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.04.1974
• Subjects: Adenylyl Cyclases - analysis; Adenylyl Cyclases - metabolism; Aluminum; Animals; Cell Membrane - enzymology; Chromatography; Chromatography, Ion Exchange; Evaluation Studies as Topic; Glucagon - pharmacology; Kinetics; Liver - cytology; Liver - enzymology; Methods; Microchemistry; Phosphorus Radioisotopes; Rats; Time Factors
• ISSN: 0003-2697; 1096-0309
• DOI: 10.1016/0003-2697(74)90222-X

A highly sensitive adenylate cyclase assay method has been developed which employs sequential chromatography on columns of Dowex cation exchange resin and aluminum oxide. With the use of [α- 32P]ATP as substrate, this method permits the nearly complete separation of cyclic [ 32P]AMP formed from the substrate and other 32P-containing compounds, i.e., 32P in the assay blanks was barely detectable. In comparative studies, this method was found to be considerably more sensitive than previously reported methods. The high sensitivity of this method permits detection of the small amounts of cyclic AMP formed at low enzyme concentrations or at early time points in kinetic studies.