Use of affinity elution chromatography for the microassay of adenosine 3':5'-monophosphate

Abstract

Using the principle of affinity elution chromatography, a microassay has been developed for the determination of picomolar and even smaller quantities of adenosine 3'.5' -monophosphate (cyclic AMP) in biological samples. In principle, cyclic AMP along with the other anions is adsorbed onto an anion exchange cellulose and subsequently, the cyclic nucleotide is specifically eluted with cyclic-AMP -dependent protein kinase. In the actual assay cyclic AMP is determined by competitive displacement of radiolabelled cyclic AMP by non-labelled cyclic nucleotide. The described procedure results in a linear dependence for the displacement of the radioactive nucleotide on cyclic AMP concentration and obviates the usual procedure of sample enrichment by lyophilization. The degree of interference of different nucleotides including cyclic GMP was found to be minimal and in this respect the use affinity elution chromatography was comparable to radioimmunoassay.