cGMP, synthesized from GTP by both membrane-bound and soluble guanlyate cyclase enzymes, is a key intracellular second messenger molecule that transduces cellular signaling events in response to a variety of hormones, autacoids and drugs. The relative abundance of cGMP within a given cell can serve as a marker for activation by agonists acting through particulate guanylate cyclase at the cell surface, or intracellular activation of soluble guanylate cyclase by nitric oxide. Cayman’s cGMP Assay is a competitive ELISA that can be used for quantification of cGMP directly obtained from cell lysates, tissue homogenates, plasma or urine. The assay has a range from 0.23-30 pmol/ml and a sensitivity (80% B/B0) of approximately 1 pmol/ml. Since the antibody used in this assay was prepared against a cGMP-carrier protein conjugate, antibody binding is increased if an acetyl group is present on the 2' hydroxyl group of the cGMP. The optional acetylation procedure for both samples and standards increases the sensitivity of the assay approximately ten-fold.