GloSensor™ cAMP Assay Technical Manual

Literature # TM076

G-protein coupled, seven-transmembrane (7-TM) receptors (GPCRs) represent a major class of drug targets, and the prevalence of these receptors in physiologically important signaling events is well known. The GloSensor™ cAMP Assay provides an extremely sensitive and easy-to-use format for the interrogation of overexpressed or endogenous GPCRs that signal via changes in the intracellular concentration of cAMP.
The assay uses genetically encoded biosensor variants with cAMP binding domains fused to mutant forms of Photinus pyralis luciferase. Upon binding to cAMP, conformational changes occur that promote large increases in light output. Following pre-equilibration with substrate, cells transiently or stably expressing a biosensor variant can be used to assay GPCR function using a live-cell,non-lytic assay format, enabling kinetic measurements of cAMP accumulation or turnover in living cells. Moreover, the assay offers a broad dynamic range, showing up to 500-fold changes in light output. Extreme sensitivity allows detection of G_i-coupled receptor activation or inverse agonist activity in the absence of artificial stimulation by compounds such as forskolin.