Wang, Fang I.; Ding, Gucci; Ng, Garmen S.; Dixon, S. Jeffrey; Chidiac, Peter published the artcile< Luciferase-based GloSensor cAMP assay: Temperature optimization and application to cell-based kinetic studies>, Recommanded Product: (S)-2-(6-Hydroxybenzo[d]thiazol-2-yl)-4,5-dihydrothiazole-4-carboxylic acid, the main research area is luciferase glosensor cAMP assay temperature optimization kinetics drug discovery; Drug discovery; GPCRs; GloSensor; High-throughput screening; Optimization.
G protein-coupled receptors (GPCRs) are an important receptor superfamily and common therapeutic targets. The second messenger cyclic adenosine monophosphate (cAMP) is a key mediator in many GPCR signaling pathways. Monitoring intracellular cAMP levels can help identify orthosteric agonists and antagonists, as well as allosteric modulators. In this regard, luminescence-based biosensors have revolutionized our ability to monitor GPCR signaling kinetics. The GloSensor cAMP assay enables real-time monitoring of signaling downstream of many GPCRs. However, it is crucial to optimize assay conditions such as temperature As well, it has not been reported whether the effects of temperature on biosensor activity are reversible. Here, we describe the temperature sensitivity and reversibility of the GloSensor cAMP assay, and which GloSensor version is optimal for measuring cytosolic cAMP. We also present a detailed protocol for monitoring cAMP levels in live cells expressing endogenous or exogenous GPCRs. Temperature optimization studies were carried out using HEK293H cells transiently transfected with the adenosine receptor A2a and the GloSensor plasmid (pGloSensor-20F or -22F). We found that preincubation and luminescence reading at room temperature were optimal as compared to higher temperatures As well, the GloSensor-22F biosensor had a superior signal-to-background ratio and the effect of temperature on biosensor activity was reversible. However, thermal instability of the biosensor may pose a problem for in vivo studies. Nevertheless, the GloSensor cAMP assay can be applied to analyze signaling by a wide range of GPCRs for drug discovery purposes.
Methods (Amsterdam, Netherlands) published new progress about Adenosine A2A receptors Role: BSU (Biological Study, Unclassified), BIOL (Biological Study). 2591-17-5 belongs to class thiazole, and the molecular formula is C11H8N2O3S2, Recommanded Product: (S)-2-(6-Hydroxybenzo[d]thiazol-2-yl)-4,5-dihydrothiazole-4-carboxylic acid.
Referemce:
Thiazole | C3H3NS – PubChem,
Thiazole | chemical compound | Britannica