The cAMP-Glo™ Assay is a homogeneous, bioluminescent and high-throughput assay for measuring cAMP levels in cells. The cAMP-Glo™ Assay monitors cAMP production in cells in response to the effects of test compounds on G protein-coupled receptors (GPCR). GPCRs that couple with adenylate cyclase will increase or decrease intracellular cAMP. The assay is based on the principle that cyclic AMP (cAMP) stimulates protein kinase A (PKA) holoenzyme activity, decreasing available ATP and leading to decreased light production in a coupled luciferase reaction.
The cAMP-Glo™ Assay can be performed in 96-, 384- or 1536-well plates. The cells are induced with a test compound for an appropriate period of time to modulate cAMP levels. After induction, cells are lysed to release cAMP, then the cAMP detection solution, which contains protein kinase A, is added. The Kinase-Glo® Reagent is then added to terminate the PKA reaction and detect the remaining ATP via a luciferase reaction. Plates are read using a microplate-reading luminometer. Luminescence can be correlated to the cAMP concentrations by using a cAMP standard curve. The half-life for the luminescent signal is greater than 4 hours. This extended signal half-life eliminates the need for luminometers with reagent injectors and allows batch-mode processing of multiple plates.
Features - Benefits
Fast and Easy to Use:
Assay can be completed in approximately 45 minutes.
Homogeneous.
Two steps following lysis of cells.
Excellent Signal-to-Noise Ratios:
Best signal:background ratio of all the cAMP assays.
Signal:Background >200 (with cAMP), >15 (on cells).
Easily scalable to 1536-well plate formats and beyond.
Proven Luminescent Technology:
Powered by Ultra-Glo™ Recombinant Luciferase.
No interference by fluorescent compounds.
Non-radioactive.
Applications
High-throughput analysis of library compounds to identify modulators of GPCR activity.
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