cAMP-Glo™ Max Assay

The cAMP-Glo™ Max Assay is a homogeneous, bioluminescent and high-throughput assay to measure cyclic AMP (cAMP) levels in cells. Compounds that modulate GPCRs coupled with adenylate cyclase typically alter intracellular cAMP levels. The cAMP-Glo™ Max Assay monitors cAMP levels in cells in response to the effect of agonists, antagonists or test compounds on G protein-coupled receptors (GPCRs). 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.

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The cAMP-Glo™ Max Assay is a homogeneous, bioluminescent and high-throughput assay to measure cyclic AMP (cAMP) levels in cells. Compounds that modulate GPCRs coupled with adenylate cyclase typically alter intracellular cAMP levels. The cAMP-Glo™ Max Assay monitors cAMP levels in cells in response to the effect of agonists, antagonists or test compounds on G protein-coupled receptors (GPCRs). 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.

This improved version combines the lysis and cAMP reaction buffers into the cAMP-Glo™ ONE Buffer. This new format streamlines the protocol and reduces the time needed to complete the assay. The new ONE Buffer is supplied at a 5X concentration, which provides increased flexibility for starting cell culture volumes.

The cAMP-Glo™ Max 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, and the cAMP released stimulates protein kinase A in the reagent (Figure 1). 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. The half-life for the luminescent signal is greater than 4 hours allowing ample time to read the plates and eliminates the need for luminometers with reagent injectors.

Features - Benefits

• Fast and Easy to Use:
  Improved—Lysis and cAMP detection steps combined (cAMP-Glo™ ONE Buffer).
  ONE Buffer—5X concentration provides better flexibility for starting cell culture volumes.
  Assay can be completed in approximately 30 minutes.
• 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.
• Easy quantitation of cAMP levels in broad range of cell types.

Notes

Cat.# V1681 contains sufficient reagents for 2 plates (96- or 384-well format). Cat.# V1682 contains sufficient reagents for 20 plates (96- or 384-well format). Cat.# V1683 contains sufficient reagents for 200 plates (96- or 384-well format).

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