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Luminescent Biosensors for Real-Time Monitoring of Intracellular cAMP

  • Brock F. BinkowskiEmail author
  • Frank Fan
  • Keith V. Wood
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 756)

Abstract

G-protein coupled, seven-transmembrane (7-TM) receptors (GPCRs) comprise a diverse class of signaling molecules involved in cellular physiology and pathology. In recent years, intracellular biosensors have been developed to monitor changes in intracellular cAMP in real time, facilitating studies on the mechanisms of GPCR activation and desensitization in living cells. However, methods based on fluorescence can show limitations in response dynamics together with being difficult to perform. Here we present the use of genetically encoded, luminescent biosensors that allow a facile, non-lytic assay format for monitoring cAMP dynamics in living cells.

Key words

cAMP Live cell assay GloSensor Biosensor G-protein-coupled receptor Inverse agonist 

References

  1. 1.
    Fan, F., Binkowski, B. F., Butler, B. L., Stecha, P. F., Lewis, M. K. and Wood, K. V. (2008) Novel genetically encoded biosensors using firefly luciferase. ACS Chem Biol 3, 346–51.PubMedCrossRefGoogle Scholar
  2. 2.
    Binkowski, B. F., Fan, F. and Wood, K. V. (2009) Live-cell luminescent assays for GPCR studies. Gen Eng Biotech 29, 30–31.Google Scholar
  3. 3.
    Binkowski, B. F., Fan, F. and Wood, K.V. (2009) Engineered luciferases for molecular sensing in living cells. Curr Opin Biotech 20, 14–18.PubMedCrossRefGoogle Scholar
  4. 4.
    Willoughby, D. and Cooper, D. M. F. (2008) Live-cell imaging of cAMP dynamics. Nat Methods 5, 29–36.PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2011

Authors and Affiliations

  • Brock F. Binkowski
    • 1
    Email author
  • Frank Fan
    • 1
  • Keith V. Wood
    • 1
  1. 1.Promega CorporationMadisonUSA

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