title: A luminescent biosensor with increased dynamic range for intracellular cAMP.
abstract: The second messenger cAMP is a key mediator of signal transduction following activation of G-protein coupled receptors. Investigations on Gs-coupled receptors would benefit from a second messenger assay that allows continuous monitoring of kinetic changes in cAMP concentration over a broad dynamic range. To accomplish this, we have evolved a luminescent biosensor for cAMP to better encompass the physiological concentration ranges present in living cells. When compared to an immunoassay, the evolved biosensor construct was able to accurately track both the magnitude and kinetics of cAMP change using a far less labor intensive format. We demonstrate the utility of this construct to detect a broad range of receptor activity, together with showing suitability for use in high-throughput screening.
authors.forename: Brock F
authors.initials: BF
authors.lastname: Binkowski
authors.forename: Braeden L
authors.initials: BL
authors.lastname: Butler
authors.forename: Peter F
authors.initials: PF
authors.lastname: Stecha
authors.forename: Christopher T
authors.initials: CT
authors.lastname: Eggers
authors.forename: Paul
authors.initials: P
authors.lastname: Otto
authors.forename: Kris
authors.initials: K
authors.lastname: Zimmerman
authors.forename: Gediminas
authors.initials: G
authors.lastname: Vidugiris
authors.forename: Monika G
authors.initials: MG
authors.lastname: Wood
authors.forename: Lance P
authors.initials: LP
authors.lastname: Encell
authors.forename: Frank
authors.initials: F
authors.lastname: Fan
authors.forename: Keith V
authors.initials: KV
authors.lastname: Wood
mesh_terms: D015374:Biosensing Techniques / Q000379:methods*; D000242:Cyclic AMP / Q000032:analysis* / Q000737:chemistry; D057809:HEK293 Cells; D057166:High-Throughput Screening Assays / Q000379:methods*; D006801:Humans; D007700:Kinetics; D008163:Luminescent Measurements; D008958:Models, Molecular; D043562:Receptors, G-Protein-Coupled / Q000378:metabolism; D015398:Signal Transduction; D013816:Thermodynamics
chemical_list: D043562:Receptors, G-Protein-Coupled; D000242:Cyclic AMP