GloSensor™ Technology for Second Messenger (cAMP/cGMP) and Protease Detection
Live Cell Biosensors for Real-Time Detection of Intracellular Signaling
GloSensor™ technology provides a platform of flexible luciferase-based biosensors for real-time (kinetic) detection of signaling events in live cells with great sensitivity, linearity and specificity. The platform is based on genetically encoded mutant forms of Photinus pyralis that are designed to change structural confirmation based on sensing specific intracellular signaling events. In the open confirmation the biosensor is inactive. Upon detection of specific signaling events conformational change of the sensor occurs, resulting in the closed, active form of luciferase and large increases in light output.
Biosensors for cAMP/cGMP Detection
The cAMP biosensors contain a cAMP binding domain in the hinge region of the protein. In the presence of cAMP second messenger the biosensor changes confirmation to the active form of the protein. The live-cell, non-lytic assay format allows kinetic measurements of cAMP accumulation or turnover in living cells, enabling sensitive assays of GPCRs that signal through cAMP. We also have biosensors for the specific detection of cGMP.
Biosensors for Protease Detection
Biosensors for protease activity detection are built to contain a specific protease cleavage site that inhibits the closed, active formation of the sensor. Cleavage by the protease relieves this constraint, creating the active form of the sensor and allowing real-time measures of specific protease activity in vitro or in live cell models.
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The overview presents the GloSensor™ technology concept, recent performance data and a comparison against time-resolved fluorescence and an immunoassay.
References for GloSensor™ Technology
Fan, F. Binkowski, B. et al. (2008) Novel genetically encoded biosensors using firefly luciferase. ACS Chem. Biol. 3, 346–51.
Binkowski, B. et al. (2009) Engineered luciferases for molecular sensing in living cells. Curr. Opin. Biotechnol. 20 14–18. (DOI:10.1016/j.copbio.2009.02.013)