ArticlePloS one2015
Fourth-generation epac-based FRET sensors for cAMP feature exceptional brightness, photostability and dynamic range: characterization of dedicated sensors for FLIM, for ratiometry and with high affinity.
Article in PloS one, 2015. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 190 papers.
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190 citing papers in PubMed.
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- Sensitive and Selective Next-Generation FRET-based PKA Biosensors.bioRxiv : the preprint server for biology · 2026Article
- Combined excitation scan-based hyperspectral imaging and digital deconvolution for the assessment of the kinetic and spatial distributions of cAMP signals.Proceedings of SPIE--the International Society for Optical Engineering · 2026Article
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130 more citing papers are in PubMed but not listed here.
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5 authors.
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Abstract
Epac-based FRET sensors have been widely used for the detection of cAMP concentrations in living cells. Originally developed by us as well as others, we have since then reported several important optimizations that make these sensors favourite among many cell biologists. We here report cloning and characterization of our fourth generation of cAMP sensors, which feature outstanding photostability, dynamic range and signal-to-noise ratio. The design is based on mTurquoise2, currently the brightest and most bleaching-resistant donor, and a new acceptor cassette that consists of a tandem of two cp173Venus fluorophores. We also report variants with a single point mutation, Q270E, in the Epac moiety, which decreases the dissociation constant of cAMP from 9.5 to 4 μM, and thus increases the affinity ~ 2.5-fold. Finally, we also prepared and characterized dedicated variants with non-emitting (dark) acceptors for single-wavelength FLIM acquisition that display an exceptional near-doubling of fluorescence lifetime upon saturation of cAMP levels. We believe this generation of cAMP outperforms all other sensors and therefore recommend these sensors for all future studies.
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