FRET and BRET-based biosensors in live cell compound screens.
Live cell compound screening with genetically encoded fluorescence or bioluminescence-based biosensors offers a potentially powerful approach to identify novel regulators of a signaling event of interest. In particular, compound screening in living cells has the added benefit that the entire signaling network remains intact, and thus the screen is not just against a single molecule of interest but against any molecule within the signaling network that may modulate the distinct signaling event reported by the biosensor in use. Furthermore, only molecules that are cell permeable or act at cell surface receptors will be identified as "hits," thus reducing further optimization of the compound in terms of cell penetration. Here we discuss a detailed protocol for using genetically encoded biosensors in living cells in a 96-well format for the execution of high throughput compound screens and the identification of small molecules which modulate a signaling event of interest.
Department of Pharmacology and Molecular Sciences, The Johns Hopkins University School of Medicine, Baltimore, MD, USA.
Katie Herbst
KH
Robinson
Jessica R
JR
Yang
Jin
J
Zhang
D015374:Biosensing Techniques / Q000379:methods*; D002452:Cell Count; D002470:Cell Survival; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D031541:Fluorescence Resonance Energy Transfer; D057809:HEK293 Cells; D006801:Humans; D008163:Luminescent Measurements; D014162:Transfection