title: FRET microscopy in the living cell: different approaches, strengths and weaknesses.
abstract: New imaging methodologies in quantitative fluorescence microscopy, such as Förster resonance energy transfer (FRET), have been developed in the last few years and are beginning to be extensively applied to biological problems. FRET is employed for the detection and quantification of protein interactions, and of biochemical activities. Herein, we review the different methods to measure FRET in microscopy, and more importantly, their strengths and weaknesses. In our opinion, fluorescence lifetime imaging microscopy (FLIM) is advantageous for detecting inter-molecular interactions quantitatively, the intensity ratio approach representing a valid and straightforward option for detecting intra-molecular FRET. Promising approaches in single molecule techniques and data analysis for quantitative and fast spatio-temporal protein-protein interaction studies open new avenues for FRET in biological research.
authors.affiliation: Department of Pediatrics, Infectious Diseases, Emory University, Atlanta, GA, USA.
authors.forename: Sergi
authors.initials: S
authors.lastname: Padilla-Parra
authors.forename: Marc
authors.initials: M
authors.lastname: Tramier
mesh_terms: D031541:Fluorescence Resonance Energy Transfer / Q000379:methods* / Q000639:trends; D049452:Green Fluorescent Proteins; D008856:Microscopy, Fluorescence / Q000379:methods* / Q000639:trends; D057054:Molecular Imaging / Q000379:methods*; D025941:Protein Interaction Mapping / Q000379:methods
chemical_list: D049452:Green Fluorescent Proteins