Localization of protein-protein interactions in live cells using confocal and spectral imaging FRET microscopy.
Microscopy has become an essential tool for cellular protein investigations. The development of new fluorescent markers such as green fluorescent proteins generated substantial opportunities to monitor protein-protein interactions qualitatively and quantitatively using advanced fluorescence microscope techniques including wide-field, confocal, multiphoton, spectral imaging, lifetime, and correlation spectroscopy. The specific aims of the investigation of protein dynamics in live specimens dictate the selection of the microscope methodology. In this article confocal and spectral imaging methods to monitor the dimerization of alpha enhancer binding protein (C/EBPalpha) in the pituitary GHFT1-5 living cell nucleus have been described. Also outline are issues involved in protein imaging using light microscopy techniques and the advantages of lifetime imaging of protein-protein interactions.
Chen
Ye
Y
Department of Biology, W M Keck Center for Cellular Imaging, Gilmer Hall, University of Virginia, Charlottesville, VA 22904, USA.
Periasamy
Ammasi
A
D000818:Animals; D022763:CCAAT-Enhancer-Binding Protein-alpha / Q000032:analysis; D002467:Cell Nucleus / Q000737:chemistry; D002478:Cells, Cultured; D019281:Dimerization; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods*; D049452:Green Fluorescent Proteins / Q000032:analysis; D051379:Mice; D018613:Microscopy, Confocal / Q000379:methods*; D010902:Pituitary Gland / Q000166:cytology; D025941:Protein Interaction Mapping / Q000379:methods*
D022763:CCAAT-Enhancer-Binding Protein-alpha; D049452:Green Fluorescent Proteins