| Title: | Revealing protein dynamics by photobleaching techniques |
| Address: | "Department of Molecular Biology, The Scripps Research Institute, La Jolla, CA, USA" |
| DOI: | 10.1385/1-59259-816-1:287 |
| ISSN/ISBN: | 1064-3745 (Print) 1064-3745 (Linking) |
| Abstract: | "Green fluorescent proteins (GFPs) are widely used tools to visualize proteins and study their intracellular distribution. One feature of working with GFP variants, photobleaching, has recently been combined with an older technique known as fluorescence recovery after photobleaching (FRAP) to study protein kinetics in vivo. During photobleaching, fluorochromes get destroyed irreversibly by repeated excitation with an intensive light source. When the photobleaching is applied to a restricted area or structure, recovery of fluorescence will be the result of active or passive diffusion from fluorescent molecules from unbleached surrounding areas. Fluorescence loss in photobleaching (FLIP) is a variant of FRAP where an area is bleached, and loss of fluorescence in surrounding areas is observed. FLIP can be used to study the dynamics of different pools of a protein or can show how a protein diffuses, or is transported through a cell or cellular structure. Here, we discuss these photobleaching fluorescent imaging techniques, illustrated with examples of these techniques applied to proteins of the Saccharomyces cerevisiae pheromone response MAPK pathway" |
| Keywords: | "Escherichia coli/genetics Fluorescence Recovery After Photobleaching/*methods Green Fluorescent Proteins Luminescent Proteins/biosynthesis/genetics *MAP Kinase Signaling System Microscopy, Confocal Mitogen-Activated Protein Kinases/genetics/metabolism Rec;" |
| Notes: | "Medlinevan Drogen, Frank Peter, Matthias eng Research Support, Non-U.S. Gov't 2004/06/03 Methods Mol Biol. 2004; 284:287-306. doi: 10.1385/1-59259-816-1:287" |
