One-step split GFP staining for sensitive protein detection and localization in mammalian cells |
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Authors: | Kaddoum Lara Magdeleine Eddy Waldo Geoffrey S Joly Etienne Cabantous Stéphanie |
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Institution: | CNRS, Institute of Pharmacology and Structural Biology (IPBS), Toulouse, France. |
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Abstract: | Although epitope tags are useful to detect intracellular proteins and follow their localization with antibodies, background and nonspecific staining often remain problematic. We describe a simple assay based on the split GFP complementation system. Proteins tagged with the 15-amino acid GFP 11 fragment are detected with a solution of the recombinant nonfluorescent complementary GFP 1-10 fragment to reconstitute a fluorescent GFP. In contrast to antibody-based staining methods, this one-step assay presents high specificity and very low background of fluorescence, thus conferring higher signal-to-noise ratios. We demonstrate that this new application of the split GFP tagging system facilitates detection of proteins displaying various subcellular localizations using flow cytometry and microscopy analysis. |
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