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Practical and reliable FRET/FLIM pair of fluorescent proteins

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Author(s): Shcherbo Dmitry | Souslova Ekaterina | Goedhart Joachim | Chepurnykh Tatyana | Gaintzeva Anna | Shemiakina Irina | Gadella Theodorus | Lukyanov Sergey | Chudakov Dmitriy

Journal: BMC Biotechnology
ISSN 1472-6750

Volume: 9;
Issue: 1;
Start page: 24;
Date: 2009;
Original page

ABSTRACT
Abstract Background In spite of a great number of monomeric fluorescent proteins developed in the recent years, the reported fluorescent protein-based FRET pairs are still characterized by a number of disadvantageous features, complicating their use as reporters in cell biology and for high-throughput cell-based screenings. Results Here we screened some of the recently developed monomeric protein pairs to find the optimal combination, which would provide high dynamic range FRET changes, along with high pH- and photo-stability, fast maturation and bright fluorescence, and reliable detection in any fluorescent imaging system. Among generated FRET pairs, we have selected TagGFP-TagRFP, combining all the mentioned desirable characteristics. On the basis of this highly efficient FRET pair, we have generated a bright, high contrast, pH- and photo-stable apoptosis reporter, named CaspeR3 (Caspase 3 Reporter). Conclusion The combined advantages suggest that the TagGFP-TagRFP is one of the most efficient green/red couples available to date for FRET/FLIM analyses to monitor interaction of proteins of interest in living cells and to generate FRET-based sensors for various applications. CaspeR3 provides reliable detection of apoptosis, and should become a popular tool both for cell biology studies and high throughput screening assays.

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