We found a match
Your institution may have access to this item. Find your institution then sign in to continue.
- Title
Evolutionary optimization of fluorescent proteins for intracellular FRET.
- Authors
Nguyen, Annalee W; Daugherty, Patrick S
- Abstract
Fluorescent proteins that exhibit Forster resonance energy transfer (FRET) have made a strong impact as they enable measurement of molecular-scale distances through changes in fluorescence. FRET-based approaches have enabled otherwise intractable measurements of molecular concentrations, binding interactions and catalytic activity, but are limited by the dynamic range and sensitivity of the donor-acceptor pair. To address this problem, we applied a quantitative evolutionary strategy using fluorescence-activated cell sorting to optimize a cyan-yellow fluorescent protein pair for FRET. The resulting pair, CyPet-YPet, exhibited a 20-fold ratiometric FRET signal change, as compared to threefold for the parental pair. The optimized FRET pair enabled high-throughput flow cytometric screening of cells undergoing caspase-3-dependent apoptosis. The CyPet-YPet energy transfer pair provides substantially improved sensitivity and dynamic range for a broad range of molecular imaging and screening applications.
- Publication
Nature biotechnology, 2005, Vol 23, Issue 3, p355
- ISSN
1087-0156
- Publication type
Research
- DOI
10.1038/nbt1066