Format

Send to

Choose Destination
Biophys J. 2006 Dec 15;91(12):L99-L101. Epub 2006 Oct 13.

Cerulean, Venus, and VenusY67C FRET reference standards.

Author information

1
Laboratory of Molecular Physiology, National Institute on Alcohol Abuse and Alcoholism, Bethesda, MD, USA.

Abstract

Förster's resonance energy transfer (FRET) can be used to study protein-protein interactions in living cells. Numerous methods to measure FRET have been devised and implemented; however, the accuracy of these methods is unknown, which makes interpretation of FRET efficiency values difficult if not impossible. This problem exists due to the lack of standards with known FRET efficiencies that can be used to validate FRET measurements. The advent of spectral variants of green fluorescent protein and easy access to cell transfection technology suggests a simple solution to this problem: the development of genetic constructs with known FRET efficiencies that can be replicated with high fidelity and freely distributed. In this study, fluorescent protein constructs with progressively larger separation distances between donors and acceptors were generated and FRET efficiencies were measured using fluorescence lifetime spectroscopy, sensitized acceptor emission, and spectral imaging. Since the results from each method were in good agreement, the FRET efficiency value of each construct could be determined with high accuracy and precision, thereby justifying their use as standards.

PMID:
17040988
PMCID:
PMC1779932
DOI:
10.1529/biophysj.106.096206
[Indexed for MEDLINE]
Free PMC Article

Supplemental Content

Full text links

Icon for Elsevier Science Icon for PubMed Central
Loading ...
Support Center