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Optics Express

Optics Express

  • Editor: C. Martijn de Sterke
  • Vol. 18, Iss. 18 — Aug. 30, 2010
  • pp: 18839–18851

Quantitative FRET measurement by high-speed fluorescence excitation and emission spectrometer

Jing Yuan, Leilei Peng, Brett E. Bouma, and Guillermo J. Tearney  »View Author Affiliations

Optics Express, Vol. 18, Issue 18, pp. 18839-18851 (2010)

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Förster resonance energy transfer (FRET) is an important method in studying biochemistry reactions. But quantifying FRET rapidly is difficult to do because of crosstalk between free donor, free acceptor and FRET fluorescent signals when only excitation or emission property of a FRET sample is measured. If FRET is studied with excitation-emission matrix (EEM) measurements, because the fluorescence intensity maxima of donor, acceptor, and FRET emissions occupy different regions within the EEM, FRET fluorescence can be easily separated out by linear unmixing. In this paper, we report a novel high-speed Fourier Fluorescence Excitation Emission spectrometer, which simultaneously measures three projections of EEM from a FRET sample, which are excitation, emission and excitation-emission cross-correlation spectra. We demonstrate that these three EEM projections can be measured and unmixed in approximately 1 ms to provide rapid quantitative FRET in the presence of free donors and acceptors. The system can be utilized to enable real-time biochemistry reaction studies.

© 2010 OSA

OCIS Codes
(300.6280) Spectroscopy : Spectroscopy, fluorescence and luminescence
(300.6300) Spectroscopy : Spectroscopy, Fourier transforms

ToC Category:

Original Manuscript: May 14, 2010
Revised Manuscript: July 11, 2010
Manuscript Accepted: July 15, 2010
Published: August 19, 2010

Virtual Issues
Vol. 5, Iss. 13 Virtual Journal for Biomedical Optics

Jing Yuan, Leilei Peng, Brett E. Bouma, and Guillermo J. Tearney, "Quantitative FRET measurement by high-speed fluorescence excitation and emission spectrometer," Opt. Express 18, 18839-18851 (2010)

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