Abstract

Frequency domain fluorescence lifetime imaging is a powerful technique that enables the observation of subtle changes in the molecular environment of a fluorescent probe. This technique works by measuring the phase delay between the optical emission and excitation of fluorophores as a function of modulation frequency. However, high-resolution measurements are time consuming, as the excitation modulation frequency must be swept, and faster low-resolution measurements at a single frequency are prone to large errors. Here, we present a low cost optical system for applications in real-time confocal lifetime imaging, which measures the phase vs. frequency spectrum without sweeping. Deemed Lifetime Imaging using Frequency-multiplexed Excitation (LIFE), this technique uses a digitally-synthesized radio frequency comb to drive an acousto-optic deflector, operated in a cat’s-eye configuration, to produce a single laser excitation beam modulated at multiple beat frequencies. We demonstrate simultaneous fluorescence lifetime measurements at 10 frequencies over a bandwidth of 48 MHz, enabling high speed frequency domain lifetime analysis of single- and multi-component sample mixtures.

© 2014 Optical Society of America

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References

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2014 (2)

D. R. Yankelevich, D. Ma, J. Liu, Y. Sun, Y. Sun, J. Bec, D. S. Elson, and L. Marcu, “Design and evaluation of a device for fast multispectral time-resolved fluorescence spectroscopy and imaging,” Rev. Sci. Instrum. 85(3), 034303 (2014).
[Crossref] [PubMed]

A. S. Kristoffersen, S. R. Erga, B. Hamre, and Ø. Frette, “Testing fluorescence lifetime standards using two-photon excitation and time-domain instrumentation: rhodamine B, coumarin 6 and lucifer yellow,” J. Fluoresc. 24(4), 1015–1024 (2014).
[Crossref] [PubMed]

2013 (1)

E. D. Diebold, B. W. Buckley, D. R. Gossett, and B. Jalali, “Digitally synthesized beat frequency multiplexing for sub-millisecond fluorescence microscopy,” Nat. Photonics 7(10), 806–810 (2013).
[Crossref]

2012 (2)

J. Ge, C. Kuang, S.-S. Lee, and F.-J. Kao, “Fluorescence lifetime imaging with pulsed diode laser enabled stimulated emission,” Opt. Express 20(27), 28216–28221 (2012).
[Crossref] [PubMed]

M. D. Lesoine, S. Bose, J. W. Petrich, and E. A. Smith, “Supercontinuum stimulated emission depletion fluorescence lifetime imaging,” J. Phys. Chem. B 116(27), 7821–7826 (2012).
[Crossref] [PubMed]

2011 (1)

2009 (1)

2008 (2)

G.-J. Kremers, E. B. van Munster, J. Goedhart, and T. W. J. Gadella, “Quantitative lifetime unmixing of multiexponentially decaying fluorophores using single-frequency fluorescence lifetime imaging microscopy,” Biophys. J. 95(1), 378–389 (2008).
[Crossref] [PubMed]

H. Murakoshi, S.-J. Lee, and R. Yasuda, “Highly sensitive and quantitative FRET-FLIM imaging in single dendritic spines using improved non-radiative YFP,” Brain Cell Biol. 36(1-4), 31–42 (2008).
[Crossref] [PubMed]

2007 (3)

R. Cicchi, D. Massi, S. Sestini, P. Carli, V. De Giorgi, T. Lotti, and F. S. Pavone, “Multidimensional non-linear laser imaging of Basal Cell Carcinoma,” Opt. Express 15(16), 10135–10148 (2007).
[Crossref] [PubMed]

E. B. van Munster, J. Goedhart, G. J. Kremers, E. M. Manders, and T. W. Gadella, “Combination of a spinning disc confocal unit with frequency-domain fluorescence lifetime imaging microscopy,” Cytometry A 71(4), 207–214 (2007).
[Crossref] [PubMed]

N. Boens, W. Qin, N. Basaric, J. Hofkens, and M. Ameloot, “Fluorescence Lifetime Standards for Time and Frequency Domain Fluorescence spectroscopy,” Analytical Chem. 79, 2137–2149 (2007).

2006 (2)

K. Shim and B. Kim, “Simple frequency-domain fluorescence-lifetime measurement system using violet laser diode,” J. Korean Phys. Soc. 49, 647–651 (2006).

A. D. Elder, J. H. Frank, J. Swartling, X. Dai, and C. F. Kaminski, “Calibration of a wide-field frequency-domain fluorescence lifetime microscopy system using light emitting diodes as light sources,” J. Microsc. 224(2), 166–180 (2006).
[Crossref] [PubMed]

2005 (2)

2004 (3)

W. Becker, A. Bergmann, G. Biscotti, K. Koenig, I. Riemann, L. Kelbauskas, and C. Biskup, “High-speed FLIM data acquisition by time-correlated single-photon counting,” Multiphot. Microsc. Biomed. Sci. IV 5323, 27–35 (2004).
[Crossref]

W. Becker, A. Bergmann, M. A. Hink, K. König, K. Benndorf, and C. Biskup, “Fluorescence lifetime imaging by time-correlated single-photon counting,” Microsc. Res. Tech. 63(1), 58–66 (2004).
[Crossref] [PubMed]

R. R. Duncan, A. Bergmann, M. A. Cousin, D. K. Apps, and M. J. Shipston, “Multi-dimensional time-correlated single photon counting (TCSPC) fluorescence lifetime imaging microscopy (FLIM) to detect FRET in cells,” J. Microsc. 215(1), 1–12 (2004).
[Crossref] [PubMed]

2003 (1)

B. J. Bacskai, J. Skoch, G. A. Hickey, R. Allen, and B. T. Hyman, “Fluorescence resonance energy transfer determinations using multiphoton fluorescence lifetime imaging microscopy to characterize amyloid-beta plaques,” J. Biomed. Opt. 8(3), 368–375 (2003).
[Crossref] [PubMed]

2002 (1)

H. C. Gerritsen, M. A. Asselbergs, A. V. Agronskaia, and W. G. Van Sark, “Fluorescence lifetime imaging in scanning microscopes: acquisition speed, photon economy and lifetime resolution,” J. Microsc. 206(3), 218–224 (2002).
[Crossref] [PubMed]

2000 (3)

A. Squire, P. J. Verveer, and P. I. H. Bastiaens, “Multiple frequency fluorescence lifetime imaging microscopy,” J. Microscopy 197, 136–149 (2000).

S. J. Lassiter, W. Stryjewski, B. L. Legendre, R. Erdmann, M. Wahl, J. Wurm, R. Peterson, L. Middendorf, and S. A. Soper, “Time-resolved fluorescence imaging of slab gels for lifetime base-calling in DNA sequencing applications,” Anal. Chem. 72(21), 5373–5382 (2000).
[Crossref] [PubMed]

M. Neumann, D. Herten, A. Dietrich, J. Wolfrum, and M. Sauer, “Capillary array scanner for time-resolved detection and identification of fluorescently labelled DNA fragments,” J. Chromatography A. 871, 299–310 (2000).

1999 (2)

M. B. Smalley, J. M. Shaver, and L. B. Mcgown, “On-the-fly fluorescence lifetime detection in HPLC using a multiharmonic Fourier transform phase-modulation spectrofluorometer,” Analytical Chem. 653466–3472 (1999).

D. Magde, G. E. Rojas, and P. G. Seybold, “Solvent dependence of the fluorescence lifetimes of Xanthene dyes,” Photochem. Photobiol. 70, 737–744 (1999).

1998 (2)

A. N. Watkins, C. M. Ingersoll, G. A. Baker, and F. V. Bright, “A parallel multiharmonic frequency-domain fluorometer for measuring excited-state decay kinetics following one-, two-, or three-photon excitation,” Anal. Chem. 70(16), 3384–3396 (1998).
[Crossref] [PubMed]

H. He, B. K. Nunnally, L. C. Li, and L. B. McGown, “On-the-fly fluorescence lifetime detection of dye-labeled DNA primers for multiplex analysis,” Anal. Chem. 70(16), 3413–3418 (1998).
[Crossref] [PubMed]

1997 (1)

R. Cubeddu, G. Canti, A. Pifferi, P. Taroni, and G. Valentini, “Fluorescence lifetime imaging of experimental tumors in hematoporphyrin derivative-sensitized mice,” Photochem. Photobiol. 66(2), 229–236 (1997).
[Crossref] [PubMed]

1993 (1)

T. W. J. Gadella, T. M. Jovin, and R. M. Clegg, “Fluorescence lifetime imaging microscopy (FLIM): Spatial resolution of microstructures on the nanosecond time scale,” Biophys. Chem. 48(2), 221–239 (1993).
[Crossref]

1992 (1)

W. Baumler, A. X. Schmalzl, A. Penzkofer, N. Fakultat, I. I. Physik, and U. Regensburg, “Fluorescence decay studies applying a cw femtosecond dye laser pumped ungated inverse time-correlated single photon counting system,” Meas. Sci. Technol. 3, 384–393 (1992).

1991 (1)

A. S. Verkman, M. Armijo, and K. Fushimi, “Construction and evaluation of a frequency-domain epifluorescence microscope for lifetime and anisotropy decay measurements in subcellular domains,” Biophys. Chem. 40(1), 117–125 (1991).
[Crossref] [PubMed]

1984 (1)

E. Gratton, M. Limkeman, J. R. Lakowicz, B. P. Maliwal, H. Cherek, and G. Laczko, “Resolution of mixtures of fluorophores using variable-frequency phase and modulation data,” Biophys. J. 46(4), 479–486 (1984).
[Crossref] [PubMed]

1983 (1)

G. Ide, “Fluorescence lifetime resolution with phase fluorometry,” Rev. Sci. Instrum. 54(7), 841 (1983).
[Crossref]

1981 (1)

E. H. J. Young and S.-K. Yao, “Design considerations for acousto-optic devices,” Proc. IEEE 69, 54–64 (1981).
[Crossref]

Agronskaia, A. V.

H. C. Gerritsen, M. A. Asselbergs, A. V. Agronskaia, and W. G. Van Sark, “Fluorescence lifetime imaging in scanning microscopes: acquisition speed, photon economy and lifetime resolution,” J. Microsc. 206(3), 218–224 (2002).
[Crossref] [PubMed]

Allen, R.

B. J. Bacskai, J. Skoch, G. A. Hickey, R. Allen, and B. T. Hyman, “Fluorescence resonance energy transfer determinations using multiphoton fluorescence lifetime imaging microscopy to characterize amyloid-beta plaques,” J. Biomed. Opt. 8(3), 368–375 (2003).
[Crossref] [PubMed]

Ameloot, M.

N. Boens, W. Qin, N. Basaric, J. Hofkens, and M. Ameloot, “Fluorescence Lifetime Standards for Time and Frequency Domain Fluorescence spectroscopy,” Analytical Chem. 79, 2137–2149 (2007).

Apps, D. K.

R. R. Duncan, A. Bergmann, M. A. Cousin, D. K. Apps, and M. J. Shipston, “Multi-dimensional time-correlated single photon counting (TCSPC) fluorescence lifetime imaging microscopy (FLIM) to detect FRET in cells,” J. Microsc. 215(1), 1–12 (2004).
[Crossref] [PubMed]

Armijo, M.

A. S. Verkman, M. Armijo, and K. Fushimi, “Construction and evaluation of a frequency-domain epifluorescence microscope for lifetime and anisotropy decay measurements in subcellular domains,” Biophys. Chem. 40(1), 117–125 (1991).
[Crossref] [PubMed]

Asselbergs, M. A.

H. C. Gerritsen, M. A. Asselbergs, A. V. Agronskaia, and W. G. Van Sark, “Fluorescence lifetime imaging in scanning microscopes: acquisition speed, photon economy and lifetime resolution,” J. Microsc. 206(3), 218–224 (2002).
[Crossref] [PubMed]

Auksorius, E.

Bacskai, B. J.

B. J. Bacskai, J. Skoch, G. A. Hickey, R. Allen, and B. T. Hyman, “Fluorescence resonance energy transfer determinations using multiphoton fluorescence lifetime imaging microscopy to characterize amyloid-beta plaques,” J. Biomed. Opt. 8(3), 368–375 (2003).
[Crossref] [PubMed]

Baker, G. A.

A. N. Watkins, C. M. Ingersoll, G. A. Baker, and F. V. Bright, “A parallel multiharmonic frequency-domain fluorometer for measuring excited-state decay kinetics following one-, two-, or three-photon excitation,” Anal. Chem. 70(16), 3384–3396 (1998).
[Crossref] [PubMed]

Basaric, N.

N. Boens, W. Qin, N. Basaric, J. Hofkens, and M. Ameloot, “Fluorescence Lifetime Standards for Time and Frequency Domain Fluorescence spectroscopy,” Analytical Chem. 79, 2137–2149 (2007).

Bastiaens, P. I. H.

A. Squire, P. J. Verveer, and P. I. H. Bastiaens, “Multiple frequency fluorescence lifetime imaging microscopy,” J. Microscopy 197, 136–149 (2000).

Baumler, W.

W. Baumler, A. X. Schmalzl, A. Penzkofer, N. Fakultat, I. I. Physik, and U. Regensburg, “Fluorescence decay studies applying a cw femtosecond dye laser pumped ungated inverse time-correlated single photon counting system,” Meas. Sci. Technol. 3, 384–393 (1992).

Bec, J.

D. R. Yankelevich, D. Ma, J. Liu, Y. Sun, Y. Sun, J. Bec, D. S. Elson, and L. Marcu, “Design and evaluation of a device for fast multispectral time-resolved fluorescence spectroscopy and imaging,” Rev. Sci. Instrum. 85(3), 034303 (2014).
[Crossref] [PubMed]

Becker, W.

W. Becker, A. Bergmann, G. Biscotti, K. Koenig, I. Riemann, L. Kelbauskas, and C. Biskup, “High-speed FLIM data acquisition by time-correlated single-photon counting,” Multiphot. Microsc. Biomed. Sci. IV 5323, 27–35 (2004).
[Crossref]

W. Becker, A. Bergmann, M. A. Hink, K. König, K. Benndorf, and C. Biskup, “Fluorescence lifetime imaging by time-correlated single-photon counting,” Microsc. Res. Tech. 63(1), 58–66 (2004).
[Crossref] [PubMed]

Benndorf, K.

W. Becker, A. Bergmann, M. A. Hink, K. König, K. Benndorf, and C. Biskup, “Fluorescence lifetime imaging by time-correlated single-photon counting,” Microsc. Res. Tech. 63(1), 58–66 (2004).
[Crossref] [PubMed]

Bergmann, A.

W. Becker, A. Bergmann, M. A. Hink, K. König, K. Benndorf, and C. Biskup, “Fluorescence lifetime imaging by time-correlated single-photon counting,” Microsc. Res. Tech. 63(1), 58–66 (2004).
[Crossref] [PubMed]

R. R. Duncan, A. Bergmann, M. A. Cousin, D. K. Apps, and M. J. Shipston, “Multi-dimensional time-correlated single photon counting (TCSPC) fluorescence lifetime imaging microscopy (FLIM) to detect FRET in cells,” J. Microsc. 215(1), 1–12 (2004).
[Crossref] [PubMed]

W. Becker, A. Bergmann, G. Biscotti, K. Koenig, I. Riemann, L. Kelbauskas, and C. Biskup, “High-speed FLIM data acquisition by time-correlated single-photon counting,” Multiphot. Microsc. Biomed. Sci. IV 5323, 27–35 (2004).
[Crossref]

Biscotti, G.

W. Becker, A. Bergmann, G. Biscotti, K. Koenig, I. Riemann, L. Kelbauskas, and C. Biskup, “High-speed FLIM data acquisition by time-correlated single-photon counting,” Multiphot. Microsc. Biomed. Sci. IV 5323, 27–35 (2004).
[Crossref]

Biskup, C.

W. Becker, A. Bergmann, G. Biscotti, K. Koenig, I. Riemann, L. Kelbauskas, and C. Biskup, “High-speed FLIM data acquisition by time-correlated single-photon counting,” Multiphot. Microsc. Biomed. Sci. IV 5323, 27–35 (2004).
[Crossref]

W. Becker, A. Bergmann, M. A. Hink, K. König, K. Benndorf, and C. Biskup, “Fluorescence lifetime imaging by time-correlated single-photon counting,” Microsc. Res. Tech. 63(1), 58–66 (2004).
[Crossref] [PubMed]

Boens, N.

N. Boens, W. Qin, N. Basaric, J. Hofkens, and M. Ameloot, “Fluorescence Lifetime Standards for Time and Frequency Domain Fluorescence spectroscopy,” Analytical Chem. 79, 2137–2149 (2007).

Bose, S.

M. D. Lesoine, S. Bose, J. W. Petrich, and E. A. Smith, “Supercontinuum stimulated emission depletion fluorescence lifetime imaging,” J. Phys. Chem. B 116(27), 7821–7826 (2012).
[Crossref] [PubMed]

Bright, F. V.

A. N. Watkins, C. M. Ingersoll, G. A. Baker, and F. V. Bright, “A parallel multiharmonic frequency-domain fluorometer for measuring excited-state decay kinetics following one-, two-, or three-photon excitation,” Anal. Chem. 70(16), 3384–3396 (1998).
[Crossref] [PubMed]

Buckley, B. W.

E. D. Diebold, B. W. Buckley, D. R. Gossett, and B. Jalali, “Digitally synthesized beat frequency multiplexing for sub-millisecond fluorescence microscopy,” Nat. Photonics 7(10), 806–810 (2013).
[Crossref]

Canti, G.

R. Cubeddu, G. Canti, A. Pifferi, P. Taroni, and G. Valentini, “Fluorescence lifetime imaging of experimental tumors in hematoporphyrin derivative-sensitized mice,” Photochem. Photobiol. 66(2), 229–236 (1997).
[Crossref] [PubMed]

Carli, P.

Cherek, H.

E. Gratton, M. Limkeman, J. R. Lakowicz, B. P. Maliwal, H. Cherek, and G. Laczko, “Resolution of mixtures of fluorophores using variable-frequency phase and modulation data,” Biophys. J. 46(4), 479–486 (1984).
[Crossref] [PubMed]

Chuang, F. S.

Cicchi, R.

Clegg, R. M.

T. W. J. Gadella, T. M. Jovin, and R. M. Clegg, “Fluorescence lifetime imaging microscopy (FLIM): Spatial resolution of microstructures on the nanosecond time scale,” Biophys. Chem. 48(2), 221–239 (1993).
[Crossref]

Courtney, P.

Cousin, M. A.

R. R. Duncan, A. Bergmann, M. A. Cousin, D. K. Apps, and M. J. Shipston, “Multi-dimensional time-correlated single photon counting (TCSPC) fluorescence lifetime imaging microscopy (FLIM) to detect FRET in cells,” J. Microsc. 215(1), 1–12 (2004).
[Crossref] [PubMed]

Cubeddu, R.

R. Cubeddu, G. Canti, A. Pifferi, P. Taroni, and G. Valentini, “Fluorescence lifetime imaging of experimental tumors in hematoporphyrin derivative-sensitized mice,” Photochem. Photobiol. 66(2), 229–236 (1997).
[Crossref] [PubMed]

Dai, X.

A. D. Elder, J. H. Frank, J. Swartling, X. Dai, and C. F. Kaminski, “Calibration of a wide-field frequency-domain fluorescence lifetime microscopy system using light emitting diodes as light sources,” J. Microsc. 224(2), 166–180 (2006).
[Crossref] [PubMed]

De Giorgi, V.

Diebold, E. D.

E. D. Diebold, B. W. Buckley, D. R. Gossett, and B. Jalali, “Digitally synthesized beat frequency multiplexing for sub-millisecond fluorescence microscopy,” Nat. Photonics 7(10), 806–810 (2013).
[Crossref]

Dietrich, A.

M. Neumann, D. Herten, A. Dietrich, J. Wolfrum, and M. Sauer, “Capillary array scanner for time-resolved detection and identification of fluorescently labelled DNA fragments,” J. Chromatography A. 871, 299–310 (2000).

Donley, E. A.

E. A. Donley, T. P. Heavner, F. Levi, M. O. Tataw, and S. R. Jefferts, “Double-pass acousto-optic modulator system,” Rev. Sci. Instrum. 76(6), 063112 (2005).
[Crossref]

Duncan, R. R.

R. R. Duncan, A. Bergmann, M. A. Cousin, D. K. Apps, and M. J. Shipston, “Multi-dimensional time-correlated single photon counting (TCSPC) fluorescence lifetime imaging microscopy (FLIM) to detect FRET in cells,” J. Microsc. 215(1), 1–12 (2004).
[Crossref] [PubMed]

Dunsby, C.

Elder, A. D.

A. D. Elder, J. H. Frank, J. Swartling, X. Dai, and C. F. Kaminski, “Calibration of a wide-field frequency-domain fluorescence lifetime microscopy system using light emitting diodes as light sources,” J. Microsc. 224(2), 166–180 (2006).
[Crossref] [PubMed]

Elson, D. S.

Erdmann, R.

S. J. Lassiter, W. Stryjewski, B. L. Legendre, R. Erdmann, M. Wahl, J. Wurm, R. Peterson, L. Middendorf, and S. A. Soper, “Time-resolved fluorescence imaging of slab gels for lifetime base-calling in DNA sequencing applications,” Anal. Chem. 72(21), 5373–5382 (2000).
[Crossref] [PubMed]

Erga, S. R.

A. S. Kristoffersen, S. R. Erga, B. Hamre, and Ø. Frette, “Testing fluorescence lifetime standards using two-photon excitation and time-domain instrumentation: rhodamine B, coumarin 6 and lucifer yellow,” J. Fluoresc. 24(4), 1015–1024 (2014).
[Crossref] [PubMed]

Fakultat, N.

W. Baumler, A. X. Schmalzl, A. Penzkofer, N. Fakultat, I. I. Physik, and U. Regensburg, “Fluorescence decay studies applying a cw femtosecond dye laser pumped ungated inverse time-correlated single photon counting system,” Meas. Sci. Technol. 3, 384–393 (1992).

Farwell, D. G.

Frank, J. H.

A. D. Elder, J. H. Frank, J. Swartling, X. Dai, and C. F. Kaminski, “Calibration of a wide-field frequency-domain fluorescence lifetime microscopy system using light emitting diodes as light sources,” J. Microsc. 224(2), 166–180 (2006).
[Crossref] [PubMed]

French, P. M. W.

Frette, Ø.

A. S. Kristoffersen, S. R. Erga, B. Hamre, and Ø. Frette, “Testing fluorescence lifetime standards using two-photon excitation and time-domain instrumentation: rhodamine B, coumarin 6 and lucifer yellow,” J. Fluoresc. 24(4), 1015–1024 (2014).
[Crossref] [PubMed]

Fushimi, K.

A. S. Verkman, M. Armijo, and K. Fushimi, “Construction and evaluation of a frequency-domain epifluorescence microscope for lifetime and anisotropy decay measurements in subcellular domains,” Biophys. Chem. 40(1), 117–125 (1991).
[Crossref] [PubMed]

Gadella, T. W.

E. B. van Munster, J. Goedhart, G. J. Kremers, E. M. Manders, and T. W. Gadella, “Combination of a spinning disc confocal unit with frequency-domain fluorescence lifetime imaging microscopy,” Cytometry A 71(4), 207–214 (2007).
[Crossref] [PubMed]

Gadella, T. W. J.

G.-J. Kremers, E. B. van Munster, J. Goedhart, and T. W. J. Gadella, “Quantitative lifetime unmixing of multiexponentially decaying fluorophores using single-frequency fluorescence lifetime imaging microscopy,” Biophys. J. 95(1), 378–389 (2008).
[Crossref] [PubMed]

T. W. J. Gadella, T. M. Jovin, and R. M. Clegg, “Fluorescence lifetime imaging microscopy (FLIM): Spatial resolution of microstructures on the nanosecond time scale,” Biophys. Chem. 48(2), 221–239 (1993).
[Crossref]

Ge, J.

Gerritsen, H. C.

H. C. Gerritsen, M. A. Asselbergs, A. V. Agronskaia, and W. G. Van Sark, “Fluorescence lifetime imaging in scanning microscopes: acquisition speed, photon economy and lifetime resolution,” J. Microsc. 206(3), 218–224 (2002).
[Crossref] [PubMed]

Goedhart, J.

G.-J. Kremers, E. B. van Munster, J. Goedhart, and T. W. J. Gadella, “Quantitative lifetime unmixing of multiexponentially decaying fluorophores using single-frequency fluorescence lifetime imaging microscopy,” Biophys. J. 95(1), 378–389 (2008).
[Crossref] [PubMed]

E. B. van Munster, J. Goedhart, G. J. Kremers, E. M. Manders, and T. W. Gadella, “Combination of a spinning disc confocal unit with frequency-domain fluorescence lifetime imaging microscopy,” Cytometry A 71(4), 207–214 (2007).
[Crossref] [PubMed]

Gossett, D. R.

E. D. Diebold, B. W. Buckley, D. R. Gossett, and B. Jalali, “Digitally synthesized beat frequency multiplexing for sub-millisecond fluorescence microscopy,” Nat. Photonics 7(10), 806–810 (2013).
[Crossref]

Grant, D. M.

Gratton, E.

E. Gratton, M. Limkeman, J. R. Lakowicz, B. P. Maliwal, H. Cherek, and G. Laczko, “Resolution of mixtures of fluorophores using variable-frequency phase and modulation data,” Biophys. J. 46(4), 479–486 (1984).
[Crossref] [PubMed]

Hamre, B.

A. S. Kristoffersen, S. R. Erga, B. Hamre, and Ø. Frette, “Testing fluorescence lifetime standards using two-photon excitation and time-domain instrumentation: rhodamine B, coumarin 6 and lucifer yellow,” J. Fluoresc. 24(4), 1015–1024 (2014).
[Crossref] [PubMed]

Han, W.-T.

He, H.

H. He, B. K. Nunnally, L. C. Li, and L. B. McGown, “On-the-fly fluorescence lifetime detection of dye-labeled DNA primers for multiplex analysis,” Anal. Chem. 70(16), 3413–3418 (1998).
[Crossref] [PubMed]

Heavner, T. P.

E. A. Donley, T. P. Heavner, F. Levi, M. O. Tataw, and S. R. Jefferts, “Double-pass acousto-optic modulator system,” Rev. Sci. Instrum. 76(6), 063112 (2005).
[Crossref]

Herten, D.

M. Neumann, D. Herten, A. Dietrich, J. Wolfrum, and M. Sauer, “Capillary array scanner for time-resolved detection and identification of fluorescently labelled DNA fragments,” J. Chromatography A. 871, 299–310 (2000).

Hickey, G. A.

B. J. Bacskai, J. Skoch, G. A. Hickey, R. Allen, and B. T. Hyman, “Fluorescence resonance energy transfer determinations using multiphoton fluorescence lifetime imaging microscopy to characterize amyloid-beta plaques,” J. Biomed. Opt. 8(3), 368–375 (2003).
[Crossref] [PubMed]

Hink, M. A.

W. Becker, A. Bergmann, M. A. Hink, K. König, K. Benndorf, and C. Biskup, “Fluorescence lifetime imaging by time-correlated single-photon counting,” Microsc. Res. Tech. 63(1), 58–66 (2004).
[Crossref] [PubMed]

Hofkens, J.

N. Boens, W. Qin, N. Basaric, J. Hofkens, and M. Ameloot, “Fluorescence Lifetime Standards for Time and Frequency Domain Fluorescence spectroscopy,” Analytical Chem. 79, 2137–2149 (2007).

Hyman, B. T.

B. J. Bacskai, J. Skoch, G. A. Hickey, R. Allen, and B. T. Hyman, “Fluorescence resonance energy transfer determinations using multiphoton fluorescence lifetime imaging microscopy to characterize amyloid-beta plaques,” J. Biomed. Opt. 8(3), 368–375 (2003).
[Crossref] [PubMed]

Ide, G.

G. Ide, “Fluorescence lifetime resolution with phase fluorometry,” Rev. Sci. Instrum. 54(7), 841 (1983).
[Crossref]

Ingersoll, C. M.

A. N. Watkins, C. M. Ingersoll, G. A. Baker, and F. V. Bright, “A parallel multiharmonic frequency-domain fluorometer for measuring excited-state decay kinetics following one-, two-, or three-photon excitation,” Anal. Chem. 70(16), 3384–3396 (1998).
[Crossref] [PubMed]

Jalali, B.

E. D. Diebold, B. W. Buckley, D. R. Gossett, and B. Jalali, “Digitally synthesized beat frequency multiplexing for sub-millisecond fluorescence microscopy,” Nat. Photonics 7(10), 806–810 (2013).
[Crossref]

Jefferts, S. R.

E. A. Donley, T. P. Heavner, F. Levi, M. O. Tataw, and S. R. Jefferts, “Double-pass acousto-optic modulator system,” Rev. Sci. Instrum. 76(6), 063112 (2005).
[Crossref]

Jovin, T. M.

T. W. J. Gadella, T. M. Jovin, and R. M. Clegg, “Fluorescence lifetime imaging microscopy (FLIM): Spatial resolution of microstructures on the nanosecond time scale,” Biophys. Chem. 48(2), 221–239 (1993).
[Crossref]

Kaminski, C. F.

A. D. Elder, J. H. Frank, J. Swartling, X. Dai, and C. F. Kaminski, “Calibration of a wide-field frequency-domain fluorescence lifetime microscopy system using light emitting diodes as light sources,” J. Microsc. 224(2), 166–180 (2006).
[Crossref] [PubMed]

Kao, F.-J.

Kelbauskas, L.

W. Becker, A. Bergmann, G. Biscotti, K. Koenig, I. Riemann, L. Kelbauskas, and C. Biskup, “High-speed FLIM data acquisition by time-correlated single-photon counting,” Multiphot. Microsc. Biomed. Sci. IV 5323, 27–35 (2004).
[Crossref]

Kim, B.

K. Shim and B. Kim, “Simple frequency-domain fluorescence-lifetime measurement system using violet laser diode,” J. Korean Phys. Soc. 49, 647–651 (2006).

Kim, D.

Kim, D. Y.

Koenig, K.

W. Becker, A. Bergmann, G. Biscotti, K. Koenig, I. Riemann, L. Kelbauskas, and C. Biskup, “High-speed FLIM data acquisition by time-correlated single-photon counting,” Multiphot. Microsc. Biomed. Sci. IV 5323, 27–35 (2004).
[Crossref]

König, K.

W. Becker, A. Bergmann, M. A. Hink, K. König, K. Benndorf, and C. Biskup, “Fluorescence lifetime imaging by time-correlated single-photon counting,” Microsc. Res. Tech. 63(1), 58–66 (2004).
[Crossref] [PubMed]

Kremers, G. J.

E. B. van Munster, J. Goedhart, G. J. Kremers, E. M. Manders, and T. W. Gadella, “Combination of a spinning disc confocal unit with frequency-domain fluorescence lifetime imaging microscopy,” Cytometry A 71(4), 207–214 (2007).
[Crossref] [PubMed]

Kremers, G.-J.

G.-J. Kremers, E. B. van Munster, J. Goedhart, and T. W. J. Gadella, “Quantitative lifetime unmixing of multiexponentially decaying fluorophores using single-frequency fluorescence lifetime imaging microscopy,” Biophys. J. 95(1), 378–389 (2008).
[Crossref] [PubMed]

Kristoffersen, A. S.

A. S. Kristoffersen, S. R. Erga, B. Hamre, and Ø. Frette, “Testing fluorescence lifetime standards using two-photon excitation and time-domain instrumentation: rhodamine B, coumarin 6 and lucifer yellow,” J. Fluoresc. 24(4), 1015–1024 (2014).
[Crossref] [PubMed]

Kuang, C.

Laczko, G.

E. Gratton, M. Limkeman, J. R. Lakowicz, B. P. Maliwal, H. Cherek, and G. Laczko, “Resolution of mixtures of fluorophores using variable-frequency phase and modulation data,” Biophys. J. 46(4), 479–486 (1984).
[Crossref] [PubMed]

Lakowicz, J. R.

E. Gratton, M. Limkeman, J. R. Lakowicz, B. P. Maliwal, H. Cherek, and G. Laczko, “Resolution of mixtures of fluorophores using variable-frequency phase and modulation data,” Biophys. J. 46(4), 479–486 (1984).
[Crossref] [PubMed]

Lassiter, S. J.

S. J. Lassiter, W. Stryjewski, B. L. Legendre, R. Erdmann, M. Wahl, J. Wurm, R. Peterson, L. Middendorf, and S. A. Soper, “Time-resolved fluorescence imaging of slab gels for lifetime base-calling in DNA sequencing applications,” Anal. Chem. 72(21), 5373–5382 (2000).
[Crossref] [PubMed]

Lee, S.-J.

H. Murakoshi, S.-J. Lee, and R. Yasuda, “Highly sensitive and quantitative FRET-FLIM imaging in single dendritic spines using improved non-radiative YFP,” Brain Cell Biol. 36(1-4), 31–42 (2008).
[Crossref] [PubMed]

Lee, S.-S.

Legendre, B. L.

S. J. Lassiter, W. Stryjewski, B. L. Legendre, R. Erdmann, M. Wahl, J. Wurm, R. Peterson, L. Middendorf, and S. A. Soper, “Time-resolved fluorescence imaging of slab gels for lifetime base-calling in DNA sequencing applications,” Anal. Chem. 72(21), 5373–5382 (2000).
[Crossref] [PubMed]

Lesoine, M. D.

M. D. Lesoine, S. Bose, J. W. Petrich, and E. A. Smith, “Supercontinuum stimulated emission depletion fluorescence lifetime imaging,” J. Phys. Chem. B 116(27), 7821–7826 (2012).
[Crossref] [PubMed]

Levi, F.

E. A. Donley, T. P. Heavner, F. Levi, M. O. Tataw, and S. R. Jefferts, “Double-pass acousto-optic modulator system,” Rev. Sci. Instrum. 76(6), 063112 (2005).
[Crossref]

Li, L. C.

H. He, B. K. Nunnally, L. C. Li, and L. B. McGown, “On-the-fly fluorescence lifetime detection of dye-labeled DNA primers for multiplex analysis,” Anal. Chem. 70(16), 3413–3418 (1998).
[Crossref] [PubMed]

Limkeman, M.

E. Gratton, M. Limkeman, J. R. Lakowicz, B. P. Maliwal, H. Cherek, and G. Laczko, “Resolution of mixtures of fluorophores using variable-frequency phase and modulation data,” Biophys. J. 46(4), 479–486 (1984).
[Crossref] [PubMed]

Liu, J.

D. R. Yankelevich, D. Ma, J. Liu, Y. Sun, Y. Sun, J. Bec, D. S. Elson, and L. Marcu, “Design and evaluation of a device for fast multispectral time-resolved fluorescence spectroscopy and imaging,” Rev. Sci. Instrum. 85(3), 034303 (2014).
[Crossref] [PubMed]

Lotti, T.

Ma, D.

D. R. Yankelevich, D. Ma, J. Liu, Y. Sun, Y. Sun, J. Bec, D. S. Elson, and L. Marcu, “Design and evaluation of a device for fast multispectral time-resolved fluorescence spectroscopy and imaging,” Rev. Sci. Instrum. 85(3), 034303 (2014).
[Crossref] [PubMed]

Magde, D.

D. Magde, G. E. Rojas, and P. G. Seybold, “Solvent dependence of the fluorescence lifetimes of Xanthene dyes,” Photochem. Photobiol. 70, 737–744 (1999).

Maliwal, B. P.

E. Gratton, M. Limkeman, J. R. Lakowicz, B. P. Maliwal, H. Cherek, and G. Laczko, “Resolution of mixtures of fluorophores using variable-frequency phase and modulation data,” Biophys. J. 46(4), 479–486 (1984).
[Crossref] [PubMed]

Manders, E. M.

E. B. van Munster, J. Goedhart, G. J. Kremers, E. M. Manders, and T. W. Gadella, “Combination of a spinning disc confocal unit with frequency-domain fluorescence lifetime imaging microscopy,” Cytometry A 71(4), 207–214 (2007).
[Crossref] [PubMed]

Marcu, L.

D. R. Yankelevich, D. Ma, J. Liu, Y. Sun, Y. Sun, J. Bec, D. S. Elson, and L. Marcu, “Design and evaluation of a device for fast multispectral time-resolved fluorescence spectroscopy and imaging,” Rev. Sci. Instrum. 85(3), 034303 (2014).
[Crossref] [PubMed]

Y. Sun, J. Phipps, D. S. Elson, H. Stoy, S. Tinling, J. Meier, B. Poirier, F. S. Chuang, D. G. Farwell, and L. Marcu, “Fluorescence lifetime imaging microscopy: in vivo application to diagnosis of oral carcinoma,” Opt. Lett. 34(13), 2081–2083 (2009).
[Crossref] [PubMed]

Massi, D.

Mcgown, L. B.

M. B. Smalley, J. M. Shaver, and L. B. Mcgown, “On-the-fly fluorescence lifetime detection in HPLC using a multiharmonic Fourier transform phase-modulation spectrofluorometer,” Analytical Chem. 653466–3472 (1999).

H. He, B. K. Nunnally, L. C. Li, and L. B. McGown, “On-the-fly fluorescence lifetime detection of dye-labeled DNA primers for multiplex analysis,” Anal. Chem. 70(16), 3413–3418 (1998).
[Crossref] [PubMed]

Meier, J.

Middendorf, L.

S. J. Lassiter, W. Stryjewski, B. L. Legendre, R. Erdmann, M. Wahl, J. Wurm, R. Peterson, L. Middendorf, and S. A. Soper, “Time-resolved fluorescence imaging of slab gels for lifetime base-calling in DNA sequencing applications,” Anal. Chem. 72(21), 5373–5382 (2000).
[Crossref] [PubMed]

Moon, S.

Munro, I.

Murakoshi, H.

H. Murakoshi, S.-J. Lee, and R. Yasuda, “Highly sensitive and quantitative FRET-FLIM imaging in single dendritic spines using improved non-radiative YFP,” Brain Cell Biol. 36(1-4), 31–42 (2008).
[Crossref] [PubMed]

Neil, M. A. A.

Neumann, M.

M. Neumann, D. Herten, A. Dietrich, J. Wolfrum, and M. Sauer, “Capillary array scanner for time-resolved detection and identification of fluorescently labelled DNA fragments,” J. Chromatography A. 871, 299–310 (2000).

Nunnally, B. K.

H. He, B. K. Nunnally, L. C. Li, and L. B. McGown, “On-the-fly fluorescence lifetime detection of dye-labeled DNA primers for multiplex analysis,” Anal. Chem. 70(16), 3413–3418 (1998).
[Crossref] [PubMed]

Nye, E.

Pavone, F. S.

Penzkofer, A.

W. Baumler, A. X. Schmalzl, A. Penzkofer, N. Fakultat, I. I. Physik, and U. Regensburg, “Fluorescence decay studies applying a cw femtosecond dye laser pumped ungated inverse time-correlated single photon counting system,” Meas. Sci. Technol. 3, 384–393 (1992).

Peterson, R.

S. J. Lassiter, W. Stryjewski, B. L. Legendre, R. Erdmann, M. Wahl, J. Wurm, R. Peterson, L. Middendorf, and S. A. Soper, “Time-resolved fluorescence imaging of slab gels for lifetime base-calling in DNA sequencing applications,” Anal. Chem. 72(21), 5373–5382 (2000).
[Crossref] [PubMed]

Petrich, J. W.

M. D. Lesoine, S. Bose, J. W. Petrich, and E. A. Smith, “Supercontinuum stimulated emission depletion fluorescence lifetime imaging,” J. Phys. Chem. B 116(27), 7821–7826 (2012).
[Crossref] [PubMed]

Phipps, J.

Physik, I. I.

W. Baumler, A. X. Schmalzl, A. Penzkofer, N. Fakultat, I. I. Physik, and U. Regensburg, “Fluorescence decay studies applying a cw femtosecond dye laser pumped ungated inverse time-correlated single photon counting system,” Meas. Sci. Technol. 3, 384–393 (1992).

Pifferi, A.

R. Cubeddu, G. Canti, A. Pifferi, P. Taroni, and G. Valentini, “Fluorescence lifetime imaging of experimental tumors in hematoporphyrin derivative-sensitized mice,” Photochem. Photobiol. 66(2), 229–236 (1997).
[Crossref] [PubMed]

Poirier, B.

Qin, W.

N. Boens, W. Qin, N. Basaric, J. Hofkens, and M. Ameloot, “Fluorescence Lifetime Standards for Time and Frequency Domain Fluorescence spectroscopy,” Analytical Chem. 79, 2137–2149 (2007).

Regensburg, U.

W. Baumler, A. X. Schmalzl, A. Penzkofer, N. Fakultat, I. I. Physik, and U. Regensburg, “Fluorescence decay studies applying a cw femtosecond dye laser pumped ungated inverse time-correlated single photon counting system,” Meas. Sci. Technol. 3, 384–393 (1992).

Requejo-Isidro, J.

Riemann, I.

W. Becker, A. Bergmann, G. Biscotti, K. Koenig, I. Riemann, L. Kelbauskas, and C. Biskup, “High-speed FLIM data acquisition by time-correlated single-photon counting,” Multiphot. Microsc. Biomed. Sci. IV 5323, 27–35 (2004).
[Crossref]

Rojas, G. E.

D. Magde, G. E. Rojas, and P. G. Seybold, “Solvent dependence of the fluorescence lifetimes of Xanthene dyes,” Photochem. Photobiol. 70, 737–744 (1999).

Sauer, M.

M. Neumann, D. Herten, A. Dietrich, J. Wolfrum, and M. Sauer, “Capillary array scanner for time-resolved detection and identification of fluorescently labelled DNA fragments,” J. Chromatography A. 871, 299–310 (2000).

Schimpf, D.

Schmalzl, A. X.

W. Baumler, A. X. Schmalzl, A. Penzkofer, N. Fakultat, I. I. Physik, and U. Regensburg, “Fluorescence decay studies applying a cw femtosecond dye laser pumped ungated inverse time-correlated single photon counting system,” Meas. Sci. Technol. 3, 384–393 (1992).

Sestini, S.

Seybold, P. G.

D. Magde, G. E. Rojas, and P. G. Seybold, “Solvent dependence of the fluorescence lifetimes of Xanthene dyes,” Photochem. Photobiol. 70, 737–744 (1999).

Shaver, J. M.

M. B. Smalley, J. M. Shaver, and L. B. Mcgown, “On-the-fly fluorescence lifetime detection in HPLC using a multiharmonic Fourier transform phase-modulation spectrofluorometer,” Analytical Chem. 653466–3472 (1999).

Shim, K.

K. Shim and B. Kim, “Simple frequency-domain fluorescence-lifetime measurement system using violet laser diode,” J. Korean Phys. Soc. 49, 647–651 (2006).

Shipston, M. J.

R. R. Duncan, A. Bergmann, M. A. Cousin, D. K. Apps, and M. J. Shipston, “Multi-dimensional time-correlated single photon counting (TCSPC) fluorescence lifetime imaging microscopy (FLIM) to detect FRET in cells,” J. Microsc. 215(1), 1–12 (2004).
[Crossref] [PubMed]

Skoch, J.

B. J. Bacskai, J. Skoch, G. A. Hickey, R. Allen, and B. T. Hyman, “Fluorescence resonance energy transfer determinations using multiphoton fluorescence lifetime imaging microscopy to characterize amyloid-beta plaques,” J. Biomed. Opt. 8(3), 368–375 (2003).
[Crossref] [PubMed]

Smalley, M. B.

M. B. Smalley, J. M. Shaver, and L. B. Mcgown, “On-the-fly fluorescence lifetime detection in HPLC using a multiharmonic Fourier transform phase-modulation spectrofluorometer,” Analytical Chem. 653466–3472 (1999).

Smith, E. A.

M. D. Lesoine, S. Bose, J. W. Petrich, and E. A. Smith, “Supercontinuum stimulated emission depletion fluorescence lifetime imaging,” J. Phys. Chem. B 116(27), 7821–7826 (2012).
[Crossref] [PubMed]

Soper, S. A.

S. J. Lassiter, W. Stryjewski, B. L. Legendre, R. Erdmann, M. Wahl, J. Wurm, R. Peterson, L. Middendorf, and S. A. Soper, “Time-resolved fluorescence imaging of slab gels for lifetime base-calling in DNA sequencing applications,” Anal. Chem. 72(21), 5373–5382 (2000).
[Crossref] [PubMed]

Squire, A.

A. Squire, P. J. Verveer, and P. I. H. Bastiaens, “Multiple frequency fluorescence lifetime imaging microscopy,” J. Microscopy 197, 136–149 (2000).

Stamp, G.

Stoy, H.

Stryjewski, W.

S. J. Lassiter, W. Stryjewski, B. L. Legendre, R. Erdmann, M. Wahl, J. Wurm, R. Peterson, L. Middendorf, and S. A. Soper, “Time-resolved fluorescence imaging of slab gels for lifetime base-calling in DNA sequencing applications,” Anal. Chem. 72(21), 5373–5382 (2000).
[Crossref] [PubMed]

Sun, Y.

D. R. Yankelevich, D. Ma, J. Liu, Y. Sun, Y. Sun, J. Bec, D. S. Elson, and L. Marcu, “Design and evaluation of a device for fast multispectral time-resolved fluorescence spectroscopy and imaging,” Rev. Sci. Instrum. 85(3), 034303 (2014).
[Crossref] [PubMed]

D. R. Yankelevich, D. Ma, J. Liu, Y. Sun, Y. Sun, J. Bec, D. S. Elson, and L. Marcu, “Design and evaluation of a device for fast multispectral time-resolved fluorescence spectroscopy and imaging,” Rev. Sci. Instrum. 85(3), 034303 (2014).
[Crossref] [PubMed]

Y. Sun, J. Phipps, D. S. Elson, H. Stoy, S. Tinling, J. Meier, B. Poirier, F. S. Chuang, D. G. Farwell, and L. Marcu, “Fluorescence lifetime imaging microscopy: in vivo application to diagnosis of oral carcinoma,” Opt. Lett. 34(13), 2081–2083 (2009).
[Crossref] [PubMed]

Swartling, J.

A. D. Elder, J. H. Frank, J. Swartling, X. Dai, and C. F. Kaminski, “Calibration of a wide-field frequency-domain fluorescence lifetime microscopy system using light emitting diodes as light sources,” J. Microsc. 224(2), 166–180 (2006).
[Crossref] [PubMed]

Taroni, P.

R. Cubeddu, G. Canti, A. Pifferi, P. Taroni, and G. Valentini, “Fluorescence lifetime imaging of experimental tumors in hematoporphyrin derivative-sensitized mice,” Photochem. Photobiol. 66(2), 229–236 (1997).
[Crossref] [PubMed]

Tataw, M. O.

E. A. Donley, T. P. Heavner, F. Levi, M. O. Tataw, and S. R. Jefferts, “Double-pass acousto-optic modulator system,” Rev. Sci. Instrum. 76(6), 063112 (2005).
[Crossref]

Tinling, S.

Valentini, G.

R. Cubeddu, G. Canti, A. Pifferi, P. Taroni, and G. Valentini, “Fluorescence lifetime imaging of experimental tumors in hematoporphyrin derivative-sensitized mice,” Photochem. Photobiol. 66(2), 229–236 (1997).
[Crossref] [PubMed]

van Munster, E. B.

G.-J. Kremers, E. B. van Munster, J. Goedhart, and T. W. J. Gadella, “Quantitative lifetime unmixing of multiexponentially decaying fluorophores using single-frequency fluorescence lifetime imaging microscopy,” Biophys. J. 95(1), 378–389 (2008).
[Crossref] [PubMed]

E. B. van Munster, J. Goedhart, G. J. Kremers, E. M. Manders, and T. W. Gadella, “Combination of a spinning disc confocal unit with frequency-domain fluorescence lifetime imaging microscopy,” Cytometry A 71(4), 207–214 (2007).
[Crossref] [PubMed]

Van Sark, W. G.

H. C. Gerritsen, M. A. Asselbergs, A. V. Agronskaia, and W. G. Van Sark, “Fluorescence lifetime imaging in scanning microscopes: acquisition speed, photon economy and lifetime resolution,” J. Microsc. 206(3), 218–224 (2002).
[Crossref] [PubMed]

Verkman, A. S.

A. S. Verkman, M. Armijo, and K. Fushimi, “Construction and evaluation of a frequency-domain epifluorescence microscope for lifetime and anisotropy decay measurements in subcellular domains,” Biophys. Chem. 40(1), 117–125 (1991).
[Crossref] [PubMed]

Verveer, P. J.

A. Squire, P. J. Verveer, and P. I. H. Bastiaens, “Multiple frequency fluorescence lifetime imaging microscopy,” J. Microscopy 197, 136–149 (2000).

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S. J. Lassiter, W. Stryjewski, B. L. Legendre, R. Erdmann, M. Wahl, J. Wurm, R. Peterson, L. Middendorf, and S. A. Soper, “Time-resolved fluorescence imaging of slab gels for lifetime base-calling in DNA sequencing applications,” Anal. Chem. 72(21), 5373–5382 (2000).
[Crossref] [PubMed]

Watkins, A. N.

A. N. Watkins, C. M. Ingersoll, G. A. Baker, and F. V. Bright, “A parallel multiharmonic frequency-domain fluorometer for measuring excited-state decay kinetics following one-, two-, or three-photon excitation,” Anal. Chem. 70(16), 3384–3396 (1998).
[Crossref] [PubMed]

Wolfrum, J.

M. Neumann, D. Herten, A. Dietrich, J. Wolfrum, and M. Sauer, “Capillary array scanner for time-resolved detection and identification of fluorescently labelled DNA fragments,” J. Chromatography A. 871, 299–310 (2000).

Won, Y.

Wurm, J.

S. J. Lassiter, W. Stryjewski, B. L. Legendre, R. Erdmann, M. Wahl, J. Wurm, R. Peterson, L. Middendorf, and S. A. Soper, “Time-resolved fluorescence imaging of slab gels for lifetime base-calling in DNA sequencing applications,” Anal. Chem. 72(21), 5373–5382 (2000).
[Crossref] [PubMed]

Yang, W.

Yankelevich, D. R.

D. R. Yankelevich, D. Ma, J. Liu, Y. Sun, Y. Sun, J. Bec, D. S. Elson, and L. Marcu, “Design and evaluation of a device for fast multispectral time-resolved fluorescence spectroscopy and imaging,” Rev. Sci. Instrum. 85(3), 034303 (2014).
[Crossref] [PubMed]

Yao, S.-K.

E. H. J. Young and S.-K. Yao, “Design considerations for acousto-optic devices,” Proc. IEEE 69, 54–64 (1981).
[Crossref]

Yasuda, R.

H. Murakoshi, S.-J. Lee, and R. Yasuda, “Highly sensitive and quantitative FRET-FLIM imaging in single dendritic spines using improved non-radiative YFP,” Brain Cell Biol. 36(1-4), 31–42 (2008).
[Crossref] [PubMed]

Young, E. H. J.

E. H. J. Young and S.-K. Yao, “Design considerations for acousto-optic devices,” Proc. IEEE 69, 54–64 (1981).
[Crossref]

Anal. Chem. (3)

S. J. Lassiter, W. Stryjewski, B. L. Legendre, R. Erdmann, M. Wahl, J. Wurm, R. Peterson, L. Middendorf, and S. A. Soper, “Time-resolved fluorescence imaging of slab gels for lifetime base-calling in DNA sequencing applications,” Anal. Chem. 72(21), 5373–5382 (2000).
[Crossref] [PubMed]

H. He, B. K. Nunnally, L. C. Li, and L. B. McGown, “On-the-fly fluorescence lifetime detection of dye-labeled DNA primers for multiplex analysis,” Anal. Chem. 70(16), 3413–3418 (1998).
[Crossref] [PubMed]

A. N. Watkins, C. M. Ingersoll, G. A. Baker, and F. V. Bright, “A parallel multiharmonic frequency-domain fluorometer for measuring excited-state decay kinetics following one-, two-, or three-photon excitation,” Anal. Chem. 70(16), 3384–3396 (1998).
[Crossref] [PubMed]

Analytical Chem. (2)

M. B. Smalley, J. M. Shaver, and L. B. Mcgown, “On-the-fly fluorescence lifetime detection in HPLC using a multiharmonic Fourier transform phase-modulation spectrofluorometer,” Analytical Chem. 653466–3472 (1999).

N. Boens, W. Qin, N. Basaric, J. Hofkens, and M. Ameloot, “Fluorescence Lifetime Standards for Time and Frequency Domain Fluorescence spectroscopy,” Analytical Chem. 79, 2137–2149 (2007).

Biophys. Chem. (2)

A. S. Verkman, M. Armijo, and K. Fushimi, “Construction and evaluation of a frequency-domain epifluorescence microscope for lifetime and anisotropy decay measurements in subcellular domains,” Biophys. Chem. 40(1), 117–125 (1991).
[Crossref] [PubMed]

T. W. J. Gadella, T. M. Jovin, and R. M. Clegg, “Fluorescence lifetime imaging microscopy (FLIM): Spatial resolution of microstructures on the nanosecond time scale,” Biophys. Chem. 48(2), 221–239 (1993).
[Crossref]

Biophys. J. (2)

E. Gratton, M. Limkeman, J. R. Lakowicz, B. P. Maliwal, H. Cherek, and G. Laczko, “Resolution of mixtures of fluorophores using variable-frequency phase and modulation data,” Biophys. J. 46(4), 479–486 (1984).
[Crossref] [PubMed]

G.-J. Kremers, E. B. van Munster, J. Goedhart, and T. W. J. Gadella, “Quantitative lifetime unmixing of multiexponentially decaying fluorophores using single-frequency fluorescence lifetime imaging microscopy,” Biophys. J. 95(1), 378–389 (2008).
[Crossref] [PubMed]

Brain Cell Biol. (1)

H. Murakoshi, S.-J. Lee, and R. Yasuda, “Highly sensitive and quantitative FRET-FLIM imaging in single dendritic spines using improved non-radiative YFP,” Brain Cell Biol. 36(1-4), 31–42 (2008).
[Crossref] [PubMed]

Cytometry A (1)

E. B. van Munster, J. Goedhart, G. J. Kremers, E. M. Manders, and T. W. Gadella, “Combination of a spinning disc confocal unit with frequency-domain fluorescence lifetime imaging microscopy,” Cytometry A 71(4), 207–214 (2007).
[Crossref] [PubMed]

J. Biomed. Opt. (1)

B. J. Bacskai, J. Skoch, G. A. Hickey, R. Allen, and B. T. Hyman, “Fluorescence resonance energy transfer determinations using multiphoton fluorescence lifetime imaging microscopy to characterize amyloid-beta plaques,” J. Biomed. Opt. 8(3), 368–375 (2003).
[Crossref] [PubMed]

J. Chromatography A. (1)

M. Neumann, D. Herten, A. Dietrich, J. Wolfrum, and M. Sauer, “Capillary array scanner for time-resolved detection and identification of fluorescently labelled DNA fragments,” J. Chromatography A. 871, 299–310 (2000).

J. Fluoresc. (1)

A. S. Kristoffersen, S. R. Erga, B. Hamre, and Ø. Frette, “Testing fluorescence lifetime standards using two-photon excitation and time-domain instrumentation: rhodamine B, coumarin 6 and lucifer yellow,” J. Fluoresc. 24(4), 1015–1024 (2014).
[Crossref] [PubMed]

J. Korean Phys. Soc. (1)

K. Shim and B. Kim, “Simple frequency-domain fluorescence-lifetime measurement system using violet laser diode,” J. Korean Phys. Soc. 49, 647–651 (2006).

J. Microsc. (3)

A. D. Elder, J. H. Frank, J. Swartling, X. Dai, and C. F. Kaminski, “Calibration of a wide-field frequency-domain fluorescence lifetime microscopy system using light emitting diodes as light sources,” J. Microsc. 224(2), 166–180 (2006).
[Crossref] [PubMed]

R. R. Duncan, A. Bergmann, M. A. Cousin, D. K. Apps, and M. J. Shipston, “Multi-dimensional time-correlated single photon counting (TCSPC) fluorescence lifetime imaging microscopy (FLIM) to detect FRET in cells,” J. Microsc. 215(1), 1–12 (2004).
[Crossref] [PubMed]

H. C. Gerritsen, M. A. Asselbergs, A. V. Agronskaia, and W. G. Van Sark, “Fluorescence lifetime imaging in scanning microscopes: acquisition speed, photon economy and lifetime resolution,” J. Microsc. 206(3), 218–224 (2002).
[Crossref] [PubMed]

J. Microscopy (1)

A. Squire, P. J. Verveer, and P. I. H. Bastiaens, “Multiple frequency fluorescence lifetime imaging microscopy,” J. Microscopy 197, 136–149 (2000).

J. Phys. Chem. B (1)

M. D. Lesoine, S. Bose, J. W. Petrich, and E. A. Smith, “Supercontinuum stimulated emission depletion fluorescence lifetime imaging,” J. Phys. Chem. B 116(27), 7821–7826 (2012).
[Crossref] [PubMed]

Meas. Sci. Technol. (1)

W. Baumler, A. X. Schmalzl, A. Penzkofer, N. Fakultat, I. I. Physik, and U. Regensburg, “Fluorescence decay studies applying a cw femtosecond dye laser pumped ungated inverse time-correlated single photon counting system,” Meas. Sci. Technol. 3, 384–393 (1992).

Microsc. Res. Tech. (1)

W. Becker, A. Bergmann, M. A. Hink, K. König, K. Benndorf, and C. Biskup, “Fluorescence lifetime imaging by time-correlated single-photon counting,” Microsc. Res. Tech. 63(1), 58–66 (2004).
[Crossref] [PubMed]

Multiphot. Microsc. Biomed. Sci. IV (1)

W. Becker, A. Bergmann, G. Biscotti, K. Koenig, I. Riemann, L. Kelbauskas, and C. Biskup, “High-speed FLIM data acquisition by time-correlated single-photon counting,” Multiphot. Microsc. Biomed. Sci. IV 5323, 27–35 (2004).
[Crossref]

Nat. Photonics (1)

E. D. Diebold, B. W. Buckley, D. R. Gossett, and B. Jalali, “Digitally synthesized beat frequency multiplexing for sub-millisecond fluorescence microscopy,” Nat. Photonics 7(10), 806–810 (2013).
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Opt. Express (3)

Opt. Lett. (2)

Photochem. Photobiol. (2)

D. Magde, G. E. Rojas, and P. G. Seybold, “Solvent dependence of the fluorescence lifetimes of Xanthene dyes,” Photochem. Photobiol. 70, 737–744 (1999).

R. Cubeddu, G. Canti, A. Pifferi, P. Taroni, and G. Valentini, “Fluorescence lifetime imaging of experimental tumors in hematoporphyrin derivative-sensitized mice,” Photochem. Photobiol. 66(2), 229–236 (1997).
[Crossref] [PubMed]

Proc. IEEE (1)

E. H. J. Young and S.-K. Yao, “Design considerations for acousto-optic devices,” Proc. IEEE 69, 54–64 (1981).
[Crossref]

Rev. Sci. Instrum. (3)

G. Ide, “Fluorescence lifetime resolution with phase fluorometry,” Rev. Sci. Instrum. 54(7), 841 (1983).
[Crossref]

E. A. Donley, T. P. Heavner, F. Levi, M. O. Tataw, and S. R. Jefferts, “Double-pass acousto-optic modulator system,” Rev. Sci. Instrum. 76(6), 063112 (2005).
[Crossref]

D. R. Yankelevich, D. Ma, J. Liu, Y. Sun, Y. Sun, J. Bec, D. S. Elson, and L. Marcu, “Design and evaluation of a device for fast multispectral time-resolved fluorescence spectroscopy and imaging,” Rev. Sci. Instrum. 85(3), 034303 (2014).
[Crossref] [PubMed]

Other (3)

FLIM X16 TCSPC Detector - LaVision BioTec GmbH,” http://www.lavisionbiotec.com/flim-x16-tcspc-detector.html .

D. Elson, N. Galletly, and C. Talbot, “Multidimensional fluorescence imaging applied to biological tissue,” in Reviews in Fluorescence (Springer, 2006), pp. 477–524.

J. R. Lakowicz, “Frequency-domain Lifetime Measurements,” in Principles of Fluorescence Spectroscopy (Springer, 2006), pp. 157–199.

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Figures (4)

Fig. 1
Fig. 1

Schematic of (a) comb generation and (b) imaging setup. The laser beam is frequency-shifted at multiple beat frequencies using the AOD (Crystal Technology, AOMO 3200-125) in a cat’s-eye configuration, driven by a frequency comb from an arbitrary waveform generator (AWG). The pinhole (PH) selects the doubly-diffracted beam from the AOD. The single beam from the comb generation setup is focused by a 10x, 0.3-NA objective lens onto the sample. Fluorescence is collected in the epi-direction and filtered by a 532 nm long pass dichroic mirror (DM) and a 580/20 band pass filter (BPF), before focusing through a 40-μm pinhole in a confocal configuration. Finally, the fluorescence signal is collected by a photomultipler tube (PMT). The electronic output from the PMT is low-pass filtered (LPF) and amplified (LNA) before being digitized along with the reference photodiode signal. (PBS: polarizing beam splitter)

Fig. 2
Fig. 2

Generation of multiple unique beat frequencies using the AOD and cat’s-eye configuration. (a) Time domain signal of RF tones { ω j } . (b) The signal in (a) drives the AOD in the cat’s-eye configuration to produce an optical beam containing frequency shifts of twice the input RF tone frequencies. (c) These RF tones are engineered to avoid beat frequency overlap, which causes phase ambiguity.

Fig. 3
Fig. 3

(a) Selected fluorescence lifetime measurements. (b) Lifetime measurements of Rhodamine B / Rhodamine 6G mixtures at various ratios in water. The error bars denote the 95% confidence interval of the signal windows at each beat frequency. A non-linear fitting of Eq. (2) was applied to the data to extract the fluorescence lifetime values.

Fig. 4
Fig. 4

Measured RB:R6G two-component mixtures at different component ratios (circles), as compared to the ideal case (dashed line).

Tables (2)

Tables Icon

Table 1 Calculated lifetime values of selected fluorescent dye solutions, with literature values

Tables Icon

Table 2 Calculated lifetimes and component ratios of RB:R6G two-component mixtures

Equations (4)

Equations on this page are rendered with MathJax. Learn more.

m τ = 1 1+ ω 2 τ 2
φ τ = tan 1 (ωτ)= cos 1 ( 1 1+ ω 2 τ 2 ),
I(t)= j=1 n | E j expi[ ( ω 0 2 ω j )t+ φ j ] | 2 = I 0 +2 j=1 n1 k=j+1 n I jk cos( 2 Ω jk t Φ jk ),
F(t)= I 0 +2 j=1 n1 k=j+1 n I jk 1+ Ω jk 2 τ 2 cos( 2 Ω jk t Φ jk φ τ,jk )

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