Abstract

We demonstrate time-gated confocal imaging as a means to separate coherent anti-Stokes Raman scattering (CARS) microscopy data from multi-photon excited endogenous fluorescence in tissue. CARS is a quasi-instantaneous process and its signal decay time is only limited by the system’s instrument response function (IRF). Signals due to two-photon-excited (TPE) tissue autofluorescence with excited state lifetimes on the nanosecond scale can be identified and separated from the CARS signal by employing time-gating techniques. We demonstrate this improved contrast on the example of CARS microscopy of intact roots of plant seedlings as well as on rat arterial tissue.

© 2007 Optical Society of America

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  1. C. L. Evans, E. O. Potma, M. Puoris'haag, D. Cote, C. P. Lin, and X. S. Xie, "Chemical imaging of tissue in vivo with video-rate coherent anti-Stokes Raman scattering microscopy," PNAS 102, 16807-16812 (2005).
    [CrossRef] [PubMed]
  2. T. B. Huff and J. X. Cheng, "In vivo coherent anti-Stokes Raman scattering imaging of sciatic nerve tissue," J. Microsc. 225, 175-182 (2007).
    [CrossRef] [PubMed]
  3. H. Kano and H. Hamaguchi, "Vibrationally resonant imaging of a single living cell by supercontinuum- based multiplex coherent anti-Stokes Raman Scattering Microspectroscopy," Opt. Express 13, 1322- 1327 (2005).
    [CrossRef] [PubMed]
  4. O. Burkacky, A. Zumbusch, C. Brackmann, and A. Enejder, "Dual-pump coherent anti-Stokes-Raman Scattering Microscopy," Opt. Lett. 31, 3656-3658 (2006).
    [CrossRef] [PubMed]
  5. X. L. Nan, E. O. Potma, and X. S. Xie, "Nonperturbative chemical imaging of organelle transport in living cells with coherent anti-stokes Raman Scattering Microscopy," Biophys. J. 91, 728-735 (2006).
    [CrossRef] [PubMed]
  6. L. Tong, Y. Lu, R. J. Lee, and J. X. Cheng, "Imaging receptor-mediated endocytosis with a polymeric nanoparticle-based coherent anti-stokes raman scattering probe," J. Phys. Chem. B 111, 9980-9985 (2007).
    [CrossRef] [PubMed]
  7. H. F. Wang, Y. Fu, P. Zickmund, R. Y. Shi, and J. X. Cheng, "Coherent anti-stokes Raman scattering imaging of axonal myelin in live spinal tissues," Biophys. J. 89, 581-591 (2005).
    [CrossRef] [PubMed]
  8. J. X. Cheng, Y. K. Jia, G. F. Zheng, and X. S. Xie, "Laser-scanning coherent anti-stokes RamanScattering Microscopy and applications to cell biology," Biophys. J. 83, 502-509 (2002).
    [CrossRef] [PubMed]
  9. J. X. Cheng, A. Volkmer, and X. S. Xie, "Theoretical and experimental characterization of Coherent anti- Stokes Raman Scattering Microscopy," J. Opt. Soc. Am. B 19, 1363-1375 (2002)
    [CrossRef]
  10. J. X. Cheng and X. S. Xie, "Coherent anti-Stokes Raman scattering microscopy: Instrumentation, theory, and applications," J. Phys. Chem. B 108, 827-840 (2004).
    [CrossRef]
  11. A. Volkmer, J. X. Cheng, and X. S. Xie, "Vibrational imaging with high sensitivity via epidetected coherent anti-Stokes Raman scattering microscopy," Phys. Rev. Lett. 8702, 4 (2001).
  12. N. Djaker, P. F. Lenne, D. Marguet, A. Colonna, C. Hadjur, and H. Rigneault, "Coherent anti-Stokes Raman scattering microscopy (CARS): Instrumentation and applications," Nuc. Instr. Meth. Phys. 571, 177-181 (2007).
    [CrossRef]
  13. L. G. Rodriguez, S. J. Lockett, and G. R. Holtom, "Coherent anti-stokes Raman scattering microscopy: A biological review," Cytometry A 69A, 779-791 (2006).
    [CrossRef]
  14. G. J. Puppels, F. F. M. Demul, C. Otto, J. Greve, M. Robertnicoud, D. J. Arndtjovin, and T. M. Jovin, "Studying single living cells and chromosomes by Confocal Raman Microspectroscopy," Nature 347, 301-303 (1990).
    [CrossRef] [PubMed]
  15. J. W. Chan, A. P. Esposito, C. E. Talley, C. W. Hollars, S. M. Lane, and T. Huser, "Reagentless identification of single bacterial spores in aqueous solution by confocal laser tweezers Raman spectroscopy," Anal. Chem. 76, 599-603 (2004).
    [CrossRef] [PubMed]
  16. N. J. Crane, M. D. Morris, M. A. Ignelzi, and G. G. Yu, "Raman imaging demonstrates FGF2-induced craniosynostosis in mouse calvaria," J. Biomed. Opt. 10, 8 (2005).
    [CrossRef]
  17. M. Kazanci, H. D. Wagner, N. I. Manjubala, H. S. Gupta, E. Paschalis, P. Roschger, and P. Fratzl, "Raman imaging of two orthogonal planes within cortical bone," Bone 41, 456-461 (2007).
    [CrossRef] [PubMed]
  18. G. J. Zhang, D. J. Moore, C. R. Flach, and R. Mendelsohn, "Vibrational microscopy and imaging of skin: from single cells to intact tissue," Anal. Bioanal. Chem. 387, 1591-1599 (2007).
    [CrossRef]
  19. G. J. Zhang, D. J. Moore, K. B. Sloan, C. R. Flach, and R. Mendelsohn, "Imaging the prodrug-to-drug transformation of a 5-fluorouracil derivative in skin by confocal Raman microscopy," J. Invest. Dermatol. 127, 1205-1209 (2007).
    [CrossRef] [PubMed]
  20. M. D. Duncan, J. Reintjes, and T. J. Manuccia, "Scanning Coherent Anti-Stokes Raman Microscope," Opt. Lett. 7, 350-352 (1982).
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  21. A. Zumbusch, G. R. Holtom, and X. S. Xie, "Three-dimensional vibrational imaging by coherent anti- Stokes Raman scattering," Phys. Rev. Lett. 82, 4142-4145 (1999).
    [CrossRef]
  22. K. Konig and I. Riemann, "High-resolution multiphoton tomography of human skin with subcellular spatial resolution and picosecond time resolution," J. Biomed. Opt. 8, 432-439 (2003).
    [CrossRef] [PubMed]
  23. K. Konig, K. Schenke-Layland, I. Riemann, and U. A. Stock, "Multiphoton autofluorescence imaging of intratissue elastic fibers," Biomaterials 26, 495-500 (2005).
    [CrossRef]
  24. M. C. Skala, J. M. Squirrell, K. M. Vrotsos, V. C. Eickhoff, A. Gendron-Fitzpatrick, K. W. Eliceiri, and N. Ramanujam, "Multiphoton microscopy of endogenous fluorescence differentiates normal, precancerous, and cancerous squamous epithelial tissues," Cancer Res. 65, 1180-1186 (2005).
    [CrossRef] [PubMed]
  25. J. A. Palero, H. S. de Bruijn, A. van der Ploeg-van den Heuvel, H. Sterenborg, and H. C. Gerritsen, "In vivo nonlinear spectral imaging in mouse skin," Opt. Express 14, 4395-4402 (2006).
    [CrossRef] [PubMed]
  26. J. M. Piepmeier, C. Y. Liu, and E. Neuwelt, "Multiphoton excitation of autofluorescence for microscopy of glioma tissue - Comments," Neurosurgery 58, 767-767 (2006).
  27. J. A. Palero, H. S. de Bruijn, A. V. van den Heuvel, H. Sterenborg, and H. C. Gerritsen, "Spectrally resolved multiphoton imaging of in vivo and excised mouse skin tissues," Biophys. J. 93, 992-1007 (2007).
    [CrossRef] [PubMed]
  28. E. S. Lee, J. Y. Lee, and Y. S. Yoo, "Nonlinear optical interference of two successive coherent anti-Stokes Raman scattering signals for biological imaging applications," J. Biomed. Opt. 12, 5 (2007).
    [CrossRef]
  29. E. O. Potma, C. L. Evans, and X. S. Xie, "Heterodyne coherent anti-Stokes Raman scattering (CARS) imaging," Opt. Lett. 31, 241-243 (2006).
    [CrossRef] [PubMed]
  30. I. Toytman, K. Cohn, T. Smith, D. Simanovskii, and D. Palanker, "Wide-field coherent anti-Stokes Raman scattering microscopy with non-phase-matching illumination," Opt. Lett. 32, 1941-1943 (2007).
    [CrossRef] [PubMed]
  31. W. Becker, A. Bergmann, M. A. Hink, K. Konig, K. Benndorf, and C. Biskup, "Fluorescence lifetime imaging by time-correlated single-photon counting," Microsc. Res. Technique 63, 58-66 (2004).
    [CrossRef]
  32. 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, 1-12 (2004).
    [CrossRef] [PubMed]
  33. M. Peter, and S. M. Ameer-Beg, "Imaging molecular interactions by multiphoton FLIM," Biology of the Cell 96, 231-236 (2004).
    [CrossRef] [PubMed]
  34. K. Suhling, P. M. W. French, and D. Phillips, "Time-resolved fluorescence microscopy," Photochem. Photobiol. Sci. 4, 13-22 (2005).
    [CrossRef]

2007 (9)

T. B. Huff and J. X. Cheng, "In vivo coherent anti-Stokes Raman scattering imaging of sciatic nerve tissue," J. Microsc. 225, 175-182 (2007).
[CrossRef] [PubMed]

L. Tong, Y. Lu, R. J. Lee, and J. X. Cheng, "Imaging receptor-mediated endocytosis with a polymeric nanoparticle-based coherent anti-stokes raman scattering probe," J. Phys. Chem. B 111, 9980-9985 (2007).
[CrossRef] [PubMed]

N. Djaker, P. F. Lenne, D. Marguet, A. Colonna, C. Hadjur, and H. Rigneault, "Coherent anti-Stokes Raman scattering microscopy (CARS): Instrumentation and applications," Nuc. Instr. Meth. Phys. 571, 177-181 (2007).
[CrossRef]

M. Kazanci, H. D. Wagner, N. I. Manjubala, H. S. Gupta, E. Paschalis, P. Roschger, and P. Fratzl, "Raman imaging of two orthogonal planes within cortical bone," Bone 41, 456-461 (2007).
[CrossRef] [PubMed]

G. J. Zhang, D. J. Moore, C. R. Flach, and R. Mendelsohn, "Vibrational microscopy and imaging of skin: from single cells to intact tissue," Anal. Bioanal. Chem. 387, 1591-1599 (2007).
[CrossRef]

G. J. Zhang, D. J. Moore, K. B. Sloan, C. R. Flach, and R. Mendelsohn, "Imaging the prodrug-to-drug transformation of a 5-fluorouracil derivative in skin by confocal Raman microscopy," J. Invest. Dermatol. 127, 1205-1209 (2007).
[CrossRef] [PubMed]

J. A. Palero, H. S. de Bruijn, A. V. van den Heuvel, H. Sterenborg, and H. C. Gerritsen, "Spectrally resolved multiphoton imaging of in vivo and excised mouse skin tissues," Biophys. J. 93, 992-1007 (2007).
[CrossRef] [PubMed]

E. S. Lee, J. Y. Lee, and Y. S. Yoo, "Nonlinear optical interference of two successive coherent anti-Stokes Raman scattering signals for biological imaging applications," J. Biomed. Opt. 12, 5 (2007).
[CrossRef]

I. Toytman, K. Cohn, T. Smith, D. Simanovskii, and D. Palanker, "Wide-field coherent anti-Stokes Raman scattering microscopy with non-phase-matching illumination," Opt. Lett. 32, 1941-1943 (2007).
[CrossRef] [PubMed]

2006 (6)

J. M. Piepmeier, C. Y. Liu, and E. Neuwelt, "Multiphoton excitation of autofluorescence for microscopy of glioma tissue - Comments," Neurosurgery 58, 767-767 (2006).

E. O. Potma, C. L. Evans, and X. S. Xie, "Heterodyne coherent anti-Stokes Raman scattering (CARS) imaging," Opt. Lett. 31, 241-243 (2006).
[CrossRef] [PubMed]

J. A. Palero, H. S. de Bruijn, A. van der Ploeg-van den Heuvel, H. Sterenborg, and H. C. Gerritsen, "In vivo nonlinear spectral imaging in mouse skin," Opt. Express 14, 4395-4402 (2006).
[CrossRef] [PubMed]

O. Burkacky, A. Zumbusch, C. Brackmann, and A. Enejder, "Dual-pump coherent anti-Stokes-Raman Scattering Microscopy," Opt. Lett. 31, 3656-3658 (2006).
[CrossRef] [PubMed]

L. G. Rodriguez, S. J. Lockett, and G. R. Holtom, "Coherent anti-stokes Raman scattering microscopy: A biological review," Cytometry A 69A, 779-791 (2006).
[CrossRef]

X. L. Nan, E. O. Potma, and X. S. Xie, "Nonperturbative chemical imaging of organelle transport in living cells with coherent anti-stokes Raman Scattering Microscopy," Biophys. J. 91, 728-735 (2006).
[CrossRef] [PubMed]

2005 (7)

H. F. Wang, Y. Fu, P. Zickmund, R. Y. Shi, and J. X. Cheng, "Coherent anti-stokes Raman scattering imaging of axonal myelin in live spinal tissues," Biophys. J. 89, 581-591 (2005).
[CrossRef] [PubMed]

C. L. Evans, E. O. Potma, M. Puoris'haag, D. Cote, C. P. Lin, and X. S. Xie, "Chemical imaging of tissue in vivo with video-rate coherent anti-Stokes Raman scattering microscopy," PNAS 102, 16807-16812 (2005).
[CrossRef] [PubMed]

N. J. Crane, M. D. Morris, M. A. Ignelzi, and G. G. Yu, "Raman imaging demonstrates FGF2-induced craniosynostosis in mouse calvaria," J. Biomed. Opt. 10, 8 (2005).
[CrossRef]

H. Kano and H. Hamaguchi, "Vibrationally resonant imaging of a single living cell by supercontinuum- based multiplex coherent anti-Stokes Raman Scattering Microspectroscopy," Opt. Express 13, 1322- 1327 (2005).
[CrossRef] [PubMed]

K. Konig, K. Schenke-Layland, I. Riemann, and U. A. Stock, "Multiphoton autofluorescence imaging of intratissue elastic fibers," Biomaterials 26, 495-500 (2005).
[CrossRef]

M. C. Skala, J. M. Squirrell, K. M. Vrotsos, V. C. Eickhoff, A. Gendron-Fitzpatrick, K. W. Eliceiri, and N. Ramanujam, "Multiphoton microscopy of endogenous fluorescence differentiates normal, precancerous, and cancerous squamous epithelial tissues," Cancer Res. 65, 1180-1186 (2005).
[CrossRef] [PubMed]

K. Suhling, P. M. W. French, and D. Phillips, "Time-resolved fluorescence microscopy," Photochem. Photobiol. Sci. 4, 13-22 (2005).
[CrossRef]

2004 (5)

W. Becker, A. Bergmann, M. A. Hink, K. Konig, K. Benndorf, and C. Biskup, "Fluorescence lifetime imaging by time-correlated single-photon counting," Microsc. Res. Technique 63, 58-66 (2004).
[CrossRef]

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, 1-12 (2004).
[CrossRef] [PubMed]

M. Peter, and S. M. Ameer-Beg, "Imaging molecular interactions by multiphoton FLIM," Biology of the Cell 96, 231-236 (2004).
[CrossRef] [PubMed]

J. W. Chan, A. P. Esposito, C. E. Talley, C. W. Hollars, S. M. Lane, and T. Huser, "Reagentless identification of single bacterial spores in aqueous solution by confocal laser tweezers Raman spectroscopy," Anal. Chem. 76, 599-603 (2004).
[CrossRef] [PubMed]

J. X. Cheng and X. S. Xie, "Coherent anti-Stokes Raman scattering microscopy: Instrumentation, theory, and applications," J. Phys. Chem. B 108, 827-840 (2004).
[CrossRef]

2003 (1)

K. Konig and I. Riemann, "High-resolution multiphoton tomography of human skin with subcellular spatial resolution and picosecond time resolution," J. Biomed. Opt. 8, 432-439 (2003).
[CrossRef] [PubMed]

2002 (2)

J. X. Cheng, Y. K. Jia, G. F. Zheng, and X. S. Xie, "Laser-scanning coherent anti-stokes RamanScattering Microscopy and applications to cell biology," Biophys. J. 83, 502-509 (2002).
[CrossRef] [PubMed]

J. X. Cheng, A. Volkmer, and X. S. Xie, "Theoretical and experimental characterization of Coherent anti- Stokes Raman Scattering Microscopy," J. Opt. Soc. Am. B 19, 1363-1375 (2002)
[CrossRef]

2001 (1)

A. Volkmer, J. X. Cheng, and X. S. Xie, "Vibrational imaging with high sensitivity via epidetected coherent anti-Stokes Raman scattering microscopy," Phys. Rev. Lett. 8702, 4 (2001).

1999 (1)

A. Zumbusch, G. R. Holtom, and X. S. Xie, "Three-dimensional vibrational imaging by coherent anti- Stokes Raman scattering," Phys. Rev. Lett. 82, 4142-4145 (1999).
[CrossRef]

1990 (1)

G. J. Puppels, F. F. M. Demul, C. Otto, J. Greve, M. Robertnicoud, D. J. Arndtjovin, and T. M. Jovin, "Studying single living cells and chromosomes by Confocal Raman Microspectroscopy," Nature 347, 301-303 (1990).
[CrossRef] [PubMed]

1982 (1)

Ameer-Beg, S. M.

M. Peter, and S. M. Ameer-Beg, "Imaging molecular interactions by multiphoton FLIM," Biology of the Cell 96, 231-236 (2004).
[CrossRef] [PubMed]

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, 1-12 (2004).
[CrossRef] [PubMed]

Arndtjovin, D. J.

G. J. Puppels, F. F. M. Demul, C. Otto, J. Greve, M. Robertnicoud, D. J. Arndtjovin, and T. M. Jovin, "Studying single living cells and chromosomes by Confocal Raman Microspectroscopy," Nature 347, 301-303 (1990).
[CrossRef] [PubMed]

Becker, W.

W. Becker, A. Bergmann, M. A. Hink, K. Konig, K. Benndorf, and C. Biskup, "Fluorescence lifetime imaging by time-correlated single-photon counting," Microsc. Res. Technique 63, 58-66 (2004).
[CrossRef]

Benndorf, K.

W. Becker, A. Bergmann, M. A. Hink, K. Konig, K. Benndorf, and C. Biskup, "Fluorescence lifetime imaging by time-correlated single-photon counting," Microsc. Res. Technique 63, 58-66 (2004).
[CrossRef]

Bergmann, A.

W. Becker, A. Bergmann, M. A. Hink, K. Konig, K. Benndorf, and C. Biskup, "Fluorescence lifetime imaging by time-correlated single-photon counting," Microsc. Res. Technique 63, 58-66 (2004).
[CrossRef]

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, 1-12 (2004).
[CrossRef] [PubMed]

Biskup, C.

W. Becker, A. Bergmann, M. A. Hink, K. Konig, K. Benndorf, and C. Biskup, "Fluorescence lifetime imaging by time-correlated single-photon counting," Microsc. Res. Technique 63, 58-66 (2004).
[CrossRef]

Brackmann, C.

Burkacky, O.

Chan, J. W.

J. W. Chan, A. P. Esposito, C. E. Talley, C. W. Hollars, S. M. Lane, and T. Huser, "Reagentless identification of single bacterial spores in aqueous solution by confocal laser tweezers Raman spectroscopy," Anal. Chem. 76, 599-603 (2004).
[CrossRef] [PubMed]

Cheng, J. X.

T. B. Huff and J. X. Cheng, "In vivo coherent anti-Stokes Raman scattering imaging of sciatic nerve tissue," J. Microsc. 225, 175-182 (2007).
[CrossRef] [PubMed]

L. Tong, Y. Lu, R. J. Lee, and J. X. Cheng, "Imaging receptor-mediated endocytosis with a polymeric nanoparticle-based coherent anti-stokes raman scattering probe," J. Phys. Chem. B 111, 9980-9985 (2007).
[CrossRef] [PubMed]

H. F. Wang, Y. Fu, P. Zickmund, R. Y. Shi, and J. X. Cheng, "Coherent anti-stokes Raman scattering imaging of axonal myelin in live spinal tissues," Biophys. J. 89, 581-591 (2005).
[CrossRef] [PubMed]

J. X. Cheng and X. S. Xie, "Coherent anti-Stokes Raman scattering microscopy: Instrumentation, theory, and applications," J. Phys. Chem. B 108, 827-840 (2004).
[CrossRef]

J. X. Cheng, A. Volkmer, and X. S. Xie, "Theoretical and experimental characterization of Coherent anti- Stokes Raman Scattering Microscopy," J. Opt. Soc. Am. B 19, 1363-1375 (2002)
[CrossRef]

J. X. Cheng, Y. K. Jia, G. F. Zheng, and X. S. Xie, "Laser-scanning coherent anti-stokes RamanScattering Microscopy and applications to cell biology," Biophys. J. 83, 502-509 (2002).
[CrossRef] [PubMed]

A. Volkmer, J. X. Cheng, and X. S. Xie, "Vibrational imaging with high sensitivity via epidetected coherent anti-Stokes Raman scattering microscopy," Phys. Rev. Lett. 8702, 4 (2001).

Cohn, K.

Colonna, A.

N. Djaker, P. F. Lenne, D. Marguet, A. Colonna, C. Hadjur, and H. Rigneault, "Coherent anti-Stokes Raman scattering microscopy (CARS): Instrumentation and applications," Nuc. Instr. Meth. Phys. 571, 177-181 (2007).
[CrossRef]

Cote, D.

C. L. Evans, E. O. Potma, M. Puoris'haag, D. Cote, C. P. Lin, and X. S. Xie, "Chemical imaging of tissue in vivo with video-rate coherent anti-Stokes Raman scattering microscopy," PNAS 102, 16807-16812 (2005).
[CrossRef] [PubMed]

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, 1-12 (2004).
[CrossRef] [PubMed]

Crane, N. J.

N. J. Crane, M. D. Morris, M. A. Ignelzi, and G. G. Yu, "Raman imaging demonstrates FGF2-induced craniosynostosis in mouse calvaria," J. Biomed. Opt. 10, 8 (2005).
[CrossRef]

de Bruijn, H. S.

J. A. Palero, H. S. de Bruijn, A. V. van den Heuvel, H. Sterenborg, and H. C. Gerritsen, "Spectrally resolved multiphoton imaging of in vivo and excised mouse skin tissues," Biophys. J. 93, 992-1007 (2007).
[CrossRef] [PubMed]

J. A. Palero, H. S. de Bruijn, A. van der Ploeg-van den Heuvel, H. Sterenborg, and H. C. Gerritsen, "In vivo nonlinear spectral imaging in mouse skin," Opt. Express 14, 4395-4402 (2006).
[CrossRef] [PubMed]

Demul, F. F. M.

G. J. Puppels, F. F. M. Demul, C. Otto, J. Greve, M. Robertnicoud, D. J. Arndtjovin, and T. M. Jovin, "Studying single living cells and chromosomes by Confocal Raman Microspectroscopy," Nature 347, 301-303 (1990).
[CrossRef] [PubMed]

Djaker, N.

N. Djaker, P. F. Lenne, D. Marguet, A. Colonna, C. Hadjur, and H. Rigneault, "Coherent anti-Stokes Raman scattering microscopy (CARS): Instrumentation and applications," Nuc. Instr. Meth. Phys. 571, 177-181 (2007).
[CrossRef]

Duncan, M. D.

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, 1-12 (2004).
[CrossRef] [PubMed]

Eickhoff, V. C.

M. C. Skala, J. M. Squirrell, K. M. Vrotsos, V. C. Eickhoff, A. Gendron-Fitzpatrick, K. W. Eliceiri, and N. Ramanujam, "Multiphoton microscopy of endogenous fluorescence differentiates normal, precancerous, and cancerous squamous epithelial tissues," Cancer Res. 65, 1180-1186 (2005).
[CrossRef] [PubMed]

Eliceiri, K. W.

M. C. Skala, J. M. Squirrell, K. M. Vrotsos, V. C. Eickhoff, A. Gendron-Fitzpatrick, K. W. Eliceiri, and N. Ramanujam, "Multiphoton microscopy of endogenous fluorescence differentiates normal, precancerous, and cancerous squamous epithelial tissues," Cancer Res. 65, 1180-1186 (2005).
[CrossRef] [PubMed]

Enejder, A.

Esposito, A. P.

J. W. Chan, A. P. Esposito, C. E. Talley, C. W. Hollars, S. M. Lane, and T. Huser, "Reagentless identification of single bacterial spores in aqueous solution by confocal laser tweezers Raman spectroscopy," Anal. Chem. 76, 599-603 (2004).
[CrossRef] [PubMed]

Evans, C. L.

E. O. Potma, C. L. Evans, and X. S. Xie, "Heterodyne coherent anti-Stokes Raman scattering (CARS) imaging," Opt. Lett. 31, 241-243 (2006).
[CrossRef] [PubMed]

C. L. Evans, E. O. Potma, M. Puoris'haag, D. Cote, C. P. Lin, and X. S. Xie, "Chemical imaging of tissue in vivo with video-rate coherent anti-Stokes Raman scattering microscopy," PNAS 102, 16807-16812 (2005).
[CrossRef] [PubMed]

Flach, C. R.

G. J. Zhang, D. J. Moore, C. R. Flach, and R. Mendelsohn, "Vibrational microscopy and imaging of skin: from single cells to intact tissue," Anal. Bioanal. Chem. 387, 1591-1599 (2007).
[CrossRef]

G. J. Zhang, D. J. Moore, K. B. Sloan, C. R. Flach, and R. Mendelsohn, "Imaging the prodrug-to-drug transformation of a 5-fluorouracil derivative in skin by confocal Raman microscopy," J. Invest. Dermatol. 127, 1205-1209 (2007).
[CrossRef] [PubMed]

Fratzl, P.

M. Kazanci, H. D. Wagner, N. I. Manjubala, H. S. Gupta, E. Paschalis, P. Roschger, and P. Fratzl, "Raman imaging of two orthogonal planes within cortical bone," Bone 41, 456-461 (2007).
[CrossRef] [PubMed]

French, P. M. W.

K. Suhling, P. M. W. French, and D. Phillips, "Time-resolved fluorescence microscopy," Photochem. Photobiol. Sci. 4, 13-22 (2005).
[CrossRef]

Fu, Y.

H. F. Wang, Y. Fu, P. Zickmund, R. Y. Shi, and J. X. Cheng, "Coherent anti-stokes Raman scattering imaging of axonal myelin in live spinal tissues," Biophys. J. 89, 581-591 (2005).
[CrossRef] [PubMed]

Gendron-Fitzpatrick, A.

M. C. Skala, J. M. Squirrell, K. M. Vrotsos, V. C. Eickhoff, A. Gendron-Fitzpatrick, K. W. Eliceiri, and N. Ramanujam, "Multiphoton microscopy of endogenous fluorescence differentiates normal, precancerous, and cancerous squamous epithelial tissues," Cancer Res. 65, 1180-1186 (2005).
[CrossRef] [PubMed]

Gerritsen, H. C.

J. A. Palero, H. S. de Bruijn, A. V. van den Heuvel, H. Sterenborg, and H. C. Gerritsen, "Spectrally resolved multiphoton imaging of in vivo and excised mouse skin tissues," Biophys. J. 93, 992-1007 (2007).
[CrossRef] [PubMed]

J. A. Palero, H. S. de Bruijn, A. van der Ploeg-van den Heuvel, H. Sterenborg, and H. C. Gerritsen, "In vivo nonlinear spectral imaging in mouse skin," Opt. Express 14, 4395-4402 (2006).
[CrossRef] [PubMed]

Greve, J.

G. J. Puppels, F. F. M. Demul, C. Otto, J. Greve, M. Robertnicoud, D. J. Arndtjovin, and T. M. Jovin, "Studying single living cells and chromosomes by Confocal Raman Microspectroscopy," Nature 347, 301-303 (1990).
[CrossRef] [PubMed]

Gupta, H. S.

M. Kazanci, H. D. Wagner, N. I. Manjubala, H. S. Gupta, E. Paschalis, P. Roschger, and P. Fratzl, "Raman imaging of two orthogonal planes within cortical bone," Bone 41, 456-461 (2007).
[CrossRef] [PubMed]

Hadjur, C.

N. Djaker, P. F. Lenne, D. Marguet, A. Colonna, C. Hadjur, and H. Rigneault, "Coherent anti-Stokes Raman scattering microscopy (CARS): Instrumentation and applications," Nuc. Instr. Meth. Phys. 571, 177-181 (2007).
[CrossRef]

Hamaguchi, H.

Hink, M. A.

W. Becker, A. Bergmann, M. A. Hink, K. Konig, K. Benndorf, and C. Biskup, "Fluorescence lifetime imaging by time-correlated single-photon counting," Microsc. Res. Technique 63, 58-66 (2004).
[CrossRef]

Hollars, C. W.

J. W. Chan, A. P. Esposito, C. E. Talley, C. W. Hollars, S. M. Lane, and T. Huser, "Reagentless identification of single bacterial spores in aqueous solution by confocal laser tweezers Raman spectroscopy," Anal. Chem. 76, 599-603 (2004).
[CrossRef] [PubMed]

Holtom, G. R.

L. G. Rodriguez, S. J. Lockett, and G. R. Holtom, "Coherent anti-stokes Raman scattering microscopy: A biological review," Cytometry A 69A, 779-791 (2006).
[CrossRef]

A. Zumbusch, G. R. Holtom, and X. S. Xie, "Three-dimensional vibrational imaging by coherent anti- Stokes Raman scattering," Phys. Rev. Lett. 82, 4142-4145 (1999).
[CrossRef]

Huff, T. B.

T. B. Huff and J. X. Cheng, "In vivo coherent anti-Stokes Raman scattering imaging of sciatic nerve tissue," J. Microsc. 225, 175-182 (2007).
[CrossRef] [PubMed]

Huser, T.

J. W. Chan, A. P. Esposito, C. E. Talley, C. W. Hollars, S. M. Lane, and T. Huser, "Reagentless identification of single bacterial spores in aqueous solution by confocal laser tweezers Raman spectroscopy," Anal. Chem. 76, 599-603 (2004).
[CrossRef] [PubMed]

Ignelzi, M. A.

N. J. Crane, M. D. Morris, M. A. Ignelzi, and G. G. Yu, "Raman imaging demonstrates FGF2-induced craniosynostosis in mouse calvaria," J. Biomed. Opt. 10, 8 (2005).
[CrossRef]

Jia, Y. K.

J. X. Cheng, Y. K. Jia, G. F. Zheng, and X. S. Xie, "Laser-scanning coherent anti-stokes RamanScattering Microscopy and applications to cell biology," Biophys. J. 83, 502-509 (2002).
[CrossRef] [PubMed]

Jovin, T. M.

G. J. Puppels, F. F. M. Demul, C. Otto, J. Greve, M. Robertnicoud, D. J. Arndtjovin, and T. M. Jovin, "Studying single living cells and chromosomes by Confocal Raman Microspectroscopy," Nature 347, 301-303 (1990).
[CrossRef] [PubMed]

Kano, H.

Kazanci, M.

M. Kazanci, H. D. Wagner, N. I. Manjubala, H. S. Gupta, E. Paschalis, P. Roschger, and P. Fratzl, "Raman imaging of two orthogonal planes within cortical bone," Bone 41, 456-461 (2007).
[CrossRef] [PubMed]

Konig, K.

K. Konig, K. Schenke-Layland, I. Riemann, and U. A. Stock, "Multiphoton autofluorescence imaging of intratissue elastic fibers," Biomaterials 26, 495-500 (2005).
[CrossRef]

W. Becker, A. Bergmann, M. A. Hink, K. Konig, K. Benndorf, and C. Biskup, "Fluorescence lifetime imaging by time-correlated single-photon counting," Microsc. Res. Technique 63, 58-66 (2004).
[CrossRef]

K. Konig and I. Riemann, "High-resolution multiphoton tomography of human skin with subcellular spatial resolution and picosecond time resolution," J. Biomed. Opt. 8, 432-439 (2003).
[CrossRef] [PubMed]

Lane, S. M.

J. W. Chan, A. P. Esposito, C. E. Talley, C. W. Hollars, S. M. Lane, and T. Huser, "Reagentless identification of single bacterial spores in aqueous solution by confocal laser tweezers Raman spectroscopy," Anal. Chem. 76, 599-603 (2004).
[CrossRef] [PubMed]

Lee, E. S.

E. S. Lee, J. Y. Lee, and Y. S. Yoo, "Nonlinear optical interference of two successive coherent anti-Stokes Raman scattering signals for biological imaging applications," J. Biomed. Opt. 12, 5 (2007).
[CrossRef]

Lee, J. Y.

E. S. Lee, J. Y. Lee, and Y. S. Yoo, "Nonlinear optical interference of two successive coherent anti-Stokes Raman scattering signals for biological imaging applications," J. Biomed. Opt. 12, 5 (2007).
[CrossRef]

Lee, R. J.

L. Tong, Y. Lu, R. J. Lee, and J. X. Cheng, "Imaging receptor-mediated endocytosis with a polymeric nanoparticle-based coherent anti-stokes raman scattering probe," J. Phys. Chem. B 111, 9980-9985 (2007).
[CrossRef] [PubMed]

Lenne, P. F.

N. Djaker, P. F. Lenne, D. Marguet, A. Colonna, C. Hadjur, and H. Rigneault, "Coherent anti-Stokes Raman scattering microscopy (CARS): Instrumentation and applications," Nuc. Instr. Meth. Phys. 571, 177-181 (2007).
[CrossRef]

Lin, C. P.

C. L. Evans, E. O. Potma, M. Puoris'haag, D. Cote, C. P. Lin, and X. S. Xie, "Chemical imaging of tissue in vivo with video-rate coherent anti-Stokes Raman scattering microscopy," PNAS 102, 16807-16812 (2005).
[CrossRef] [PubMed]

Liu, C. Y.

J. M. Piepmeier, C. Y. Liu, and E. Neuwelt, "Multiphoton excitation of autofluorescence for microscopy of glioma tissue - Comments," Neurosurgery 58, 767-767 (2006).

Lockett, S. J.

L. G. Rodriguez, S. J. Lockett, and G. R. Holtom, "Coherent anti-stokes Raman scattering microscopy: A biological review," Cytometry A 69A, 779-791 (2006).
[CrossRef]

Lu, Y.

L. Tong, Y. Lu, R. J. Lee, and J. X. Cheng, "Imaging receptor-mediated endocytosis with a polymeric nanoparticle-based coherent anti-stokes raman scattering probe," J. Phys. Chem. B 111, 9980-9985 (2007).
[CrossRef] [PubMed]

Manjubala, N. I.

M. Kazanci, H. D. Wagner, N. I. Manjubala, H. S. Gupta, E. Paschalis, P. Roschger, and P. Fratzl, "Raman imaging of two orthogonal planes within cortical bone," Bone 41, 456-461 (2007).
[CrossRef] [PubMed]

Manuccia, T. J.

Marguet, D.

N. Djaker, P. F. Lenne, D. Marguet, A. Colonna, C. Hadjur, and H. Rigneault, "Coherent anti-Stokes Raman scattering microscopy (CARS): Instrumentation and applications," Nuc. Instr. Meth. Phys. 571, 177-181 (2007).
[CrossRef]

Mendelsohn, R.

G. J. Zhang, D. J. Moore, C. R. Flach, and R. Mendelsohn, "Vibrational microscopy and imaging of skin: from single cells to intact tissue," Anal. Bioanal. Chem. 387, 1591-1599 (2007).
[CrossRef]

G. J. Zhang, D. J. Moore, K. B. Sloan, C. R. Flach, and R. Mendelsohn, "Imaging the prodrug-to-drug transformation of a 5-fluorouracil derivative in skin by confocal Raman microscopy," J. Invest. Dermatol. 127, 1205-1209 (2007).
[CrossRef] [PubMed]

Moore, D. J.

G. J. Zhang, D. J. Moore, K. B. Sloan, C. R. Flach, and R. Mendelsohn, "Imaging the prodrug-to-drug transformation of a 5-fluorouracil derivative in skin by confocal Raman microscopy," J. Invest. Dermatol. 127, 1205-1209 (2007).
[CrossRef] [PubMed]

G. J. Zhang, D. J. Moore, C. R. Flach, and R. Mendelsohn, "Vibrational microscopy and imaging of skin: from single cells to intact tissue," Anal. Bioanal. Chem. 387, 1591-1599 (2007).
[CrossRef]

Morris, M. D.

N. J. Crane, M. D. Morris, M. A. Ignelzi, and G. G. Yu, "Raman imaging demonstrates FGF2-induced craniosynostosis in mouse calvaria," J. Biomed. Opt. 10, 8 (2005).
[CrossRef]

Nan, X. L.

X. L. Nan, E. O. Potma, and X. S. Xie, "Nonperturbative chemical imaging of organelle transport in living cells with coherent anti-stokes Raman Scattering Microscopy," Biophys. J. 91, 728-735 (2006).
[CrossRef] [PubMed]

Neuwelt, E.

J. M. Piepmeier, C. Y. Liu, and E. Neuwelt, "Multiphoton excitation of autofluorescence for microscopy of glioma tissue - Comments," Neurosurgery 58, 767-767 (2006).

Otto, C.

G. J. Puppels, F. F. M. Demul, C. Otto, J. Greve, M. Robertnicoud, D. J. Arndtjovin, and T. M. Jovin, "Studying single living cells and chromosomes by Confocal Raman Microspectroscopy," Nature 347, 301-303 (1990).
[CrossRef] [PubMed]

Palanker, D.

Palero, J. A.

J. A. Palero, H. S. de Bruijn, A. V. van den Heuvel, H. Sterenborg, and H. C. Gerritsen, "Spectrally resolved multiphoton imaging of in vivo and excised mouse skin tissues," Biophys. J. 93, 992-1007 (2007).
[CrossRef] [PubMed]

J. A. Palero, H. S. de Bruijn, A. van der Ploeg-van den Heuvel, H. Sterenborg, and H. C. Gerritsen, "In vivo nonlinear spectral imaging in mouse skin," Opt. Express 14, 4395-4402 (2006).
[CrossRef] [PubMed]

Paschalis, E.

M. Kazanci, H. D. Wagner, N. I. Manjubala, H. S. Gupta, E. Paschalis, P. Roschger, and P. Fratzl, "Raman imaging of two orthogonal planes within cortical bone," Bone 41, 456-461 (2007).
[CrossRef] [PubMed]

Peter, M.

M. Peter, and S. M. Ameer-Beg, "Imaging molecular interactions by multiphoton FLIM," Biology of the Cell 96, 231-236 (2004).
[CrossRef] [PubMed]

Phillips, D.

K. Suhling, P. M. W. French, and D. Phillips, "Time-resolved fluorescence microscopy," Photochem. Photobiol. Sci. 4, 13-22 (2005).
[CrossRef]

Piepmeier, J. M.

J. M. Piepmeier, C. Y. Liu, and E. Neuwelt, "Multiphoton excitation of autofluorescence for microscopy of glioma tissue - Comments," Neurosurgery 58, 767-767 (2006).

Potma, E. O.

E. O. Potma, C. L. Evans, and X. S. Xie, "Heterodyne coherent anti-Stokes Raman scattering (CARS) imaging," Opt. Lett. 31, 241-243 (2006).
[CrossRef] [PubMed]

X. L. Nan, E. O. Potma, and X. S. Xie, "Nonperturbative chemical imaging of organelle transport in living cells with coherent anti-stokes Raman Scattering Microscopy," Biophys. J. 91, 728-735 (2006).
[CrossRef] [PubMed]

C. L. Evans, E. O. Potma, M. Puoris'haag, D. Cote, C. P. Lin, and X. S. Xie, "Chemical imaging of tissue in vivo with video-rate coherent anti-Stokes Raman scattering microscopy," PNAS 102, 16807-16812 (2005).
[CrossRef] [PubMed]

Puoris'haag, M.

C. L. Evans, E. O. Potma, M. Puoris'haag, D. Cote, C. P. Lin, and X. S. Xie, "Chemical imaging of tissue in vivo with video-rate coherent anti-Stokes Raman scattering microscopy," PNAS 102, 16807-16812 (2005).
[CrossRef] [PubMed]

Puppels, G. J.

G. J. Puppels, F. F. M. Demul, C. Otto, J. Greve, M. Robertnicoud, D. J. Arndtjovin, and T. M. Jovin, "Studying single living cells and chromosomes by Confocal Raman Microspectroscopy," Nature 347, 301-303 (1990).
[CrossRef] [PubMed]

Ramanujam, N.

M. C. Skala, J. M. Squirrell, K. M. Vrotsos, V. C. Eickhoff, A. Gendron-Fitzpatrick, K. W. Eliceiri, and N. Ramanujam, "Multiphoton microscopy of endogenous fluorescence differentiates normal, precancerous, and cancerous squamous epithelial tissues," Cancer Res. 65, 1180-1186 (2005).
[CrossRef] [PubMed]

Reintjes, J.

Riemann, I.

K. Konig, K. Schenke-Layland, I. Riemann, and U. A. Stock, "Multiphoton autofluorescence imaging of intratissue elastic fibers," Biomaterials 26, 495-500 (2005).
[CrossRef]

K. Konig and I. Riemann, "High-resolution multiphoton tomography of human skin with subcellular spatial resolution and picosecond time resolution," J. Biomed. Opt. 8, 432-439 (2003).
[CrossRef] [PubMed]

Rigneault, H.

N. Djaker, P. F. Lenne, D. Marguet, A. Colonna, C. Hadjur, and H. Rigneault, "Coherent anti-Stokes Raman scattering microscopy (CARS): Instrumentation and applications," Nuc. Instr. Meth. Phys. 571, 177-181 (2007).
[CrossRef]

Robertnicoud, M.

G. J. Puppels, F. F. M. Demul, C. Otto, J. Greve, M. Robertnicoud, D. J. Arndtjovin, and T. M. Jovin, "Studying single living cells and chromosomes by Confocal Raman Microspectroscopy," Nature 347, 301-303 (1990).
[CrossRef] [PubMed]

Rodriguez, L. G.

L. G. Rodriguez, S. J. Lockett, and G. R. Holtom, "Coherent anti-stokes Raman scattering microscopy: A biological review," Cytometry A 69A, 779-791 (2006).
[CrossRef]

Roschger, P.

M. Kazanci, H. D. Wagner, N. I. Manjubala, H. S. Gupta, E. Paschalis, P. Roschger, and P. Fratzl, "Raman imaging of two orthogonal planes within cortical bone," Bone 41, 456-461 (2007).
[CrossRef] [PubMed]

Schenke-Layland, K.

K. Konig, K. Schenke-Layland, I. Riemann, and U. A. Stock, "Multiphoton autofluorescence imaging of intratissue elastic fibers," Biomaterials 26, 495-500 (2005).
[CrossRef]

Shi, R. Y.

H. F. Wang, Y. Fu, P. Zickmund, R. Y. Shi, and J. X. Cheng, "Coherent anti-stokes Raman scattering imaging of axonal myelin in live spinal tissues," Biophys. J. 89, 581-591 (2005).
[CrossRef] [PubMed]

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, 1-12 (2004).
[CrossRef] [PubMed]

Simanovskii, D.

Skala, M. C.

M. C. Skala, J. M. Squirrell, K. M. Vrotsos, V. C. Eickhoff, A. Gendron-Fitzpatrick, K. W. Eliceiri, and N. Ramanujam, "Multiphoton microscopy of endogenous fluorescence differentiates normal, precancerous, and cancerous squamous epithelial tissues," Cancer Res. 65, 1180-1186 (2005).
[CrossRef] [PubMed]

Sloan, K. B.

G. J. Zhang, D. J. Moore, K. B. Sloan, C. R. Flach, and R. Mendelsohn, "Imaging the prodrug-to-drug transformation of a 5-fluorouracil derivative in skin by confocal Raman microscopy," J. Invest. Dermatol. 127, 1205-1209 (2007).
[CrossRef] [PubMed]

Smith, T.

Squirrell, J. M.

M. C. Skala, J. M. Squirrell, K. M. Vrotsos, V. C. Eickhoff, A. Gendron-Fitzpatrick, K. W. Eliceiri, and N. Ramanujam, "Multiphoton microscopy of endogenous fluorescence differentiates normal, precancerous, and cancerous squamous epithelial tissues," Cancer Res. 65, 1180-1186 (2005).
[CrossRef] [PubMed]

Sterenborg, H.

J. A. Palero, H. S. de Bruijn, A. V. van den Heuvel, H. Sterenborg, and H. C. Gerritsen, "Spectrally resolved multiphoton imaging of in vivo and excised mouse skin tissues," Biophys. J. 93, 992-1007 (2007).
[CrossRef] [PubMed]

J. A. Palero, H. S. de Bruijn, A. van der Ploeg-van den Heuvel, H. Sterenborg, and H. C. Gerritsen, "In vivo nonlinear spectral imaging in mouse skin," Opt. Express 14, 4395-4402 (2006).
[CrossRef] [PubMed]

Stock, U. A.

K. Konig, K. Schenke-Layland, I. Riemann, and U. A. Stock, "Multiphoton autofluorescence imaging of intratissue elastic fibers," Biomaterials 26, 495-500 (2005).
[CrossRef]

Suhling, K.

K. Suhling, P. M. W. French, and D. Phillips, "Time-resolved fluorescence microscopy," Photochem. Photobiol. Sci. 4, 13-22 (2005).
[CrossRef]

Talley, C. E.

J. W. Chan, A. P. Esposito, C. E. Talley, C. W. Hollars, S. M. Lane, and T. Huser, "Reagentless identification of single bacterial spores in aqueous solution by confocal laser tweezers Raman spectroscopy," Anal. Chem. 76, 599-603 (2004).
[CrossRef] [PubMed]

Tong, L.

L. Tong, Y. Lu, R. J. Lee, and J. X. Cheng, "Imaging receptor-mediated endocytosis with a polymeric nanoparticle-based coherent anti-stokes raman scattering probe," J. Phys. Chem. B 111, 9980-9985 (2007).
[CrossRef] [PubMed]

Toytman, I.

van den Heuvel, A. V.

J. A. Palero, H. S. de Bruijn, A. V. van den Heuvel, H. Sterenborg, and H. C. Gerritsen, "Spectrally resolved multiphoton imaging of in vivo and excised mouse skin tissues," Biophys. J. 93, 992-1007 (2007).
[CrossRef] [PubMed]

van der Ploeg-van den Heuvel, A.

Volkmer, A.

J. X. Cheng, A. Volkmer, and X. S. Xie, "Theoretical and experimental characterization of Coherent anti- Stokes Raman Scattering Microscopy," J. Opt. Soc. Am. B 19, 1363-1375 (2002)
[CrossRef]

A. Volkmer, J. X. Cheng, and X. S. Xie, "Vibrational imaging with high sensitivity via epidetected coherent anti-Stokes Raman scattering microscopy," Phys. Rev. Lett. 8702, 4 (2001).

Vrotsos, K. M.

M. C. Skala, J. M. Squirrell, K. M. Vrotsos, V. C. Eickhoff, A. Gendron-Fitzpatrick, K. W. Eliceiri, and N. Ramanujam, "Multiphoton microscopy of endogenous fluorescence differentiates normal, precancerous, and cancerous squamous epithelial tissues," Cancer Res. 65, 1180-1186 (2005).
[CrossRef] [PubMed]

Wagner, H. D.

M. Kazanci, H. D. Wagner, N. I. Manjubala, H. S. Gupta, E. Paschalis, P. Roschger, and P. Fratzl, "Raman imaging of two orthogonal planes within cortical bone," Bone 41, 456-461 (2007).
[CrossRef] [PubMed]

Wang, H. F.

H. F. Wang, Y. Fu, P. Zickmund, R. Y. Shi, and J. X. Cheng, "Coherent anti-stokes Raman scattering imaging of axonal myelin in live spinal tissues," Biophys. J. 89, 581-591 (2005).
[CrossRef] [PubMed]

Xie, X. S.

E. O. Potma, C. L. Evans, and X. S. Xie, "Heterodyne coherent anti-Stokes Raman scattering (CARS) imaging," Opt. Lett. 31, 241-243 (2006).
[CrossRef] [PubMed]

X. L. Nan, E. O. Potma, and X. S. Xie, "Nonperturbative chemical imaging of organelle transport in living cells with coherent anti-stokes Raman Scattering Microscopy," Biophys. J. 91, 728-735 (2006).
[CrossRef] [PubMed]

C. L. Evans, E. O. Potma, M. Puoris'haag, D. Cote, C. P. Lin, and X. S. Xie, "Chemical imaging of tissue in vivo with video-rate coherent anti-Stokes Raman scattering microscopy," PNAS 102, 16807-16812 (2005).
[CrossRef] [PubMed]

J. X. Cheng and X. S. Xie, "Coherent anti-Stokes Raman scattering microscopy: Instrumentation, theory, and applications," J. Phys. Chem. B 108, 827-840 (2004).
[CrossRef]

J. X. Cheng, A. Volkmer, and X. S. Xie, "Theoretical and experimental characterization of Coherent anti- Stokes Raman Scattering Microscopy," J. Opt. Soc. Am. B 19, 1363-1375 (2002)
[CrossRef]

J. X. Cheng, Y. K. Jia, G. F. Zheng, and X. S. Xie, "Laser-scanning coherent anti-stokes RamanScattering Microscopy and applications to cell biology," Biophys. J. 83, 502-509 (2002).
[CrossRef] [PubMed]

A. Volkmer, J. X. Cheng, and X. S. Xie, "Vibrational imaging with high sensitivity via epidetected coherent anti-Stokes Raman scattering microscopy," Phys. Rev. Lett. 8702, 4 (2001).

A. Zumbusch, G. R. Holtom, and X. S. Xie, "Three-dimensional vibrational imaging by coherent anti- Stokes Raman scattering," Phys. Rev. Lett. 82, 4142-4145 (1999).
[CrossRef]

Yoo, Y. S.

E. S. Lee, J. Y. Lee, and Y. S. Yoo, "Nonlinear optical interference of two successive coherent anti-Stokes Raman scattering signals for biological imaging applications," J. Biomed. Opt. 12, 5 (2007).
[CrossRef]

Yu, G. G.

N. J. Crane, M. D. Morris, M. A. Ignelzi, and G. G. Yu, "Raman imaging demonstrates FGF2-induced craniosynostosis in mouse calvaria," J. Biomed. Opt. 10, 8 (2005).
[CrossRef]

Zhang, G. J.

G. J. Zhang, D. J. Moore, C. R. Flach, and R. Mendelsohn, "Vibrational microscopy and imaging of skin: from single cells to intact tissue," Anal. Bioanal. Chem. 387, 1591-1599 (2007).
[CrossRef]

G. J. Zhang, D. J. Moore, K. B. Sloan, C. R. Flach, and R. Mendelsohn, "Imaging the prodrug-to-drug transformation of a 5-fluorouracil derivative in skin by confocal Raman microscopy," J. Invest. Dermatol. 127, 1205-1209 (2007).
[CrossRef] [PubMed]

Zheng, G. F.

J. X. Cheng, Y. K. Jia, G. F. Zheng, and X. S. Xie, "Laser-scanning coherent anti-stokes RamanScattering Microscopy and applications to cell biology," Biophys. J. 83, 502-509 (2002).
[CrossRef] [PubMed]

Zickmund, P.

H. F. Wang, Y. Fu, P. Zickmund, R. Y. Shi, and J. X. Cheng, "Coherent anti-stokes Raman scattering imaging of axonal myelin in live spinal tissues," Biophys. J. 89, 581-591 (2005).
[CrossRef] [PubMed]

Zumbusch, A.

O. Burkacky, A. Zumbusch, C. Brackmann, and A. Enejder, "Dual-pump coherent anti-Stokes-Raman Scattering Microscopy," Opt. Lett. 31, 3656-3658 (2006).
[CrossRef] [PubMed]

A. Zumbusch, G. R. Holtom, and X. S. Xie, "Three-dimensional vibrational imaging by coherent anti- Stokes Raman scattering," Phys. Rev. Lett. 82, 4142-4145 (1999).
[CrossRef]

Anal. Bioanal. Chem. (1)

G. J. Zhang, D. J. Moore, C. R. Flach, and R. Mendelsohn, "Vibrational microscopy and imaging of skin: from single cells to intact tissue," Anal. Bioanal. Chem. 387, 1591-1599 (2007).
[CrossRef]

Anal. Chem. (1)

J. W. Chan, A. P. Esposito, C. E. Talley, C. W. Hollars, S. M. Lane, and T. Huser, "Reagentless identification of single bacterial spores in aqueous solution by confocal laser tweezers Raman spectroscopy," Anal. Chem. 76, 599-603 (2004).
[CrossRef] [PubMed]

Biology of the Cell (1)

M. Peter, and S. M. Ameer-Beg, "Imaging molecular interactions by multiphoton FLIM," Biology of the Cell 96, 231-236 (2004).
[CrossRef] [PubMed]

Biomaterials (1)

K. Konig, K. Schenke-Layland, I. Riemann, and U. A. Stock, "Multiphoton autofluorescence imaging of intratissue elastic fibers," Biomaterials 26, 495-500 (2005).
[CrossRef]

Biophys. J. (4)

J. A. Palero, H. S. de Bruijn, A. V. van den Heuvel, H. Sterenborg, and H. C. Gerritsen, "Spectrally resolved multiphoton imaging of in vivo and excised mouse skin tissues," Biophys. J. 93, 992-1007 (2007).
[CrossRef] [PubMed]

X. L. Nan, E. O. Potma, and X. S. Xie, "Nonperturbative chemical imaging of organelle transport in living cells with coherent anti-stokes Raman Scattering Microscopy," Biophys. J. 91, 728-735 (2006).
[CrossRef] [PubMed]

H. F. Wang, Y. Fu, P. Zickmund, R. Y. Shi, and J. X. Cheng, "Coherent anti-stokes Raman scattering imaging of axonal myelin in live spinal tissues," Biophys. J. 89, 581-591 (2005).
[CrossRef] [PubMed]

J. X. Cheng, Y. K. Jia, G. F. Zheng, and X. S. Xie, "Laser-scanning coherent anti-stokes RamanScattering Microscopy and applications to cell biology," Biophys. J. 83, 502-509 (2002).
[CrossRef] [PubMed]

Bone (1)

M. Kazanci, H. D. Wagner, N. I. Manjubala, H. S. Gupta, E. Paschalis, P. Roschger, and P. Fratzl, "Raman imaging of two orthogonal planes within cortical bone," Bone 41, 456-461 (2007).
[CrossRef] [PubMed]

Cancer Res. (1)

M. C. Skala, J. M. Squirrell, K. M. Vrotsos, V. C. Eickhoff, A. Gendron-Fitzpatrick, K. W. Eliceiri, and N. Ramanujam, "Multiphoton microscopy of endogenous fluorescence differentiates normal, precancerous, and cancerous squamous epithelial tissues," Cancer Res. 65, 1180-1186 (2005).
[CrossRef] [PubMed]

Cytometry A (1)

L. G. Rodriguez, S. J. Lockett, and G. R. Holtom, "Coherent anti-stokes Raman scattering microscopy: A biological review," Cytometry A 69A, 779-791 (2006).
[CrossRef]

J. Biomed. Opt. (3)

N. J. Crane, M. D. Morris, M. A. Ignelzi, and G. G. Yu, "Raman imaging demonstrates FGF2-induced craniosynostosis in mouse calvaria," J. Biomed. Opt. 10, 8 (2005).
[CrossRef]

E. S. Lee, J. Y. Lee, and Y. S. Yoo, "Nonlinear optical interference of two successive coherent anti-Stokes Raman scattering signals for biological imaging applications," J. Biomed. Opt. 12, 5 (2007).
[CrossRef]

K. Konig and I. Riemann, "High-resolution multiphoton tomography of human skin with subcellular spatial resolution and picosecond time resolution," J. Biomed. Opt. 8, 432-439 (2003).
[CrossRef] [PubMed]

J. Invest. Dermatol. (1)

G. J. Zhang, D. J. Moore, K. B. Sloan, C. R. Flach, and R. Mendelsohn, "Imaging the prodrug-to-drug transformation of a 5-fluorouracil derivative in skin by confocal Raman microscopy," J. Invest. Dermatol. 127, 1205-1209 (2007).
[CrossRef] [PubMed]

J. Microsc. (2)

T. B. Huff and J. X. Cheng, "In vivo coherent anti-Stokes Raman scattering imaging of sciatic nerve tissue," J. Microsc. 225, 175-182 (2007).
[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, 1-12 (2004).
[CrossRef] [PubMed]

J. Opt. Soc. Am. B (1)

J. Phys. Chem. B (2)

L. Tong, Y. Lu, R. J. Lee, and J. X. Cheng, "Imaging receptor-mediated endocytosis with a polymeric nanoparticle-based coherent anti-stokes raman scattering probe," J. Phys. Chem. B 111, 9980-9985 (2007).
[CrossRef] [PubMed]

J. X. Cheng and X. S. Xie, "Coherent anti-Stokes Raman scattering microscopy: Instrumentation, theory, and applications," J. Phys. Chem. B 108, 827-840 (2004).
[CrossRef]

Microsc. Res. and Technique (1)

W. Becker, A. Bergmann, M. A. Hink, K. Konig, K. Benndorf, and C. Biskup, "Fluorescence lifetime imaging by time-correlated single-photon counting," Microsc. Res. Technique 63, 58-66 (2004).
[CrossRef]

Nature (1)

G. J. Puppels, F. F. M. Demul, C. Otto, J. Greve, M. Robertnicoud, D. J. Arndtjovin, and T. M. Jovin, "Studying single living cells and chromosomes by Confocal Raman Microspectroscopy," Nature 347, 301-303 (1990).
[CrossRef] [PubMed]

Neurosurgery (1)

J. M. Piepmeier, C. Y. Liu, and E. Neuwelt, "Multiphoton excitation of autofluorescence for microscopy of glioma tissue - Comments," Neurosurgery 58, 767-767 (2006).

Nuc. Instr. Meth. Phys. (1)

N. Djaker, P. F. Lenne, D. Marguet, A. Colonna, C. Hadjur, and H. Rigneault, "Coherent anti-Stokes Raman scattering microscopy (CARS): Instrumentation and applications," Nuc. Instr. Meth. Phys. 571, 177-181 (2007).
[CrossRef]

Opt. Express (2)

Opt. Lett. (4)

Photochem. Photobiol. Sci. (1)

K. Suhling, P. M. W. French, and D. Phillips, "Time-resolved fluorescence microscopy," Photochem. Photobiol. Sci. 4, 13-22 (2005).
[CrossRef]

Phys. Rev. Lett. (2)

A. Zumbusch, G. R. Holtom, and X. S. Xie, "Three-dimensional vibrational imaging by coherent anti- Stokes Raman scattering," Phys. Rev. Lett. 82, 4142-4145 (1999).
[CrossRef]

A. Volkmer, J. X. Cheng, and X. S. Xie, "Vibrational imaging with high sensitivity via epidetected coherent anti-Stokes Raman scattering microscopy," Phys. Rev. Lett. 8702, 4 (2001).

PNAS (1)

C. L. Evans, E. O. Potma, M. Puoris'haag, D. Cote, C. P. Lin, and X. S. Xie, "Chemical imaging of tissue in vivo with video-rate coherent anti-Stokes Raman scattering microscopy," PNAS 102, 16807-16812 (2005).
[CrossRef] [PubMed]

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Figures (5)

Fig. 1.
Fig. 1.

(A) Schematics of our experimental CARS microscopy system. An optical parametric oscillator is synchronously pumped by a Nd:YVO4 laser (10W, 7 ps, 1064 nm, 76 MHz) to produce the excitation beam, while the Nd:YVO4 laser provides the probe beam. CARS signals are detected in epi-mode by a single photon counting avalanche photodiode (SPAD). BS: beam splitter, DM: dichroic mirror, BE: beam expander, OPO: optical parametric oscillator, TCSPC: time-correlated single photon couting, PC: personal computer. (B) TCSPC concept: the arrival time of a photon is detected with 35 ps resolution with respect to the laser pulse. (C) Photon arrival time distribution histogram of the 2845 cm-1 CARS signal from aliphatic C-H bonds generated in rat artery by scanning the CARS beams and collecting photon arrival time data for every pixel, which are fitted by a single exponential decay function. The fit values for each photon arrival time are shown in the distribution histogram. The majority of photons arrives with a time constant of 310 ps, which reflects the instrument response function of our system – limited by the timing jitter of the SPAD. The microscopic photon arrival time is determined by the TimeHarp200 histogramming electronics, whereby the start-stop times are binned to a 4096 channel histogram with an adjustable resolution per channel and a minimum resolution corresponding to ~30 ps. CARS and TPE fluorescence images are generated by the SymphoTime software by setting the respective time-gates, i.e. by selecting a subset of photons collected during a given time range within the microscopic photon arrival (TCSPC) time window, corresponding to CARS only and TPE fluorescence photons, respectively.

Fig. 2.
Fig. 2.

(A) Transmitted light micrograph of the lower stem near the roots (hypocotyls region) of a living Arabidopsis thaliana seedling. (B) Intensity CARS image of the same area of the hypocotyls shown in (A). The areas highlighted by arrows in the image are areas where emission spectra were taken and correspond to the spectra shown in (C). Image scale is 40 µm×40 µm. (C) Local emission spectra obtained by positioning the laser beams to different parts of the plant cell as indicated in (B). (D) Two-photon-excited confocal autofluorescence image of the same plant area, where just the picosecond OPO beam (817 nm) was used to excite fluorescence. (E) Same as (D), but with just the Nd:YVO4. (F) Difference image, where the fluorescence images shown on (D) and (E) were subtracted from the “CARS” image in (B). This should result in a CARS image essentially free of background fluorescence contributions except for those due to simultaneous absorption of 1 photon from the pump beam and 1 photon from the Stokes beam. All CARS images were taken at 256×256 pixels with 2 ms dwell time per pixel, resulting in a total image acquisition time of 6.93 min.

Fig. 3.
Fig. 3.

Time-correlated CARS micrographs of the hypocotyl of an Arabidopsis thaliana seedling. (A) Intensity CARS image showing all photons collected per pixel without arrival time information. Image scale is 40µm×40µm. (B) This image shows the same area as in A, but encodes the photon arrival time in false color. Note that some well-confined areas of the image indicate very short photon arrivals times (shown in dark-blue to purple color). (C) Normalized arrival time decays for different regions highlighted by the arrows in (B), as well as averaged over the entire image. The CARS images were taken at 256×256 pixels with 2 ms dwell time per pixel, resulting in a total image acquisition time of 6.93 min.

Fig. 4.
Fig. 4.

Time-gated CARS microscopy images of the same hypocotyls as shown in Figs. 2 and 3. (A) Intensity image with a time gate set to 0–0.6 ns. This short time gate separates the instantaneously arriving CARS photons from the delayed photons due to fluorescence emission. The gate width is set to cover all photons that are emitted within the time set by the instrument response function. (B) Intensity image with time gate set between 2–10 ns. This image shows only two-photon excited fluorescence. (C) Normalized arrival time decays for different regions overlaid with the schematic times gates used for isolating “early” and “late” photons resulting in the images shown in (A) and (B). The CARS images were taken at 256×256 pixels with 2 ms dwell time per pixel, resulting in a total image acquisition time of 6.93 min.

Fig. 5.
Fig. 5.

CARS micrograph of rat arterial tissue (cross-section) showing the effect of time-gating on animal tissue. The fat white outline drawn in the image indicates the border between the tissue and the beginning of the interior of the artery (the“lumen”). (A) CARS image encoding the photon arrival time on a false color scale. Image scale is 30×30 µm. (B) Normalized photon arrival time histograms obtained by isolating areas with short photon arrival times from areas with long photon arrival times as highlighted by white circles in (A). (C) Intensity image with a time gate set to 0–0.6 ns, covering the instantaneously emitted CARS photons. (D) Intensity image with time gate set to 2–10 ns, which captures all delayed photons due to fluorescence emission. Some areas of the sample shown by brightly red-colored spots lead to saturation of the detection electronics, and photon arrival times could not be properly represented for these areas. These parts were set to black in (C) and (D). All CARS images are acquired at 256×256 pixel resolution with 0.6 ms dwell time per pixel and a total image acquisition time of 2.03 min.

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