Abstract

Supercontinuum-based multiplex coherent anti-Stokes Raman scattering (CARS) microspectroscopy has been applied to vibrational imaging of a living fission yeast cell. We have successfully extracted only a vibrationally resonant CARS image from a characteristic spectral profile in the C-H stretching vibrational region. Using our simple but sensitive analysis, the vibrational contrast is significantly improved in comparison with a CARS imaging at a fixed Raman shift. The CARS image of a living yeast cell indicates several areas at which the signal is remarkably strong. They are considered to arise from mitochondria.

© 2005 Optical Society of America

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References

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  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]
  2. M. Hashimoto, T. Araki, and S. Kawata, "Molecular vibration imaging in the fingerprint region by use of coherent anti-Stokes Raman scattering microscopy with a collinear configuration," Opt. Lett. 25, 1768-1770 (2000).
    [CrossRef]
  3. J.-X. Cheng, Y. K. Jia, G. Zheng, and X. S. Xie, "Laser-scanning coherent anti-Stokes Raman scattering microscopy and applications to cell biology," Biophys. J. 83, 502-509 (2002).
    [CrossRef] [PubMed]
  4. M. Müller and J. M. Schins, "Imaging the thermodynamic state of lipid membranes with multiplex CARS microscopy," J. Phys. Chem. B 106, 3715-3723 (2002).
    [CrossRef]
  5. H. N. Paulsen, K. M. Hilligsoe, J. Thogersen, S. R. Keiding, and J. J. Larsen, "Coherent anti-Stokes Raman scattering microscopy with a photonic crystal fiber based light source," Opt. Lett. 28, 1123-1125 (2003).
    [CrossRef] [PubMed]
  6. T. Ichimura, N. Hayazawa, M. Hashimoto, Y. Inouye, and S. Kawata, "Tip-enhanced coherent anti-Stokes Raman scattering for vibrational nanoimaging," Phys. Rev. Lett. 92, 220801-220804 (2004).
    [CrossRef] [PubMed]
  7. 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]
  8. H. Kano and H. Hamaguchi, "Near-infrared coherent anti-Stokes Raman scattering microscopy using supercontinuum generated from a photonic crystal fiber," Appl. Phys. B B80, 243-246 (2005).
    [CrossRef]
  9. J.-X. Cheng, L. D. Book, and X. S. Xie, "Polarization coherent anti-Stokes Raman scattering microscopy," Opt. Lett. 26, 1341-1343 (2001).
    [CrossRef]
  10. A. Volkmer, L. D. Book, and X. S. Xie, "Time-resolved coherent anti-Stokes Raman scattering microscopy: imaging based on Raman free induction decay," Appl. Phys. Lett. 80, 1505-1507 (2002).
    [CrossRef]
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    [CrossRef]
  12. C. Otto, A. Voroshilov, S. G. Kruglik, and J. Greve, "Vibrational bands of luminescent zinc(II)-octaethylporphyrin using a polarization-sensitive "microscopic" multiplex CARS technique," J. Raman Spectrosc. 32, 495-501 (2001).
    [CrossRef]
  13. J.-X. Cheng, A. Volkmer, L. D. Book, and X. S. Xie, "Multiplex coherent anti-Stokes Raman scattering microspectroscopy and study of lipid vesicles," J. Phys. Chem. B 106, 8493-8498 (2002).
    [CrossRef]
  14. C. L. Evans, E. O. Potma, and X. S. Xie, "Coherent anti-Stokes Raman scattering spectral interferometry: determination of the real and imaginary components of nonlinear susceptibility chi(3) for vibrational microscopy," Opt. Lett. 29, 2923-2925 (2004).
    [CrossRef]
  15. T. W. Kee and M. T. Cicerone, "Simple approach to one-laser, broadband coherent anti-Stokes Raman scattering microscopy," Opt. Lett. 29, 2701-2703 (2004).
    [CrossRef] [PubMed]
  16. H. Kano and H. Hamaguchi, "Ultrabroadband (>2500 cm-1) multiplex coherent anti-Stokes Raman scattering microspectroscopy using a supercontinuum generated from a photonic crystal fiber," Appl. Phys. Lett. (accepted).
    [PubMed]
  17. J. K. Ranka, R. S. Windeler, and A. J. Stentz, "Visible continuum generation in air-silica microstructure optical fibers with anomalous dispersion at 800 nm," Opt. Lett. 25, 25-27 (2000).
    [CrossRef]
  18. M. L. Hu, C. Y. Wang, L. Chai, and A. M. Zheltikov "Frequency-tunable anti-Stokes line emission by eigenmodes of a birefringent microstructure fiber," Opt. Exp. 12, 1932-1937 (2004), http://www.opticsexpress.org/abstract.cfm?URI=OPEX-12-9-1932.
    [CrossRef]
  19. M. L. Hu, C. Y. Wang, Y. Li, Z. Wang, L. Chai, and A. M. Zheltikov "Multiplex frequency conversion of unamplified 30-fs Ti: sapphire laser pulses by an array of waveguiding wires in a random-hole microstructure fiber," Opt. Exp. 12, 6129-6134 (2004), http://www.opticsexpress.org/abstract.cfm?URI=OPEX-12-25-6129.
    [CrossRef]
  20. H. Kano and H. Hamaguchi, "Characterization of a supercontinuum generated from a photonic crystal fiber and its application to coherent Raman spectroscopy," Opt. Lett. 28, 2360-2362 (2003).
    [CrossRef] [PubMed]
  21. H. Kano and H. Hamaguchi, "Femtosecond coherent anti-Stokes Raman scattering spectroscopy using a supercontinuum generated from a photonic crystal fiber," Appl. Phys. Lett. 85, 4298-4300 (2004).
    [CrossRef]
  22. T. Nagahara, K. Imura, and H. Okamoto, "Time-resolved scanning near-field optical microscopy with supercontinuum light pulses generated in microstructure fiber," Rev. Sci. Instrum. 75, 4528-4533 (2004).
    [CrossRef]
  23. R. Holzwarth, T. Udem, T. W. Hänsch, J. C. Knight, W. J. Wadsworth, and P. S. J. Russell, "Optical frequency synthesizer for precision spectroscopy," Phys. Rev. Lett. 85, 2264-2267 (2000).
    [CrossRef] [PubMed]
  24. 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. 87, 023901-023904 (2001).
    [CrossRef]
  25. G. W. H. Wurpel, J. M. Schins, and M. Müller, "Direct measurement of chain order in single phospholipids mono- and bilayers with multiplex CARS," J. Phys. Chem. B 108, 3400-3403 (2004).
    [CrossRef]
  26. Y.-S. Huang, T. Karashima, M. Yamamoto, and H. Hamaguchi, "Molecular-level pursuit of yeast mitosis by time- and space-resolved Raman spectroscopy," J. Raman Spectrosc. 34, 1-3 (2003).
    [CrossRef]
  27. Y.-S. Huang, T. Karashima, M. Yamamoto, T. Ogura, and H. Hamaguchi, "Raman spectroscopic signature of life in a living yeast cell," J. Raman Spectrosc. 35, 525-526 (2004).
    [CrossRef]

Appl. Phys. B

H. Kano and H. Hamaguchi, "Near-infrared coherent anti-Stokes Raman scattering microscopy using supercontinuum generated from a photonic crystal fiber," Appl. Phys. B B80, 243-246 (2005).
[CrossRef]

Appl. Phys. Lett.

A. Volkmer, L. D. Book, and X. S. Xie, "Time-resolved coherent anti-Stokes Raman scattering microscopy: imaging based on Raman free induction decay," Appl. Phys. Lett. 80, 1505-1507 (2002).
[CrossRef]

H. Kano and H. Hamaguchi, "Ultrabroadband (>2500 cm-1) multiplex coherent anti-Stokes Raman scattering microspectroscopy using a supercontinuum generated from a photonic crystal fiber," Appl. Phys. Lett. (accepted).
[PubMed]

H. Kano and H. Hamaguchi, "Femtosecond coherent anti-Stokes Raman scattering spectroscopy using a supercontinuum generated from a photonic crystal fiber," Appl. Phys. Lett. 85, 4298-4300 (2004).
[CrossRef]

Biophys. J.

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

J. Phys. Chem. B

M. Müller and J. M. Schins, "Imaging the thermodynamic state of lipid membranes with multiplex CARS microscopy," J. Phys. Chem. B 106, 3715-3723 (2002).
[CrossRef]

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]

G. W. H. Wurpel, J. M. Schins, and M. Müller, "Direct measurement of chain order in single phospholipids mono- and bilayers with multiplex CARS," J. Phys. Chem. B 108, 3400-3403 (2004).
[CrossRef]

J.-X. Cheng, A. Volkmer, L. D. Book, and X. S. Xie, "Multiplex coherent anti-Stokes Raman scattering microspectroscopy and study of lipid vesicles," J. Phys. Chem. B 106, 8493-8498 (2002).
[CrossRef]

J. Raman Spectrosc.

Y.-S. Huang, T. Karashima, M. Yamamoto, and H. Hamaguchi, "Molecular-level pursuit of yeast mitosis by time- and space-resolved Raman spectroscopy," J. Raman Spectrosc. 34, 1-3 (2003).
[CrossRef]

Y.-S. Huang, T. Karashima, M. Yamamoto, T. Ogura, and H. Hamaguchi, "Raman spectroscopic signature of life in a living yeast cell," J. Raman Spectrosc. 35, 525-526 (2004).
[CrossRef]

C. Otto, A. Voroshilov, S. G. Kruglik, and J. Greve, "Vibrational bands of luminescent zinc(II)-octaethylporphyrin using a polarization-sensitive "microscopic" multiplex CARS technique," J. Raman Spectrosc. 32, 495-501 (2001).
[CrossRef]

Opt. Exp.

M. L. Hu, C. Y. Wang, L. Chai, and A. M. Zheltikov "Frequency-tunable anti-Stokes line emission by eigenmodes of a birefringent microstructure fiber," Opt. Exp. 12, 1932-1937 (2004), http://www.opticsexpress.org/abstract.cfm?URI=OPEX-12-9-1932.
[CrossRef]

M. L. Hu, C. Y. Wang, Y. Li, Z. Wang, L. Chai, and A. M. Zheltikov "Multiplex frequency conversion of unamplified 30-fs Ti: sapphire laser pulses by an array of waveguiding wires in a random-hole microstructure fiber," Opt. Exp. 12, 6129-6134 (2004), http://www.opticsexpress.org/abstract.cfm?URI=OPEX-12-25-6129.
[CrossRef]

Opt. Lett.

J. K. Ranka, R. S. Windeler, and A. J. Stentz, "Visible continuum generation in air-silica microstructure optical fibers with anomalous dispersion at 800 nm," Opt. Lett. 25, 25-27 (2000).
[CrossRef]

M. Hashimoto, T. Araki, and S. Kawata, "Molecular vibration imaging in the fingerprint region by use of coherent anti-Stokes Raman scattering microscopy with a collinear configuration," Opt. Lett. 25, 1768-1770 (2000).
[CrossRef]

J.-X. Cheng, L. D. Book, and X. S. Xie, "Polarization coherent anti-Stokes Raman scattering microscopy," Opt. Lett. 26, 1341-1343 (2001).
[CrossRef]

G. W. H. Wurpel, J. M. Schins, and M. Müller, "Chemical specificity in three-dimensional imaging with multiplex coherent anti-Stokes Raman scattering microscopy," Opt. Lett. 27, 1093-1095 (2002).
[CrossRef]

H. N. Paulsen, K. M. Hilligsoe, J. Thogersen, S. R. Keiding, and J. J. Larsen, "Coherent anti-Stokes Raman scattering microscopy with a photonic crystal fiber based light source," Opt. Lett. 28, 1123-1125 (2003).
[CrossRef] [PubMed]

H. Kano and H. Hamaguchi, "Characterization of a supercontinuum generated from a photonic crystal fiber and its application to coherent Raman spectroscopy," Opt. Lett. 28, 2360-2362 (2003).
[CrossRef] [PubMed]

T. W. Kee and M. T. Cicerone, "Simple approach to one-laser, broadband coherent anti-Stokes Raman scattering microscopy," Opt. Lett. 29, 2701-2703 (2004).
[CrossRef] [PubMed]

C. L. Evans, E. O. Potma, and X. S. Xie, "Coherent anti-Stokes Raman scattering spectral interferometry: determination of the real and imaginary components of nonlinear susceptibility chi(3) for vibrational microscopy," Opt. Lett. 29, 2923-2925 (2004).
[CrossRef]

Phys. Rev. Lett.

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]

T. Ichimura, N. Hayazawa, M. Hashimoto, Y. Inouye, and S. Kawata, "Tip-enhanced coherent anti-Stokes Raman scattering for vibrational nanoimaging," Phys. Rev. Lett. 92, 220801-220804 (2004).
[CrossRef] [PubMed]

R. Holzwarth, T. Udem, T. W. Hänsch, J. C. Knight, W. J. Wadsworth, and P. S. J. Russell, "Optical frequency synthesizer for precision spectroscopy," Phys. Rev. Lett. 85, 2264-2267 (2000).
[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. 87, 023901-023904 (2001).
[CrossRef]

Rev. Sci. Instrum.

T. Nagahara, K. Imura, and H. Okamoto, "Time-resolved scanning near-field optical microscopy with supercontinuum light pulses generated in microstructure fiber," Rev. Sci. Instrum. 75, 4528-4533 (2004).
[CrossRef]

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Figures (3)

Fig. 1.
Fig. 1.

Typical spectral profile of the CARS signal of a living yeast cell (red solid) and surrounding water (blue dotted)

Fig. 2.
Fig. 2.

CARS lateral images of living yeast cells at the Raman shift of 2856 cm-1 (a) and 2200 cm-1 (b). The black and white crosses in (a) correspond to the positions at which the spectral profiles are shown in Fig. 1.

Fig. 3.
Fig. 3.

Vibrationally resonant CARS imaging using the differentiation method (a) and a method using a conventional lineshape function described in Eq. (1) (b). Note that the contrast and signal-to-noise ratio is dramatically improved in comparison with Fig. 2(a).

Equations (1)

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I ( ω ) = A NR e i ϕ + R A R Γ R Γ R i ( ω Ω R ) 2 .

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