Abstract

We present a systematic characterization of coherent anti-Stokes Raman scattering (CARS) microscopy. CARS signal generation in a heterogeneous sample under a tight-focusing condition is formulated by the Green’s function method. The CARS radiation pattern and the forward- and backward-detected CARS signals from a three-dimensional Raman scatterer are calculated. The coherent nature of CARS image formation and its consequences for image contrast and spatial resolution are investigated. Experimental implementations of CARS microscopy with collinearly copropagating and counterpropagating excitation beams, forward and backward data collection, and polarization-sensitive detection are described. Finally, CARS images of unstained live cells with forward detection, epidetection, and polarization-sensitive detection are presented and compared.

© 2002 Optical Society of America

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    [CrossRef]
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    [CrossRef]
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    [CrossRef]
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    [CrossRef]
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    [CrossRef]
  58. M. A. Yuratich and D. C. Hanna, “Coherent anti-Stokes Raman spectroscopy (CARS) selection rules, depolarization ratios and rotational structure,” Mol. Phys. 33, 671–682 (1977).
    [CrossRef]
  59. As our calculation did not consider the contribution from the solvent surrounding the scatterer, the forward radiation pattern is a suitable description of polarization CARS for which the nonresonant background from the solvent is suppressed. The forward CARS generated by a pair of parallel-polarized pump and Stokes beams is a coherent addition of the signal from the scatterer and that from the solvent and is always highly directional.
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2001

E. O. Potma, W. P. de Boeij, P. J. M. van Haastert, and D. A. Wiersma, “Real-time visualisation of intracelluar hydrodynamics in single living cells,” Proc. Natl. Acad. Sci. U.S.A. 98, 1577–1582 (2001).
[CrossRef]

J. X. Cheng, A. Volkmer, L. D. Book, and X. S. Xie, “An epi-detected coherent anti-Stokes Raman scattering (E-CARS) microscope with high spectral resolution and high sensitivity,” J. Phys. Chem. B 105, 1277–1280 (2001).
[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. 87, 0239011–0239014 (2001).
[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]

2000

1999

P. J. Campagnola, M.-D. Wei, A. Lewis, and L. M. Loew, “High-resolution nonlinear optical imaging of live cells by second harmonic generation,” Biophys. J. 77, 3341–3349 (1999).
[CrossRef] [PubMed]

M. Florsheimer, C. Brillert, and H. Fuchs, “Chemical imaging of interfaces by sum frequency microscopy,” Langmuir 15, 5437–5439 (1999).
[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]

1998

1997

Y. Barad, H. Eisenberg, M. Horowitz, and Y. Silberberg, “Nonlinear scanning laser microscopy by third-harmonic generation,” Appl. Phys. Lett. 70, 922–924 (1997).
[CrossRef]

L. Novotny, “Allowed and forbidden light in near-field optics. II. Interacting dipolar particles,” J. Opt. Soc. Am. A 14, 105–113 (1997).
[CrossRef]

1996

C. Xu, W. Zipfel, J. B. Shear, R. M. Williams, and W. W. Webb, “Multiphoton fluorescence excitation: new spectral windows for biological nonlinear microscopy,” Proc. Natl. Acad. Sci. U.S.A. 93, 10,763–10,768 (1996).
[CrossRef]

1993

S. Hell, G. Reiner, C. Cremer, and E. H. K. Stelzer, “Aberrations in confocal fluorescence microscopy induced by mismatches in refractive index,” J. Microsc. (Oxford) 169, 391–405 (1993).
[CrossRef]

1991

W. Kaabar and R. Devonshire, “A versatile model of CARS signal generation: optimum beam diameter ratios for different phase-matching geometries,” Chem. Phys. Lett. 186, 522–530 (1991).
[CrossRef]

1990

W. Denk, J. H. Strickler, and W. W. Webb, “Two-photon laser scanning fluorescence microscopy,” Science 248, 73–76 (1990).
[CrossRef] [PubMed]

1986

1985

S. Velsko and R. M. Hochstrasser, “Studies of vibrational relaxation in low-temperature molecular crystal using coherent Raman spectroscopy,” J. Phys. Chem. 89, 2240–2253 (1985).
[CrossRef]

1984

1982

1981

1980

1979

J.-L. Oudar, R. W. Smith, and Y. R. Shen, “Polarization-sensitive coherent anti-Stokes Raman spectroscopy,” Appl. Phys. Lett. 34, 758–760 (1979).
[CrossRef]

1977

S. A. Akhmanov, A. F. Bunkin, S. G. Ivanov, and N. I. Koroteev, “Coherent ellipsometry of Raman scattering of light,” JETP 25, 416–420 (1977).

M. A. Yuratich and D. C. Hanna, “Coherent anti-Stokes Raman spectroscopy (CARS) selection rules, depolarization ratios and rotational structure,” Mol. Phys. 33, 671–682 (1977).
[CrossRef]

W. M. Shaub, A. B. Harvey, and G. C. Bjorklund, “Power generation in coherent anti-Stokes Raman spectroscopy with focused laser beams,” J. Chem. Phys. 67, 2547–2550 (1977).
[CrossRef]

1976

H. Lotem, R. T. Lynch, Jr., and N. Bloembergen, “Interference between Raman resonances in four-wave difference mixing,” Phys. Rev. A 14, 1748–1755 (1976).
[CrossRef]

R. T. Lynch, Jr., S. D. Kramer, H. Lotem, and N. Bloembergen, “Double resonance interference in third-order light mixing,” Opt. Commun. 16, 372–375 (1976).
[CrossRef]

J. J. Song, G. L. Eesley, and M. D. Levenson, “Background suppression in coherent Raman spectroscopy,” Appl. Phys. Lett. 29, 567–569 (1976).
[CrossRef]

1975

G. C. Bjorklund, “Effects of focusing on third-order nonlinear processes in isotropic media,” IEEE J. Quantum Electron. QE-11, 287–296 (1975).
[CrossRef]

1965

P. D. Maker and R. W. Terhune, “Study of optical effects due to an induced polarization third order in the electric field strength,” Phys. Rev. 137, A801–A818 (1965).
[CrossRef]

1962

D. A. Kleinman, “Nonlinear dielectric polarization in optical media,” Phys. Rev. 126, 1977–1979 (1962).
[CrossRef]

1959

B. Richards and E. Wolf, “Electromagnetic diffraction in optical systems. II. Structure of the image field in an aplanatic system,” Proc. R. Soc. London, Ser. A 253, 358–379 (1959).
[CrossRef]

Akhmanov, S. A.

S. A. Akhmanov, A. F. Bunkin, S. G. Ivanov, and N. I. Koroteev, “Coherent ellipsometry of Raman scattering of light,” JETP 25, 416–420 (1977).

Araki, T.

Barad, Y.

Y. Barad, H. Eisenberg, M. Horowitz, and Y. Silberberg, “Nonlinear scanning laser microscopy by third-harmonic generation,” Appl. Phys. Lett. 70, 922–924 (1997).
[CrossRef]

Bjorklund, G. C.

W. M. Shaub, A. B. Harvey, and G. C. Bjorklund, “Power generation in coherent anti-Stokes Raman spectroscopy with focused laser beams,” J. Chem. Phys. 67, 2547–2550 (1977).
[CrossRef]

G. C. Bjorklund, “Effects of focusing on third-order nonlinear processes in isotropic media,” IEEE J. Quantum Electron. QE-11, 287–296 (1975).
[CrossRef]

Bloembergen, N.

H. Lotem, R. T. Lynch, Jr., and N. Bloembergen, “Interference between Raman resonances in four-wave difference mixing,” Phys. Rev. A 14, 1748–1755 (1976).
[CrossRef]

R. T. Lynch, Jr., S. D. Kramer, H. Lotem, and N. Bloembergen, “Double resonance interference in third-order light mixing,” Opt. Commun. 16, 372–375 (1976).
[CrossRef]

Book, L. D.

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

J. X. Cheng, A. Volkmer, L. D. Book, and X. S. Xie, “An epi-detected coherent anti-Stokes Raman scattering (E-CARS) microscope with high spectral resolution and high sensitivity,” J. Phys. Chem. B 105, 1277–1280 (2001).
[CrossRef]

Brakenhoff, G. J.

M. Müller, J. Squier, C. A. de Lange, and G. J. Brakenhoff, “CARS microscopy with folded BOXCARS phasematching,” J. Microsc. (Oxford) 197, 150–158 (2000).
[CrossRef]

M. Müller, J. Squier, K. R. Wilson, and G. J. Brakenhoff, “3D-microscopy of transparent objects using third-harmonic generation,” J. Microsc. (Oxford) 191, 266–274 (1998).
[CrossRef]

Brillert, C.

M. Florsheimer, C. Brillert, and H. Fuchs, “Chemical imaging of interfaces by sum frequency microscopy,” Langmuir 15, 5437–5439 (1999).
[CrossRef]

Bunkin, A. F.

S. A. Akhmanov, A. F. Bunkin, S. G. Ivanov, and N. I. Koroteev, “Coherent ellipsometry of Raman scattering of light,” JETP 25, 416–420 (1977).

Campagnola, P. J.

P. J. Campagnola, M.-D. Wei, A. Lewis, and L. M. Loew, “High-resolution nonlinear optical imaging of live cells by second harmonic generation,” Biophys. J. 77, 3341–3349 (1999).
[CrossRef] [PubMed]

Cheng, J. X.

J. X. Cheng, A. Volkmer, L. D. Book, and X. S. Xie, “An epi-detected coherent anti-Stokes Raman scattering (E-CARS) microscope with high spectral resolution and high sensitivity,” J. Phys. Chem. B 105, 1277–1280 (2001).
[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]

Cheng, J.-X.

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, 0239011–0239014 (2001).
[CrossRef]

Cremer, C.

S. Hell, G. Reiner, C. Cremer, and E. H. K. Stelzer, “Aberrations in confocal fluorescence microscopy induced by mismatches in refractive index,” J. Microsc. (Oxford) 169, 391–405 (1993).
[CrossRef]

Davis, L. C.

de Boeij, W. P.

E. O. Potma, W. P. de Boeij, P. J. M. van Haastert, and D. A. Wiersma, “Real-time visualisation of intracelluar hydrodynamics in single living cells,” Proc. Natl. Acad. Sci. U.S.A. 98, 1577–1582 (2001).
[CrossRef]

E. O. Potma, W. P. de Boeij, and D. A. Wiersma, “Nonlinear coherent four-wave mixing in optical microscopy,” J. Opt. Soc. Am. B 17, 1678–1684 (2000).
[CrossRef]

de Grauw, C. J.

de Lange, C. A.

M. Müller, J. Squier, C. A. de Lange, and G. J. Brakenhoff, “CARS microscopy with folded BOXCARS phasematching,” J. Microsc. (Oxford) 197, 150–158 (2000).
[CrossRef]

Denk, W.

W. Denk, J. H. Strickler, and W. W. Webb, “Two-photon laser scanning fluorescence microscopy,” Science 248, 73–76 (1990).
[CrossRef] [PubMed]

Devonshire, R.

W. Kaabar and R. Devonshire, “A versatile model of CARS signal generation: optimum beam diameter ratios for different phase-matching geometries,” Chem. Phys. Lett. 186, 522–530 (1991).
[CrossRef]

Duncan, M. D.

Eckbreth, A. C.

Eesley, G. L.

J. J. Song, G. L. Eesley, and M. D. Levenson, “Background suppression in coherent Raman spectroscopy,” Appl. Phys. Lett. 29, 567–569 (1976).
[CrossRef]

Eisenberg, H.

Y. Barad, H. Eisenberg, M. Horowitz, and Y. Silberberg, “Nonlinear scanning laser microscopy by third-harmonic generation,” Appl. Phys. Lett. 70, 922–924 (1997).
[CrossRef]

Falk, J.

S. G. Guha and J. Falk, “The effects of focusing on the efficiency of coherent anti-Stokes Raman scattering,” J. Chem. Phys. 75, 2599–2562 (1981).
[CrossRef]

Florsheimer, M.

M. Florsheimer, C. Brillert, and H. Fuchs, “Chemical imaging of interfaces by sum frequency microscopy,” Langmuir 15, 5437–5439 (1999).
[CrossRef]

Fuchs, H.

M. Florsheimer, C. Brillert, and H. Fuchs, “Chemical imaging of interfaces by sum frequency microscopy,” Langmuir 15, 5437–5439 (1999).
[CrossRef]

Gauderon, R.

Greve, J.

Guha, S. G.

S. G. Guha and J. Falk, “The effects of focusing on the efficiency of coherent anti-Stokes Raman scattering,” J. Chem. Phys. 75, 2599–2562 (1981).
[CrossRef]

Hall, R. J.

Hanna, D. C.

M. A. Yuratich and D. C. Hanna, “Coherent anti-Stokes Raman spectroscopy (CARS) selection rules, depolarization ratios and rotational structure,” Mol. Phys. 33, 671–682 (1977).
[CrossRef]

Harvey, A. B.

W. M. Shaub, A. B. Harvey, and G. C. Bjorklund, “Power generation in coherent anti-Stokes Raman spectroscopy with focused laser beams,” J. Chem. Phys. 67, 2547–2550 (1977).
[CrossRef]

Hashimoto, M.

Hell, S.

S. Hell, G. Reiner, C. Cremer, and E. H. K. Stelzer, “Aberrations in confocal fluorescence microscopy induced by mismatches in refractive index,” J. Microsc. (Oxford) 169, 391–405 (1993).
[CrossRef]

Hell, S. W.

M. Straub, P. Lodemann, P. Holroyd, R. Jahn, and S. W. Hell, “Live cell imaging by multifocal multiphoton microscopy,” Eur. J. Cell. Biol. 79, 726–734 (2000).
[CrossRef] [PubMed]

Hochstrasser, R. M.

S. Velsko and R. M. Hochstrasser, “Studies of vibrational relaxation in low-temperature molecular crystal using coherent Raman spectroscopy,” J. Phys. Chem. 89, 2240–2253 (1985).
[CrossRef]

Holroyd, P.

M. Straub, P. Lodemann, P. Holroyd, R. Jahn, and S. W. Hell, “Live cell imaging by multifocal multiphoton microscopy,” Eur. J. Cell. Biol. 79, 726–734 (2000).
[CrossRef] [PubMed]

Holtom, G. R.

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]

Horowitz, M.

Y. Barad, H. Eisenberg, M. Horowitz, and Y. Silberberg, “Nonlinear scanning laser microscopy by third-harmonic generation,” Appl. Phys. Lett. 70, 922–924 (1997).
[CrossRef]

Ivanov, S. G.

S. A. Akhmanov, A. F. Bunkin, S. G. Ivanov, and N. I. Koroteev, “Coherent ellipsometry of Raman scattering of light,” JETP 25, 416–420 (1977).

Jahn, R.

M. Straub, P. Lodemann, P. Holroyd, R. Jahn, and S. W. Hell, “Live cell imaging by multifocal multiphoton microscopy,” Eur. J. Cell. Biol. 79, 726–734 (2000).
[CrossRef] [PubMed]

Kaabar, W.

W. Kaabar and R. Devonshire, “A versatile model of CARS signal generation: optimum beam diameter ratios for different phase-matching geometries,” Chem. Phys. Lett. 186, 522–530 (1991).
[CrossRef]

Kamga, F. M.

Kawata, S.

Kleinman, D. A.

D. A. Kleinman, “Nonlinear dielectric polarization in optical media,” Phys. Rev. 126, 1977–1979 (1962).
[CrossRef]

Koroteev, N. I.

S. A. Akhmanov, A. F. Bunkin, S. G. Ivanov, and N. I. Koroteev, “Coherent ellipsometry of Raman scattering of light,” JETP 25, 416–420 (1977).

Kramer, S. D.

R. T. Lynch, Jr., S. D. Kramer, H. Lotem, and N. Bloembergen, “Double resonance interference in third-order light mixing,” Opt. Commun. 16, 372–375 (1976).
[CrossRef]

Lee, S. W.

Levenson, M. D.

J. J. Song, G. L. Eesley, and M. D. Levenson, “Background suppression in coherent Raman spectroscopy,” Appl. Phys. Lett. 29, 567–569 (1976).
[CrossRef]

Lewis, A.

P. J. Campagnola, M.-D. Wei, A. Lewis, and L. M. Loew, “High-resolution nonlinear optical imaging of live cells by second harmonic generation,” Biophys. J. 77, 3341–3349 (1999).
[CrossRef] [PubMed]

Ling, H.

Lodemann, P.

M. Straub, P. Lodemann, P. Holroyd, R. Jahn, and S. W. Hell, “Live cell imaging by multifocal multiphoton microscopy,” Eur. J. Cell. Biol. 79, 726–734 (2000).
[CrossRef] [PubMed]

Loew, L. M.

P. J. Campagnola, M.-D. Wei, A. Lewis, and L. M. Loew, “High-resolution nonlinear optical imaging of live cells by second harmonic generation,” Biophys. J. 77, 3341–3349 (1999).
[CrossRef] [PubMed]

Lotem, H.

R. T. Lynch, Jr., S. D. Kramer, H. Lotem, and N. Bloembergen, “Double resonance interference in third-order light mixing,” Opt. Commun. 16, 372–375 (1976).
[CrossRef]

H. Lotem, R. T. Lynch, Jr., and N. Bloembergen, “Interference between Raman resonances in four-wave difference mixing,” Phys. Rev. A 14, 1748–1755 (1976).
[CrossRef]

Lukins, P. B.

Lynch Jr., R. T.

R. T. Lynch, Jr., S. D. Kramer, H. Lotem, and N. Bloembergen, “Double resonance interference in third-order light mixing,” Opt. Commun. 16, 372–375 (1976).
[CrossRef]

H. Lotem, R. T. Lynch, Jr., and N. Bloembergen, “Interference between Raman resonances in four-wave difference mixing,” Phys. Rev. A 14, 1748–1755 (1976).
[CrossRef]

Maker, P. D.

P. D. Maker and R. W. Terhune, “Study of optical effects due to an induced polarization third order in the electric field strength,” Phys. Rev. 137, A801–A818 (1965).
[CrossRef]

Manuccia, T. J.

Marko, K. A.

Mertz, J.

Moreaux, L.

Müller, M.

M. Müller, J. Squier, C. A. de Lange, and G. J. Brakenhoff, “CARS microscopy with folded BOXCARS phasematching,” J. Microsc. (Oxford) 197, 150–158 (2000).
[CrossRef]

M. Müller, J. Squier, K. R. Wilson, and G. J. Brakenhoff, “3D-microscopy of transparent objects using third-harmonic generation,” J. Microsc. (Oxford) 191, 266–274 (1998).
[CrossRef]

Novotny, L.

Otto, C.

Oudar, J.-L.

J.-L. Oudar, R. W. Smith, and Y. R. Shen, “Polarization-sensitive coherent anti-Stokes Raman spectroscopy,” Appl. Phys. Lett. 34, 758–760 (1979).
[CrossRef]

Potma, E. O.

E. O. Potma, W. P. de Boeij, P. J. M. van Haastert, and D. A. Wiersma, “Real-time visualisation of intracelluar hydrodynamics in single living cells,” Proc. Natl. Acad. Sci. U.S.A. 98, 1577–1582 (2001).
[CrossRef]

E. O. Potma, W. P. de Boeij, and D. A. Wiersma, “Nonlinear coherent four-wave mixing in optical microscopy,” J. Opt. Soc. Am. B 17, 1678–1684 (2000).
[CrossRef]

Reiner, G.

S. Hell, G. Reiner, C. Cremer, and E. H. K. Stelzer, “Aberrations in confocal fluorescence microscopy induced by mismatches in refractive index,” J. Microsc. (Oxford) 169, 391–405 (1993).
[CrossRef]

Reintjes, J.

Richards, B.

B. Richards and E. Wolf, “Electromagnetic diffraction in optical systems. II. Structure of the image field in an aplanatic system,” Proc. R. Soc. London, Ser. A 253, 358–379 (1959).
[CrossRef]

Rimai, L.

Sandre, O.

Sceats, M. G.

Shaub, W. M.

W. M. Shaub, A. B. Harvey, and G. C. Bjorklund, “Power generation in coherent anti-Stokes Raman spectroscopy with focused laser beams,” J. Chem. Phys. 67, 2547–2550 (1977).
[CrossRef]

Shear, J. B.

C. Xu, W. Zipfel, J. B. Shear, R. M. Williams, and W. W. Webb, “Multiphoton fluorescence excitation: new spectral windows for biological nonlinear microscopy,” Proc. Natl. Acad. Sci. U.S.A. 93, 10,763–10,768 (1996).
[CrossRef]

Shen, Y. R.

J.-L. Oudar, R. W. Smith, and Y. R. Shen, “Polarization-sensitive coherent anti-Stokes Raman spectroscopy,” Appl. Phys. Lett. 34, 758–760 (1979).
[CrossRef]

Sheppard, C. J. R.

Shirley, J. A.

Sijtsema, N. M.

Silberberg, Y.

Y. Barad, H. Eisenberg, M. Horowitz, and Y. Silberberg, “Nonlinear scanning laser microscopy by third-harmonic generation,” Appl. Phys. Lett. 70, 922–924 (1997).
[CrossRef]

Smith, R. W.

J.-L. Oudar, R. W. Smith, and Y. R. Shen, “Polarization-sensitive coherent anti-Stokes Raman spectroscopy,” Appl. Phys. Lett. 34, 758–760 (1979).
[CrossRef]

Song, J. J.

J. J. Song, G. L. Eesley, and M. D. Levenson, “Background suppression in coherent Raman spectroscopy,” Appl. Phys. Lett. 29, 567–569 (1976).
[CrossRef]

Squier, J.

M. Müller, J. Squier, C. A. de Lange, and G. J. Brakenhoff, “CARS microscopy with folded BOXCARS phasematching,” J. Microsc. (Oxford) 197, 150–158 (2000).
[CrossRef]

M. Müller, J. Squier, K. R. Wilson, and G. J. Brakenhoff, “3D-microscopy of transparent objects using third-harmonic generation,” J. Microsc. (Oxford) 191, 266–274 (1998).
[CrossRef]

Stelzer, E. H. K.

S. Hell, G. Reiner, C. Cremer, and E. H. K. Stelzer, “Aberrations in confocal fluorescence microscopy induced by mismatches in refractive index,” J. Microsc. (Oxford) 169, 391–405 (1993).
[CrossRef]

Straub, M.

M. Straub, P. Lodemann, P. Holroyd, R. Jahn, and S. W. Hell, “Live cell imaging by multifocal multiphoton microscopy,” Eur. J. Cell. Biol. 79, 726–734 (2000).
[CrossRef] [PubMed]

Strickler, J. H.

W. Denk, J. H. Strickler, and W. W. Webb, “Two-photon laser scanning fluorescence microscopy,” Science 248, 73–76 (1990).
[CrossRef] [PubMed]

Teets, R. E.

Terhune, R. W.

P. D. Maker and R. W. Terhune, “Study of optical effects due to an induced polarization third order in the electric field strength,” Phys. Rev. 137, A801–A818 (1965).
[CrossRef]

van Haastert, P. J. M.

E. O. Potma, W. P. de Boeij, P. J. M. van Haastert, and D. A. Wiersma, “Real-time visualisation of intracelluar hydrodynamics in single living cells,” Proc. Natl. Acad. Sci. U.S.A. 98, 1577–1582 (2001).
[CrossRef]

Velsko, S.

S. Velsko and R. M. Hochstrasser, “Studies of vibrational relaxation in low-temperature molecular crystal using coherent Raman spectroscopy,” J. Phys. Chem. 89, 2240–2253 (1985).
[CrossRef]

Volkmer, A.

J. X. Cheng, A. Volkmer, L. D. Book, and X. S. Xie, “An epi-detected coherent anti-Stokes Raman scattering (E-CARS) microscope with high spectral resolution and high sensitivity,” J. Phys. Chem. B 105, 1277–1280 (2001).
[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. 87, 0239011–0239014 (2001).
[CrossRef]

Webb, W. W.

C. Xu, W. Zipfel, J. B. Shear, R. M. Williams, and W. W. Webb, “Multiphoton fluorescence excitation: new spectral windows for biological nonlinear microscopy,” Proc. Natl. Acad. Sci. U.S.A. 93, 10,763–10,768 (1996).
[CrossRef]

W. Denk, J. H. Strickler, and W. W. Webb, “Two-photon laser scanning fluorescence microscopy,” Science 248, 73–76 (1990).
[CrossRef] [PubMed]

Wei, M.-D.

P. J. Campagnola, M.-D. Wei, A. Lewis, and L. M. Loew, “High-resolution nonlinear optical imaging of live cells by second harmonic generation,” Biophys. J. 77, 3341–3349 (1999).
[CrossRef] [PubMed]

Wiersma, D. A.

E. O. Potma, W. P. de Boeij, P. J. M. van Haastert, and D. A. Wiersma, “Real-time visualisation of intracelluar hydrodynamics in single living cells,” Proc. Natl. Acad. Sci. U.S.A. 98, 1577–1582 (2001).
[CrossRef]

E. O. Potma, W. P. de Boeij, and D. A. Wiersma, “Nonlinear coherent four-wave mixing in optical microscopy,” J. Opt. Soc. Am. B 17, 1678–1684 (2000).
[CrossRef]

Williams, R. M.

C. Xu, W. Zipfel, J. B. Shear, R. M. Williams, and W. W. Webb, “Multiphoton fluorescence excitation: new spectral windows for biological nonlinear microscopy,” Proc. Natl. Acad. Sci. U.S.A. 93, 10,763–10,768 (1996).
[CrossRef]

Wilson, K. R.

M. Müller, J. Squier, K. R. Wilson, and G. J. Brakenhoff, “3D-microscopy of transparent objects using third-harmonic generation,” J. Microsc. (Oxford) 191, 266–274 (1998).
[CrossRef]

Wolf, E.

B. Richards and E. Wolf, “Electromagnetic diffraction in optical systems. II. Structure of the image field in an aplanatic system,” Proc. R. Soc. London, Ser. A 253, 358–379 (1959).
[CrossRef]

Wouters, S. D.

Xie, X. S.

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

J. X. Cheng, A. Volkmer, L. D. Book, and X. S. Xie, “An epi-detected coherent anti-Stokes Raman scattering (E-CARS) microscope with high spectral resolution and high sensitivity,” J. Phys. Chem. B 105, 1277–1280 (2001).
[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. 87, 0239011–0239014 (2001).
[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]

Xu, C.

C. Xu, W. Zipfel, J. B. Shear, R. M. Williams, and W. W. Webb, “Multiphoton fluorescence excitation: new spectral windows for biological nonlinear microscopy,” Proc. Natl. Acad. Sci. U.S.A. 93, 10,763–10,768 (1996).
[CrossRef]

Yuratich, M. A.

M. A. Yuratich and D. C. Hanna, “Coherent anti-Stokes Raman spectroscopy (CARS) selection rules, depolarization ratios and rotational structure,” Mol. Phys. 33, 671–682 (1977).
[CrossRef]

Zipfel, W.

C. Xu, W. Zipfel, J. B. Shear, R. M. Williams, and W. W. Webb, “Multiphoton fluorescence excitation: new spectral windows for biological nonlinear microscopy,” Proc. Natl. Acad. Sci. U.S.A. 93, 10,763–10,768 (1996).
[CrossRef]

Zumbusch, A.

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]

Appl. Opt.

Appl. Phys. Lett.

Y. Barad, H. Eisenberg, M. Horowitz, and Y. Silberberg, “Nonlinear scanning laser microscopy by third-harmonic generation,” Appl. Phys. Lett. 70, 922–924 (1997).
[CrossRef]

J. J. Song, G. L. Eesley, and M. D. Levenson, “Background suppression in coherent Raman spectroscopy,” Appl. Phys. Lett. 29, 567–569 (1976).
[CrossRef]

J.-L. Oudar, R. W. Smith, and Y. R. Shen, “Polarization-sensitive coherent anti-Stokes Raman spectroscopy,” Appl. Phys. Lett. 34, 758–760 (1979).
[CrossRef]

Appl. Spectrosc.

Biophys. J.

P. J. Campagnola, M.-D. Wei, A. Lewis, and L. M. Loew, “High-resolution nonlinear optical imaging of live cells by second harmonic generation,” Biophys. J. 77, 3341–3349 (1999).
[CrossRef] [PubMed]

Chem. Phys. Lett.

W. Kaabar and R. Devonshire, “A versatile model of CARS signal generation: optimum beam diameter ratios for different phase-matching geometries,” Chem. Phys. Lett. 186, 522–530 (1991).
[CrossRef]

Eur. J. Cell. Biol.

M. Straub, P. Lodemann, P. Holroyd, R. Jahn, and S. W. Hell, “Live cell imaging by multifocal multiphoton microscopy,” Eur. J. Cell. Biol. 79, 726–734 (2000).
[CrossRef] [PubMed]

IEEE J. Quantum Electron.

G. C. Bjorklund, “Effects of focusing on third-order nonlinear processes in isotropic media,” IEEE J. Quantum Electron. QE-11, 287–296 (1975).
[CrossRef]

J. Chem. Phys.

W. M. Shaub, A. B. Harvey, and G. C. Bjorklund, “Power generation in coherent anti-Stokes Raman spectroscopy with focused laser beams,” J. Chem. Phys. 67, 2547–2550 (1977).
[CrossRef]

S. G. Guha and J. Falk, “The effects of focusing on the efficiency of coherent anti-Stokes Raman scattering,” J. Chem. Phys. 75, 2599–2562 (1981).
[CrossRef]

J. Microsc. (Oxford)

M. Müller, J. Squier, C. A. de Lange, and G. J. Brakenhoff, “CARS microscopy with folded BOXCARS phasematching,” J. Microsc. (Oxford) 197, 150–158 (2000).
[CrossRef]

M. Müller, J. Squier, K. R. Wilson, and G. J. Brakenhoff, “3D-microscopy of transparent objects using third-harmonic generation,” J. Microsc. (Oxford) 191, 266–274 (1998).
[CrossRef]

S. Hell, G. Reiner, C. Cremer, and E. H. K. Stelzer, “Aberrations in confocal fluorescence microscopy induced by mismatches in refractive index,” J. Microsc. (Oxford) 169, 391–405 (1993).
[CrossRef]

J. Opt. Soc. Am. A

J. Opt. Soc. Am. B

J. Phys. Chem.

S. Velsko and R. M. Hochstrasser, “Studies of vibrational relaxation in low-temperature molecular crystal using coherent Raman spectroscopy,” J. Phys. Chem. 89, 2240–2253 (1985).
[CrossRef]

J. Phys. Chem. B

J. X. Cheng, A. Volkmer, L. D. Book, and X. S. Xie, “An epi-detected coherent anti-Stokes Raman scattering (E-CARS) microscope with high spectral resolution and high sensitivity,” J. Phys. Chem. B 105, 1277–1280 (2001).
[CrossRef]

JETP

S. A. Akhmanov, A. F. Bunkin, S. G. Ivanov, and N. I. Koroteev, “Coherent ellipsometry of Raman scattering of light,” JETP 25, 416–420 (1977).

Langmuir

M. Florsheimer, C. Brillert, and H. Fuchs, “Chemical imaging of interfaces by sum frequency microscopy,” Langmuir 15, 5437–5439 (1999).
[CrossRef]

Mol. Phys.

M. A. Yuratich and D. C. Hanna, “Coherent anti-Stokes Raman spectroscopy (CARS) selection rules, depolarization ratios and rotational structure,” Mol. Phys. 33, 671–682 (1977).
[CrossRef]

Opt. Commun.

R. T. Lynch, Jr., S. D. Kramer, H. Lotem, and N. Bloembergen, “Double resonance interference in third-order light mixing,” Opt. Commun. 16, 372–375 (1976).
[CrossRef]

Opt. Lett.

Phys. Rev.

D. A. Kleinman, “Nonlinear dielectric polarization in optical media,” Phys. Rev. 126, 1977–1979 (1962).
[CrossRef]

P. D. Maker and R. W. Terhune, “Study of optical effects due to an induced polarization third order in the electric field strength,” Phys. Rev. 137, A801–A818 (1965).
[CrossRef]

Phys. Rev. A

H. Lotem, R. T. Lynch, Jr., and N. Bloembergen, “Interference between Raman resonances in four-wave difference mixing,” Phys. Rev. A 14, 1748–1755 (1976).
[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]

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, 0239011–0239014 (2001).
[CrossRef]

Proc. Natl. Acad. Sci. U.S.A.

E. O. Potma, W. P. de Boeij, P. J. M. van Haastert, and D. A. Wiersma, “Real-time visualisation of intracelluar hydrodynamics in single living cells,” Proc. Natl. Acad. Sci. U.S.A. 98, 1577–1582 (2001).
[CrossRef]

C. Xu, W. Zipfel, J. B. Shear, R. M. Williams, and W. W. Webb, “Multiphoton fluorescence excitation: new spectral windows for biological nonlinear microscopy,” Proc. Natl. Acad. Sci. U.S.A. 93, 10,763–10,768 (1996).
[CrossRef]

Proc. R. Soc. London, Ser. A

B. Richards and E. Wolf, “Electromagnetic diffraction in optical systems. II. Structure of the image field in an aplanatic system,” Proc. R. Soc. London, Ser. A 253, 358–379 (1959).
[CrossRef]

Science

W. Denk, J. H. Strickler, and W. W. Webb, “Two-photon laser scanning fluorescence microscopy,” Science 248, 73–76 (1990).
[CrossRef] [PubMed]

Other

Y. R. Shen, The Principles of Nonlinear Optics (Wiley, New York, 1984).

M. D. Levenson and S. S. Kano, Introduction to Nonlinear Laser Spectroscopy (Academic, San Diego, Calif., 1988).

J. Hoyland, “Fluorescent probes in practice—potential artifacts,” in Fluorescent and Luminescent Probes for Biological Activity, W. T. Mason, ed. (Academic, San Diego, Calif., 1999), p. 108.

H. J. Humecki, Practical Guide to Infrared and Microspectroscopy, Vol. 19 of Practical Spectroscopy, E. G. Brame, Jr., ed. (Marcel Dekker, New York, 1995).

G. Turrell and J. Corset, Raman Microscopy Development and Applications (Academic, San Diego, Calif., 1996).

R. J. H. Clark and R. E. Hester, eds., Advances in Non-linear Spectroscopy (Wiley, New York, 1988), Vol. 15.

E. Kohen and J. G. Hirschberg, Cell Structure and Function by Microspectrofluorometry (Academic, San Diego, Calif., 1989).

J. B. Pawley, Handbook of Biological Confocal Microscopy (Plenum, New York, 1995).

R. P. Haugland, Handbook of Fluorescent Probes and Research Chemicals, 6th ed. (Molecular Probes, Eugene, Ore., 1996).

T. Wilson and C. Sheppard, Theory and Practice of Scanning Optical Microscopy (Academic, Orlando, Fla., 1984).

S. Maeda, T. Kamisuki, and Y. Adachi, “Condensed phase CARS,” in Advances in Non-linear Spectroscopy, R. J. H. Clark and R. E. Hester, eds. (Wiley, New York, 1988), p. 253.

R. Brakel and F. W. Schneider, “Polarization CARS spectroscopy,” in Advances in Non-linear Spectroscopy, R. J. H. Clark and R. E. Hester, eds. (Wiley, New York, 1988), p. 149.

A. E. Siegman, Lasers (University Science Books, Mill Valley, Calif., 1986).

L. Novotny, Lecture Notes on “Nano-Optics” (University of Rochester, Rochester, N.Y., 2000).

The most significant term for CARS radiation involving the z-polarized field is χzxxz(3) Epx Epx (Esz)*, which is ~1% of χxxxx(3) Epx Epx (Esx)* with χzxxz(3)xxxx(3) /3. Thus the radiation power from the former term is only 0.01% of that from the latter term. Moreover, the radiation from the former term is maximized in the z=0 plane, so little signal can be detected in the forward or the backward direction. Experimentally, we observed that the nonresonant CARS signal was highly polarized along the same direction as the parallel-polarized pump and Stokes beams. This observation verifies the validity of our assumption.

R. W. Boyd, Nonlinear Optics (Academic, Boston, Mass., 1992).

S. Mukamel, Principles of Nonlinear Optical Spectroscopy (Oxford U. Press, New York, 1995).

W. C. Chew, Waves and Fields in Inhomogeneous Media, 2nd ed. (Institute of Electrical and Electronics Engineers, New York, 1995).

As our calculation did not consider the contribution from the solvent surrounding the scatterer, the forward radiation pattern is a suitable description of polarization CARS for which the nonresonant background from the solvent is suppressed. The forward CARS generated by a pair of parallel-polarized pump and Stokes beams is a coherent addition of the signal from the scatterer and that from the solvent and is always highly directional.

D. Lin-Vien, N. B. Colthup, W. G. Fateley, and J. G. Grasselli, Handbook of Infrared and Raman Characteristic Frequencies of Organic Molecules (Academic, San Diego, Calif., 1991).

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Figures (12)

Fig. 1
Fig. 1

(a) Illustration of the tight focusing of the incident Gaussian beams and the CARS radiation from a spherical sample with the definitions of the parameters used in the calculation. (b) Illustration of the polarization vectors for the excitation beams and the induced nonlinear polarization.

Fig. 2
Fig. 2

(a) Intensity distribution on a log scale and (b) axial phase shift of the focal field of a Gaussian beam focused by an objective lens of NA=1.4.

Fig. 3
Fig. 3

(a) Far-field CARS radiation pattern from spherical scatterers centered at the focus with different diameters. (b) Far-field CARS radiation pattern from scatterers centered at focus with the same volume but different shapes (rod, sphere, and disk). The rod has a diameter of 0.2 λp and an axial length of 2.0 λp. The sphere has a diameter of 0.78 λp. The disk has a diameter of 0.89 λp and a thickness of 0.1 λp. Shown in parentheses are the intensity ratios between samples of different sizes and different shapes. The radiation field is polarized along the x axis. The signals were calculated with the assumption of tightly focused (NA=1.4) incident beams copropagating along the +z axis and polarized along the x axis.

Fig. 4
Fig. 4

(a) Forward- and backward-detected signals as a function of the diameter of a spherical sample in a copropagating beam geometry. (b) The same as in (a) but for a hemispherical sample located in the z>0 region. (c) Forward- and backward-detected signals as a function of the diameter of a spherical sample in a counterpropagating beam geometry.

Fig. 5
Fig. 5

Forward-detected CARS signals as a function of diameter D of a spherical scatterer calculated with copropagating pump and Stokes beams and objective lenses with different NAs.

Fig. 6
Fig. 6

(a) Lateral image intensity profile as a function of the detuning from the vibrational resonance frequency, (ωp-ωs)-ωR, for a 1-µm spherical scatterer embedded in a nonlinear medium. (b), (c) Lateral and axial intensity profiles of two identical spherical scatterers with diameter 0.2 λp and separated by a distance d along the x and the z axes, respectively.

Fig. 7
Fig. 7

Schematic of the configurations for F-CARS, P-CARS, E-CARS, and C-CARS microscopes. P’s, polarizers; OL’s, objective lenses; S’s, samples; F’s, filters; HW, half-wave plate; D’s, dichroic mirrors.

Fig. 8
Fig. 8

(a) F-CARS and spontaneous Raman spectra of a polystyrene film spin coated onto a coverslip. The F-CARS spectrum was taken with an average pump power of 780 µW and an average Stokes power of 390 µW at a repetition rate of 100 kHz. The pump frequency was fixed at 13 325 cm-1. The Raman spectrum was recorded on a Raman spectrometer (Jobin-Yvon–Spec, LabRam). The F-CARS spectrum (solid curve) was simulated by Eq. (16). The parameters for the two Raman bands were set as ω1=1601 cm-1, ω2=1582 cm-1, Γ1=Γ2=3.5 cm-1, A2/A1=0.3, and χNR(3)/A1=0.55 cm. The pulse widths of the pump and the Stokes beams were chosen to be 2.9 cm-1. Shown below the spectra are the F-CARS images of 1-µm polystyrene beads upon a coverslip and covered with water. The pump and Stokes powers were 0.6 and 0.3 mW, respectively, at a repetition rate of 400 kHz. The size of each image was 5 µm×3 µm, and the acquisition time was 96 s for each image.

Fig. 9
Fig. 9

(a) P-CARS spectrum of a polystyrene film taken at an average pump power of 2 mW and an average Stokes power of 1 mW at a repetition rate of 400 kHz. The pump frequency was fixed at 13 325 cm-1. Shown below the spectrum are P-CARS images of 1-µm polystyrene beads upon a coverslip and covered with water. The pump and Stokes powers were 1.2 and 0.6 mW, respectively, at a repetition rate of 400 kHz. The size of the image was 4 µm×4 µm, and the acquisition time was 160 s for each image.

Fig. 10
Fig. 10

(a) E-CARS image of 0.2-µm beads embedded in 2% agarose gel; pump frequency, 14 183 cm-1. The pump and Stokes powers were 2.5 and 1.8 mW at an 800-kHz repetition rate. The image size was 3 µm×2 µm, and the acquisition time was 80 s. (b) C-CARS image of 0.5-µm beads in water. The pump and Stokes beams were two 110-fs pulse trains at 800 and 917 nm, respectively. The pump and Stokes powers were 100 and 50 µW, respectively, at a repetition rate of 250 kHz. The image size was 3 µm×2 µm, and the acquisition time was 126 s.

Fig. 11
Fig. 11

(a) F-CARS image of unstained epithelial cells. ωp-ωs was tuned to 1579 cm-1, with the pump frequency at 13 330 cm-1. The pump and Stokes powers were 0.4 and 0.2 mW at a 400-kHz repetition rate. The image size was 72 µm×72 µm, and the acquisition time was 12 min. (b) E-CARS image of unstained epithelial cells. ωp-ωs was tuned to 1570 cm-1, with the pump frequency at 13 333 cm-1. The pump and Stokes powers were 2.0 and 1.0 mW at a 400-kHz repetition rate. The image size was 75 µm×75 µm, and the acquisition time was 12 min. (c) P-CARS image of an unstained epithelial cell. ωp-ωs was tuned to 1650 cm-1, with the pump frequency at 13 324 cm-1. The pump and Stokes powers were 1.8 and 1.0 mW at a 400-kHz repetition rate. The image size was 50 µm×30 µm, and the acquisition time was 8 min. Shown below each image is the intensity profile along the line indicated by the two arrows.

Fig. 12
Fig. 12

Calculated ratio of forward to backward CARS radiation power as a function of diameter D for a spherical sample centered at the focus. The tightly focused field (NA=1.4) was calculated from the angular spectrum representation [Eq. (4)] and from the paraxial approximation [Eq. (A1)].

Equations (23)

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Ep(r, t)=Ep(r)exp(-iωpt)+c.c.,
Es(r, t)=Es(r)exp(-iωst)+c.c.
Ejinc(α)=Ej0 exp(-f2 sin2 α/w02),
Ej(ρ, φ, z)=ikjf exp(-ikjf)2 I00+I02 cos 2φI02 sin 2φ-i2I01 cos φ,
I0m=0αmax Ejinc(α)sin αcos αgm(α)Jm(kjρ sin α)×exp(kz cos α)dα,
××E(r, t)+n2c2 E(r, t)t2=-4πc2 2PNL(r, t)t2.
PNL(r, t)=P(3)(r)exp(-iωast)+c.c.,
E(r, t)=Eas(r)exp(-iωast)+c.c.
××Eas(r)-kas2Eas(r)=4πωas2c2 P(3)(r).
2Eas(r)+kas2Eas(r)=-4πωas2c2 Iˆ+kas2·P(3)(r),
Eas(R)=-4πωas2c2 V dVIˆ+kas2G(R-r)·P(3)(r).
G(R-r)=exp(ikas|R-r|)/4π|R-r|.
Eas(R)=-ωas2c2 exp(ikas|R|)|R| V dV exp-ikasR·r|R|×000cos Θ cos Φcos Θ sin Φ-sin Θ-sin Φcos Φ0×Px(3)(r)Py(3)(r)Pz(3)(r)iˆRiˆΘiˆΦ,
pCARS=nasc8π Θ1Θ2 dΘ 02π dΦ|Eas(R)|2R2 sin Θ.
P(3)(r)=3χ1111(3)(ωas, r)Ep2(r)Es*(r)iˆx.
χ1111(3)(ωas)=ηNR+Atωt-2ωp-iΓt+Atωt-(ωp+ωs)-iΓt+Atωt-2ωs-iΓi+RARωR-(ωp-ωs)-iΓR.
PxNR=3χ1111(3)NREp2Es* cos ϕ,
PyNR=3χ1221(3)NREp2Es* sin ϕ.
PxR=3χ1111(3)REp2Es* cos ϕ,PyR=3χ1221(3)REp2Es* sin ϕ.
PNR(3)(r)=3χ1111(3)NR(ωas)Ep2(r)Es*(r)cos ϕ/cos ϕ,
P(3)(r)=3χ1111(3)R(ωas)Ep2(r)Es*(r)cos ϕ sin θ(1-ρR/ρNR).
P2/(PNR2/r)=r[χ1111(3)R/χ1111(3)NR]2(1-ρR/ρNR)2 sin2 θ cos2 θ.
E(ρ, z)=E0 w0w(z)×exp-ρ2w(z)2expikz-η(z)+kρ22R(z),

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