Abstract

We demonstrate coherent anti-Stokes Raman scattering (CARS) microscopy of lipid-rich structures using a single unamplified femtosecond Ti:sapphire laser and a photonic crystal fiber (PCF) with two closely lying zero dispersion wavelengths (ZDW) for the Stokes source. The primary enabling factor for the fast data acquisition (84 μs per pixel) in the proof-of-principle CARS images, is the low noise supercontinuum (SC) generated in this type of PCF, in contrast to SC generated in a PCF with one ZDW. The dependence of the Stokes pulse on average input power, pump wavelength, pulse duration and polarization is experimentally characterized. We show that it is possible to control the spectral shape of the SC by tuning the pump wavelength of the input pulse and the consequence for CARS microscopy is discussed.

© 2007 Optical Society of America

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  1. 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]
  2. F. Ganikhanov, S. Carrasco, X. S. Xie, M. Katz, W. Seitz and D. Kopf, "Broadly tunable dual-wavelength light source for coherent anti-Stokes Raman scattering microscopy," Opt. Lett. 31, 1292-1294 (2006).
    [CrossRef] [PubMed]
  3. C. L. Evans, E. O., Potma, M Puoris’haag., D. Côté, C. P. Lin, and S. Xie, "Chemical imaging of tissue in vivo with video-rate coherent anti-Stokes Raman scattering microscopy,". Proc. Natl. Acad. Sci. USA 102, 16807-16812 (2005).
    [CrossRef] [PubMed]
  4. T. 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]
  5. 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]
  6. E. R. Andresen, H. N. Paulsen, V. Birkedal, J. Thogersen and S. R. Keiding, "Broadband multiplex coherent anti-Stokes Raman scattering microscopy employing photonic crystal fibers," J. Opt. Soc. Am. B 22, 1934 - 1938 (2005).
    [CrossRef]
  7. E. R. Andresen, C. K. Nielsen, J. Thøgersen and S. R. Keiding "Fiber laser-based light source for coherent anti-Stokes Raman scattering microspectroscopy," Opt. Express 15, 4848 - 4856 (2007).
    [CrossRef] [PubMed]
  8. N. R. Newbury, B. R. Washburn, K. L. Corwin, and R. S. Windeler, "Noise amplification during supercontinuum generation in microstructure fiber," Opt. Lett. 28, 944 - 946 (2003).
    [CrossRef] [PubMed]
  9. K. L. Corwin, N. R. Newbury, J. M. Dudley, S. Coen, S. A. Diddams, K. Weber, and R. S. Windeler "Fundamental Noise Limitations to Supercontinuum Generation in Microstructure Fiber," Phys. Rev. Lett.  90, 113904 (2003).
    [CrossRef] [PubMed]
  10. J. M. Dudley, G. Genty, S. Coen, "Supercontinuum generation in photonic crystal fiber," Rev. Mod. Phys 78, 1135 - 1184 (2006).
    [CrossRef]
  11. K. Hilligsoe, T. Andersen, H. Paulsen, C. Nielsen, K. Molmer, S. Keiding, R. Kristiansen, K. Hansen and J. Larsen, "Supercontinuum generation in a photonic crystal fiber with two zero dispersion wavelengths," Opt. Express 12, 1045 - 1054 (2004).
    [CrossRef] [PubMed]
  12. A. Aguirre, N. Nisizawa, J. Fujimoto, W. Seitz, M. Lederer, and D. Kopf, "Continuum generation in a novel photonic crystal fiber for ultrahigh resolution optical coherence tomography at 800 nm and 1300 nm," Opt. Express 14, 1145-1160 (2006).
    [CrossRef] [PubMed]
  13. A. Apolonski, B. Povazay, A. Unterhuber, W. Drexler, W. J. Wadsworth, J. C. Knight, and P. St. J. Russell, "Spectral shaping of supercontinuum in a cobweb photonic-crystal fiber with sub-20-fs pulses," J. Opt. Soc. Am. B 19, 2165-2170 (2002).
    [CrossRef]
  14. M. H. Frosz, P. Falk, and O. Bang, "The role of the second zero-dispersion wavelength in generation of supercontinua and bright-bright soliton-pairs across the zero-dispersion wavelength," Opt. Express 13, 6181-6192 (2005).
    [CrossRef] [PubMed]
  15. H. Zhang, S. Yu, J. Zhang, and W. Gu, "Effect of frequency chirp on supercontinuum generation in photonic crystal fibers with two zero-dispersion wavelengths," Opt. Express 15, 1147-1154 (2007).
    [CrossRef] [PubMed]
  16. R. L. Fork, O. E. Martinez, and J. P. Gordon, "Negative dispersion using pair of prisms," Opt. Lett. 9, 150-152 (1984).
    [CrossRef] [PubMed]
  17. H. Wang and A.M. Rollins, "Optimization of dual-band continuum light source for ultrahigh-resolution optical coherence tomography," Appl. Opt. 46, 1787-1794 (2007).
    [CrossRef] [PubMed]
  18. M. Pezolet and D Georgescauld, "Raman Spectroscopy of Nerve Fibers. A study of membrane lipids under steady state conditions," Biophys. J. 47, 367-372 (1985).
    [CrossRef] [PubMed]
  19. K. P. Knutsen, J. C. Johnson, A. E. Miller, P. B. Petersen, and R. J. Saykally, "High spectral resolution CARS spectroscopy using chirped pulses," Chem. Phys. Lett. 387, 436-441 (2004).
    [CrossRef]
  20. E.H.K. Stelzer, "The Intermediate Optical System in Confocal Microscopes," in The handbook of biological confocal microscopy, J. Pawley, ed. (IMR Press: Madison. 1989).
  21. H. Wang, Y. Fu, P. Zickmund, Riyi Shi and Ji-Xin Cheng, "Coherent anti-Stokes Raman scattering imaging of axonal myelin in live spinal tissues," Biophys. J. 89, 581-591 (2005).
    [CrossRef] [PubMed]
  22. H. Kano and H. Hamaguchi, "In-vivo multi-nonlinear optical imaging of a living cell using a supercontinuum light source generated from a photonic crystal fiber," Opt. Express 14, 2798 - 2804 (2006).
    [CrossRef] [PubMed]
  23. J. A. Palero, V. O. Boer, J. C. Vijverberg, H. C. Gerritsen, and H. J. C. M. Sterenborg, "Short-wavelength twophoton excitation fluorescence microscopy of tryptophan with a photonic crystal fiber based light source," Opt. Express 13, 5363-5368 (2005).
    [CrossRef] [PubMed]

2007 (3)

2006 (4)

2005 (6)

2004 (4)

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]

T. 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]

K. Hilligsoe, T. Andersen, H. Paulsen, C. Nielsen, K. Molmer, S. Keiding, R. Kristiansen, K. Hansen and J. Larsen, "Supercontinuum generation in a photonic crystal fiber with two zero dispersion wavelengths," Opt. Express 12, 1045 - 1054 (2004).
[CrossRef] [PubMed]

K. P. Knutsen, J. C. Johnson, A. E. Miller, P. B. Petersen, and R. J. Saykally, "High spectral resolution CARS spectroscopy using chirped pulses," Chem. Phys. Lett. 387, 436-441 (2004).
[CrossRef]

2003 (2)

N. R. Newbury, B. R. Washburn, K. L. Corwin, and R. S. Windeler, "Noise amplification during supercontinuum generation in microstructure fiber," Opt. Lett. 28, 944 - 946 (2003).
[CrossRef] [PubMed]

K. L. Corwin, N. R. Newbury, J. M. Dudley, S. Coen, S. A. Diddams, K. Weber, and R. S. Windeler "Fundamental Noise Limitations to Supercontinuum Generation in Microstructure Fiber," Phys. Rev. Lett.  90, 113904 (2003).
[CrossRef] [PubMed]

2002 (1)

1985 (1)

M. Pezolet and D Georgescauld, "Raman Spectroscopy of Nerve Fibers. A study of membrane lipids under steady state conditions," Biophys. J. 47, 367-372 (1985).
[CrossRef] [PubMed]

1984 (1)

Aguirre, A.

Andersen, T.

Andresen, E. R.

Apolonski, A.

Bang, O.

Birkedal, V.

Boer, V. O.

Carrasco, S.

Cheng, J. X.

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]

Cicerone, M. T.

Coen, S.

J. M. Dudley, G. Genty, S. Coen, "Supercontinuum generation in photonic crystal fiber," Rev. Mod. Phys 78, 1135 - 1184 (2006).
[CrossRef]

K. L. Corwin, N. R. Newbury, J. M. Dudley, S. Coen, S. A. Diddams, K. Weber, and R. S. Windeler "Fundamental Noise Limitations to Supercontinuum Generation in Microstructure Fiber," Phys. Rev. Lett.  90, 113904 (2003).
[CrossRef] [PubMed]

Corwin, K. L.

K. L. Corwin, N. R. Newbury, J. M. Dudley, S. Coen, S. A. Diddams, K. Weber, and R. S. Windeler "Fundamental Noise Limitations to Supercontinuum Generation in Microstructure Fiber," Phys. Rev. Lett.  90, 113904 (2003).
[CrossRef] [PubMed]

N. R. Newbury, B. R. Washburn, K. L. Corwin, and R. S. Windeler, "Noise amplification during supercontinuum generation in microstructure fiber," Opt. Lett. 28, 944 - 946 (2003).
[CrossRef] [PubMed]

Diddams, S. A.

K. L. Corwin, N. R. Newbury, J. M. Dudley, S. Coen, S. A. Diddams, K. Weber, and R. S. Windeler "Fundamental Noise Limitations to Supercontinuum Generation in Microstructure Fiber," Phys. Rev. Lett.  90, 113904 (2003).
[CrossRef] [PubMed]

Drexler, W.

Dudley, J. M.

J. M. Dudley, G. Genty, S. Coen, "Supercontinuum generation in photonic crystal fiber," Rev. Mod. Phys 78, 1135 - 1184 (2006).
[CrossRef]

K. L. Corwin, N. R. Newbury, J. M. Dudley, S. Coen, S. A. Diddams, K. Weber, and R. S. Windeler "Fundamental Noise Limitations to Supercontinuum Generation in Microstructure Fiber," Phys. Rev. Lett.  90, 113904 (2003).
[CrossRef] [PubMed]

Evans, C. L.

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

Falk, P.

Fork, R. L.

Frosz, M. H.

Fu, Y.

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

Fujimoto, J.

Ganikhanov, F.

Genty, G.

J. M. Dudley, G. Genty, S. Coen, "Supercontinuum generation in photonic crystal fiber," Rev. Mod. Phys 78, 1135 - 1184 (2006).
[CrossRef]

Georgescauld, D

M. Pezolet and D Georgescauld, "Raman Spectroscopy of Nerve Fibers. A study of membrane lipids under steady state conditions," Biophys. J. 47, 367-372 (1985).
[CrossRef] [PubMed]

Gerritsen, H. C.

Gordon, J. P.

Gu, W.

Hamaguchi, H.

Hansen, K.

Hilligsoe, K.

Johnson, J. C.

K. P. Knutsen, J. C. Johnson, A. E. Miller, P. B. Petersen, and R. J. Saykally, "High spectral resolution CARS spectroscopy using chirped pulses," Chem. Phys. Lett. 387, 436-441 (2004).
[CrossRef]

Kano, H.

Katz, M.

Kee, T.

Keiding, S.

Keiding, S. R.

Knight, J. C.

Knutsen, K. P.

K. P. Knutsen, J. C. Johnson, A. E. Miller, P. B. Petersen, and R. J. Saykally, "High spectral resolution CARS spectroscopy using chirped pulses," Chem. Phys. Lett. 387, 436-441 (2004).
[CrossRef]

Kopf, D.

Kristiansen, R.

Larsen, J.

Lederer, M.

Martinez, O. E.

Miller, A. E.

K. P. Knutsen, J. C. Johnson, A. E. Miller, P. B. Petersen, and R. J. Saykally, "High spectral resolution CARS spectroscopy using chirped pulses," Chem. Phys. Lett. 387, 436-441 (2004).
[CrossRef]

Molmer, K.

Newbury, N. R.

N. R. Newbury, B. R. Washburn, K. L. Corwin, and R. S. Windeler, "Noise amplification during supercontinuum generation in microstructure fiber," Opt. Lett. 28, 944 - 946 (2003).
[CrossRef] [PubMed]

K. L. Corwin, N. R. Newbury, J. M. Dudley, S. Coen, S. A. Diddams, K. Weber, and R. S. Windeler "Fundamental Noise Limitations to Supercontinuum Generation in Microstructure Fiber," Phys. Rev. Lett.  90, 113904 (2003).
[CrossRef] [PubMed]

Nielsen, C.

Nielsen, C. K.

Nisizawa, N.

Palero, J. A.

Paulsen, H.

Paulsen, H. N.

Petersen, P. B.

K. P. Knutsen, J. C. Johnson, A. E. Miller, P. B. Petersen, and R. J. Saykally, "High spectral resolution CARS spectroscopy using chirped pulses," Chem. Phys. Lett. 387, 436-441 (2004).
[CrossRef]

Pezolet, M.

M. Pezolet and D Georgescauld, "Raman Spectroscopy of Nerve Fibers. A study of membrane lipids under steady state conditions," Biophys. J. 47, 367-372 (1985).
[CrossRef] [PubMed]

Povazay, B.

Rollins, A.M.

Russell, P. St. J.

Saykally, R. J.

K. P. Knutsen, J. C. Johnson, A. E. Miller, P. B. Petersen, and R. J. Saykally, "High spectral resolution CARS spectroscopy using chirped pulses," Chem. Phys. Lett. 387, 436-441 (2004).
[CrossRef]

Seitz, W.

Sterenborg, H. J. C. M.

Thogersen, J.

Thøgersen, J.

Unterhuber, A.

Vijverberg, J. C.

Wadsworth, W. J.

Wang, H.

H. Wang and A.M. Rollins, "Optimization of dual-band continuum light source for ultrahigh-resolution optical coherence tomography," Appl. Opt. 46, 1787-1794 (2007).
[CrossRef] [PubMed]

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

Washburn, B. R.

Weber, K.

K. L. Corwin, N. R. Newbury, J. M. Dudley, S. Coen, S. A. Diddams, K. Weber, and R. S. Windeler "Fundamental Noise Limitations to Supercontinuum Generation in Microstructure Fiber," Phys. Rev. Lett.  90, 113904 (2003).
[CrossRef] [PubMed]

Windeler, R. S.

K. L. Corwin, N. R. Newbury, J. M. Dudley, S. Coen, S. A. Diddams, K. Weber, and R. S. Windeler "Fundamental Noise Limitations to Supercontinuum Generation in Microstructure Fiber," Phys. Rev. Lett.  90, 113904 (2003).
[CrossRef] [PubMed]

N. R. Newbury, B. R. Washburn, K. L. Corwin, and R. S. Windeler, "Noise amplification during supercontinuum generation in microstructure fiber," Opt. Lett. 28, 944 - 946 (2003).
[CrossRef] [PubMed]

Xie, X. S.

F. Ganikhanov, S. Carrasco, X. S. Xie, M. Katz, W. Seitz and D. Kopf, "Broadly tunable dual-wavelength light source for coherent anti-Stokes Raman scattering microscopy," Opt. Lett. 31, 1292-1294 (2006).
[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]

Yu, S.

Zhang, H.

Zhang, J.

Zickmund, P.

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

Appl. Opt. (1)

Biophys. J. (2)

M. Pezolet and D Georgescauld, "Raman Spectroscopy of Nerve Fibers. A study of membrane lipids under steady state conditions," Biophys. J. 47, 367-372 (1985).
[CrossRef] [PubMed]

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

Chem. Phys. Lett. (1)

K. P. Knutsen, J. C. Johnson, A. E. Miller, P. B. Petersen, and R. J. Saykally, "High spectral resolution CARS spectroscopy using chirped pulses," Chem. Phys. Lett. 387, 436-441 (2004).
[CrossRef]

J. Opt. Soc. Am. B (2)

J. Phys. Chem B (1)

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]

Opt. Express (8)

M. H. Frosz, P. Falk, and O. Bang, "The role of the second zero-dispersion wavelength in generation of supercontinua and bright-bright soliton-pairs across the zero-dispersion wavelength," Opt. Express 13, 6181-6192 (2005).
[CrossRef] [PubMed]

A. Aguirre, N. Nisizawa, J. Fujimoto, W. Seitz, M. Lederer, and D. Kopf, "Continuum generation in a novel photonic crystal fiber for ultrahigh resolution optical coherence tomography at 800 nm and 1300 nm," Opt. Express 14, 1145-1160 (2006).
[CrossRef] [PubMed]

H. Kano and H. Hamaguchi, "In-vivo multi-nonlinear optical imaging of a living cell using a supercontinuum light source generated from a photonic crystal fiber," Opt. Express 14, 2798 - 2804 (2006).
[CrossRef] [PubMed]

K. Hilligsoe, T. Andersen, H. Paulsen, C. Nielsen, K. Molmer, S. Keiding, R. Kristiansen, K. Hansen and J. Larsen, "Supercontinuum generation in a photonic crystal fiber with two zero dispersion wavelengths," Opt. Express 12, 1045 - 1054 (2004).
[CrossRef] [PubMed]

E. R. Andresen, C. K. Nielsen, J. Thøgersen and S. R. Keiding "Fiber laser-based light source for coherent anti-Stokes Raman scattering microspectroscopy," Opt. Express 15, 4848 - 4856 (2007).
[CrossRef] [PubMed]

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]

J. A. Palero, V. O. Boer, J. C. Vijverberg, H. C. Gerritsen, and H. J. C. M. Sterenborg, "Short-wavelength twophoton excitation fluorescence microscopy of tryptophan with a photonic crystal fiber based light source," Opt. Express 13, 5363-5368 (2005).
[CrossRef] [PubMed]

H. Zhang, S. Yu, J. Zhang, and W. Gu, "Effect of frequency chirp on supercontinuum generation in photonic crystal fibers with two zero-dispersion wavelengths," Opt. Express 15, 1147-1154 (2007).
[CrossRef] [PubMed]

Opt. Lett. (4)

Phys. Rev. Lett. (1)

K. L. Corwin, N. R. Newbury, J. M. Dudley, S. Coen, S. A. Diddams, K. Weber, and R. S. Windeler "Fundamental Noise Limitations to Supercontinuum Generation in Microstructure Fiber," Phys. Rev. Lett.  90, 113904 (2003).
[CrossRef] [PubMed]

Proc. Natl. Acad. Sci. USA (1)

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

Rev. Mod. Phys (1)

J. M. Dudley, G. Genty, S. Coen, "Supercontinuum generation in photonic crystal fiber," Rev. Mod. Phys 78, 1135 - 1184 (2006).
[CrossRef]

Other (1)

E.H.K. Stelzer, "The Intermediate Optical System in Confocal Microscopes," in The handbook of biological confocal microscopy, J. Pawley, ed. (IMR Press: Madison. 1989).

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Figures (5)

Fig 1.
Fig 1.

(Color online) Measured spectral output of the PCF as a function of average power coupled into the core. The vertical straight lines indicate the position of the two ZDWs. Excitation pulse is at 810 nm and is ~ 100 fs wide.

Fig. 2.
Fig. 2.

(Color online) Dependence of SC on pump wavelength. Output power in the PCF is constant at 140 mW and pulse duration is ~ 100 fs.

Fig. 3.
Fig. 3.

(Color online) Dependence of SC on frequency chirped (indicated as up and down chirp) pulses. Pump wavelength is at 810 nm and output power of the PCF is 140 mW.

Fig. 4.
Fig. 4.

(Color online) Schematic of the CARS microscope setup. PCF- Photonic Crystal Fiber with two ZDWs; Iso - Faraday Isolator; DM- Dichroic mirror; Com -Prism compressor; SM - Scanning mirror assembly; HW - Half wave plate; BP - Band Pass filter; BS - Beam Splitter; Obj - Objective; PMT - Photo Multiplier Tube; Fiber - multimode fiber to guide the CARS signal into the PMT.

Fig. 5.
Fig. 5.

Proof-of-principle forward - CARS images at Raman shift of 2840 cm-1 from (a) isolated unstained live rat dorsal root axons (b) lipid droplets in unlabeled 3T3 L1 adipocyte cell culture (c) sebaceous gland (in pseudo colour) in a mouse ear at a depth of ~ 40 μm and (d) adipocyte cells at a depth of 100 μm in a mouse ear imaged in the forward mode through several hundred microns of tissue. (e) same as in (d) but with Stokes beam blocked.

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