Abstract

We report an approach to coherent anti-Stokes Raman scattering (CARS) microscopy that significantly suppresses the nonresonant background for high-contrast vibrational imaging. We employ an elliptically polarized pump field with an aspect ratio of 1:3 of the ellipse combined with a linearly polarized Stokes field for elliptically polarized CARS (EP-CARS) microscopy. The EP-CARS technique yields a 6–7-fold improvement in signal-to-background ratios compared with normal CARS, while providing an approximately 1.25-fold improvement in resonant CARS signal compared with normal polarization-sensitive CARS microscopy. We demonstrate this technique by imaging 1.5μm polystyrene beads in water and lipid droplets in unstained fibroblast cells.

© 2008 Optical Society of America

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  1. A. Zumbushch, G. R. Holtom, and X. S. Xie, Phys. Rev. Lett. 82, 4142 (1999).
    [CrossRef]
  2. H. Wang, Y. Fu, P. Zickmund, R. Shi, and J. X. Cheng, Biophys. J. 89, 581 (2005).
    [CrossRef] [PubMed]
  3. Y. R. Shen, The Principles of Nonlinear Optics (Wiley, 1984).
  4. F. Lu, W. Zheng, and Z. Huang, Appl. Phys. Lett. 92, 123901 (2008).
    [CrossRef]
  5. A. Volkmer, J. X. Cheng, and X. S. Xie, Phys. Rev. Lett. 87, 023901 (2001).
    [CrossRef]
  6. J. X. Cheng, L. D. Book, and X. S. Xie, Opt. Lett. 26, 1341 (2001).
    [CrossRef]
  7. A. Volkmer, L. D. Book, and X. S. Xie, Appl. Phys. Lett. 80, 1505 (2002).
    [CrossRef]
  8. F. Ganikhanov, C. L. Evans, B. G. Saar, and X. S. Xie, Opt. Lett. 31, 1872 (2006).
    [CrossRef] [PubMed]
  9. F. Lu, W. Zheng, C. Sheppard, and Z. Huang, Opt. Lett. 33, 602 (2008).
    [CrossRef] [PubMed]
  10. E. O. Potma, C. L. Evans, and X. S. Xie, Opt. Lett. 31, 241 (2006).
    [CrossRef] [PubMed]

2008 (2)

F. Lu, W. Zheng, and Z. Huang, Appl. Phys. Lett. 92, 123901 (2008).
[CrossRef]

F. Lu, W. Zheng, C. Sheppard, and Z. Huang, Opt. Lett. 33, 602 (2008).
[CrossRef] [PubMed]

2006 (2)

2005 (1)

H. Wang, Y. Fu, P. Zickmund, R. Shi, and J. X. Cheng, Biophys. J. 89, 581 (2005).
[CrossRef] [PubMed]

2002 (1)

A. Volkmer, L. D. Book, and X. S. Xie, Appl. Phys. Lett. 80, 1505 (2002).
[CrossRef]

2001 (2)

J. X. Cheng, L. D. Book, and X. S. Xie, Opt. Lett. 26, 1341 (2001).
[CrossRef]

A. Volkmer, J. X. Cheng, and X. S. Xie, Phys. Rev. Lett. 87, 023901 (2001).
[CrossRef]

1999 (1)

A. Zumbushch, G. R. Holtom, and X. S. Xie, Phys. Rev. Lett. 82, 4142 (1999).
[CrossRef]

1984 (1)

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

Book, L. D.

A. Volkmer, L. D. Book, and X. S. Xie, Appl. Phys. Lett. 80, 1505 (2002).
[CrossRef]

J. X. Cheng, L. D. Book, and X. S. Xie, Opt. Lett. 26, 1341 (2001).
[CrossRef]

Cheng, J. X.

H. Wang, Y. Fu, P. Zickmund, R. Shi, and J. X. Cheng, Biophys. J. 89, 581 (2005).
[CrossRef] [PubMed]

A. Volkmer, J. X. Cheng, and X. S. Xie, Phys. Rev. Lett. 87, 023901 (2001).
[CrossRef]

J. X. Cheng, L. D. Book, and X. S. Xie, Opt. Lett. 26, 1341 (2001).
[CrossRef]

Evans, C. L.

Fu, Y.

H. Wang, Y. Fu, P. Zickmund, R. Shi, and J. X. Cheng, Biophys. J. 89, 581 (2005).
[CrossRef] [PubMed]

Ganikhanov, F.

Holtom, G. R.

A. Zumbushch, G. R. Holtom, and X. S. Xie, Phys. Rev. Lett. 82, 4142 (1999).
[CrossRef]

Huang, Z.

F. Lu, W. Zheng, C. Sheppard, and Z. Huang, Opt. Lett. 33, 602 (2008).
[CrossRef] [PubMed]

F. Lu, W. Zheng, and Z. Huang, Appl. Phys. Lett. 92, 123901 (2008).
[CrossRef]

Lu, F.

F. Lu, W. Zheng, and Z. Huang, Appl. Phys. Lett. 92, 123901 (2008).
[CrossRef]

F. Lu, W. Zheng, C. Sheppard, and Z. Huang, Opt. Lett. 33, 602 (2008).
[CrossRef] [PubMed]

Potma, E. O.

Saar, B. G.

Shen, Y. R.

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

Sheppard, C.

Shi, R.

H. Wang, Y. Fu, P. Zickmund, R. Shi, and J. X. Cheng, Biophys. J. 89, 581 (2005).
[CrossRef] [PubMed]

Volkmer, A.

A. Volkmer, L. D. Book, and X. S. Xie, Appl. Phys. Lett. 80, 1505 (2002).
[CrossRef]

A. Volkmer, J. X. Cheng, and X. S. Xie, Phys. Rev. Lett. 87, 023901 (2001).
[CrossRef]

Wang, H.

H. Wang, Y. Fu, P. Zickmund, R. Shi, and J. X. Cheng, Biophys. J. 89, 581 (2005).
[CrossRef] [PubMed]

Xie, X. S.

E. O. Potma, C. L. Evans, and X. S. Xie, Opt. Lett. 31, 241 (2006).
[CrossRef] [PubMed]

F. Ganikhanov, C. L. Evans, B. G. Saar, and X. S. Xie, Opt. Lett. 31, 1872 (2006).
[CrossRef] [PubMed]

A. Volkmer, L. D. Book, and X. S. Xie, Appl. Phys. Lett. 80, 1505 (2002).
[CrossRef]

A. Volkmer, J. X. Cheng, and X. S. Xie, Phys. Rev. Lett. 87, 023901 (2001).
[CrossRef]

J. X. Cheng, L. D. Book, and X. S. Xie, Opt. Lett. 26, 1341 (2001).
[CrossRef]

A. Zumbushch, G. R. Holtom, and X. S. Xie, Phys. Rev. Lett. 82, 4142 (1999).
[CrossRef]

Zheng, W.

F. Lu, W. Zheng, C. Sheppard, and Z. Huang, Opt. Lett. 33, 602 (2008).
[CrossRef] [PubMed]

F. Lu, W. Zheng, and Z. Huang, Appl. Phys. Lett. 92, 123901 (2008).
[CrossRef]

Zickmund, P.

H. Wang, Y. Fu, P. Zickmund, R. Shi, and J. X. Cheng, Biophys. J. 89, 581 (2005).
[CrossRef] [PubMed]

Zumbushch, A.

A. Zumbushch, G. R. Holtom, and X. S. Xie, Phys. Rev. Lett. 82, 4142 (1999).
[CrossRef]

Appl. Phys. Lett. (2)

F. Lu, W. Zheng, and Z. Huang, Appl. Phys. Lett. 92, 123901 (2008).
[CrossRef]

A. Volkmer, L. D. Book, and X. S. Xie, Appl. Phys. Lett. 80, 1505 (2002).
[CrossRef]

Biophys. J. (1)

H. Wang, Y. Fu, P. Zickmund, R. Shi, and J. X. Cheng, Biophys. J. 89, 581 (2005).
[CrossRef] [PubMed]

Opt. Lett. (4)

Phys. Rev. Lett. (2)

A. Volkmer, J. X. Cheng, and X. S. Xie, Phys. Rev. Lett. 87, 023901 (2001).
[CrossRef]

A. Zumbushch, G. R. Holtom, and X. S. Xie, Phys. Rev. Lett. 82, 4142 (1999).
[CrossRef]

Other (1)

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

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Figures (4)

Fig. 1
Fig. 1

Polarization vectors of the elliptically polarized pump field comprising the two linear components of E p 1 and E p 2 , the linearly polarized Stokes field E s , and the generated CARS radiations ( P 1221 ( 3 ) n r , P 1221 ( 3 ) r , P 1111 ( 3 ) n r , P 1111 ( 3 ) r , P 2211 ( 3 ) , and P 2121 ( 3 ) ), as well as the orientation of the polarization analyzer A along the x axis.

Fig. 2
Fig. 2

Schematic of the elliptically polarized coherent anti-Stokes Raman scattering (EP-CARS) microscope. An elliptically polarized pump beam ( ω p ) and a linearly polarized Stokes beam ( ω s ) are collinearly combined and delivered into a laser-scanning confocal microscope for CARS imaging. BS, beam splitter; M, mirror; P, polarizer; HW, half-wave plate; QW, quarter-wave plate; DM, dichroic mirror; MO, microscope objective; F, filter set; L, lens; PMT, photomultiplier tube.

Fig. 3
Fig. 3

Comparison of CARS images (aromatic C–H stretching vibration centered at 3054 cm 1 , FWHM, 150 cm 1 ) of 1.5 μ m polystyrene beads immersed in water: (a) normal CARS, (b) EP-CARS, and (c) normal P-CARS. (d)–(f) are the corresponding intensity profiles across the lines indicated in images (a)–(c). The average powers of the pump beam ( 835 nm ) and Stokes beam ( 1121 nm ) on the sample are 2 and 0.5 mW , respectively.

Fig. 4
Fig. 4

CARS images (symmetric C H 2 stretching vibration centered at 2845 cm 1 ; FWHM, 150 cm 1 ) of lipid droplets in an unstained fibroblast cell: (a) normal CARS and (b) EP-CARS. (c) and (d) are the corresponding intensity profiles across the lines indicated in images (a) and (b), respectively. The average powers of the pump beam ( 835 nm ) and Stokes beam ( 1095 nm ) on the cell are 1.6 and 0.4 mW , respectively.

Equations (5)

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E p 2 = α E p 1 e i π 2 ,
P x ( 3 ) = P 1111 ( 3 ) n r + P 1111 ( 3 ) r + P 1221 ( 3 ) n r + P 1221 ( 3 ) r = 3 χ 1111 ( 3 ) n r E p 1 E p 1 E s * + 3 χ 1111 ( 3 ) r E p 1 E p 1 E s * + 3 χ 1221 ( 3 ) n r E p 2 E p 2 E s * + 3 χ 1221 ( 3 ) r E p 2 E p 2 E s * .
P x ( 3 ) = 3 [ ( 1 ρ n r α 2 ) χ 1111 ( 3 ) n r + ( 1 ρ r α 2 ) χ 1111 ( 3 ) r ] E p 1 E p 1 E s * .
I EP - CARS P x ( 3 ) 2 = 9 ( 1 3 ρ r ) χ 1111 ( 3 ) r E p 1 E p 1 E s * 2 .
I P - CARS 3 E p 2 E s * χ 1111 ( 3 ) r ( cos ϕ sin θ ρ r sin ϕ cos θ ) 2 = 0.8 I EP - CARS .

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