Abstract

We demonstrate the three-dimensional (3D) imaging capabilities and chemical specificity of multiplex coherent anti-Stokes Raman scattering microscopy. The simultaneous acquisition of a significant part of the vibrational spectrum at each specimen position permits straightforward differentiation among chemical species. 3D imaging is illustrated with a lipid multilamellar vesicle, and lateral and axial resolutions are determined.

© 2002 Optical Society of America

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2002

M. Müller and J. M. Schins, J. Phys. Chem. B 106, 3715 (2002).
[CrossRef]

2001

J. X. Cheng, A. Volkmer, L. D. Book, and X. S. Xie, J. Phys. Chem. B 105, 1277 (2001).
[CrossRef]

E. O. Potma, W. P. de Boeij, P. J. M. van Haastert, and D. A. Wiersma, Proc. Natl. Acad. Sci. (USA) 98, 1577 (2001).
[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]

2000

M. Müller, J. Squier, C. A. de Lange, and G. J. Brakenhoff, J. Microsc. 197, 150 (2000).
[CrossRef]

M. Hashimoto and T. Araki, Opt. Lett. 25, 1768 (2000).
[CrossRef]

1999

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

1982

Araki, T.

Book, L. D.

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

J. X. Cheng, A. Volkmer, L. D. Book, and X. S. Xie, J. Phys. Chem. B 105, 1277 (2001).
[CrossRef]

Brakenhoff, G. J.

M. Müller, J. Squier, C. A. de Lange, and G. J. Brakenhoff, J. Microsc. 197, 150 (2000).
[CrossRef]

Cheng, J. X.

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]

J. X. Cheng, A. Volkmer, L. D. Book, and X. S. Xie, J. Phys. Chem. B 105, 1277 (2001).
[CrossRef]

de Boeij, W. P.

E. O. Potma, W. P. de Boeij, P. J. M. van Haastert, and D. A. Wiersma, Proc. Natl. Acad. Sci. (USA) 98, 1577 (2001).
[CrossRef]

de Lange, C. A.

M. Müller, J. Squier, C. A. de Lange, and G. J. Brakenhoff, J. Microsc. 197, 150 (2000).
[CrossRef]

Duncan, M. D.

Hashimoto, M.

Holtom, G. R.

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

Manuccia, T. J.

Müller, M.

M. Müller and J. M. Schins, J. Phys. Chem. B 106, 3715 (2002).
[CrossRef]

M. Müller, J. Squier, C. A. de Lange, and G. J. Brakenhoff, J. Microsc. 197, 150 (2000).
[CrossRef]

Potma, E. O.

E. O. Potma, W. P. de Boeij, P. J. M. van Haastert, and D. A. Wiersma, Proc. Natl. Acad. Sci. (USA) 98, 1577 (2001).
[CrossRef]

Reintjes, J.

Schins, J. M.

M. Müller and J. M. Schins, J. Phys. Chem. B 106, 3715 (2002).
[CrossRef]

Squier, J.

M. Müller, J. Squier, C. A. de Lange, and G. J. Brakenhoff, J. Microsc. 197, 150 (2000).
[CrossRef]

van Haastert, P. J. M.

E. O. Potma, W. P. de Boeij, P. J. M. van Haastert, and D. A. Wiersma, Proc. Natl. Acad. Sci. (USA) 98, 1577 (2001).
[CrossRef]

Volkmer, A.

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

J. X. Cheng, A. Volkmer, L. D. Book, and X. S. Xie, J. Phys. Chem. B 105, 1277 (2001).
[CrossRef]

Wiersma, D. A.

E. O. Potma, W. P. de Boeij, P. J. M. van Haastert, and D. A. Wiersma, Proc. Natl. Acad. Sci. (USA) 98, 1577 (2001).
[CrossRef]

Xie, X. S.

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

J. X. Cheng, A. Volkmer, L. D. Book, and X. S. Xie, J. Phys. Chem. B 105, 1277 (2001).
[CrossRef]

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

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

Zumbusch, A.

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

J. Microsc.

M. Müller, J. Squier, C. A. de Lange, and G. J. Brakenhoff, J. Microsc. 197, 150 (2000).
[CrossRef]

J. Phys. Chem. B

M. Müller and J. M. Schins, J. Phys. Chem. B 106, 3715 (2002).
[CrossRef]

J. X. Cheng, A. Volkmer, L. D. Book, and X. S. Xie, J. Phys. Chem. B 105, 1277 (2001).
[CrossRef]

Opt. Lett.

Phys. Rev. Lett.

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

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

Proc. Natl. Acad. Sci. (USA)

E. O. Potma, W. P. de Boeij, P. J. M. van Haastert, and D. A. Wiersma, Proc. Natl. Acad. Sci. (USA) 98, 1577 (2001).
[CrossRef]

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Figures (3)

Fig. 1
Fig. 1

Measured (filled circles) and fitted (continuous curves) CARS spectra of (a) liquid absolute ethanol and (b) a 1-M urea solution. (c) Axial edge function obtained by scanning from the glass coverslip into ethanol. A fit of all spectra gives the resonant amplitude of the peak at -882 cm-1 (filled circles). The z resolution is measured from the smoothed first derivative (continuous). (d) Similarly, the x resolution was determined by measurement of the lateral edge function from the center of an 7.8µm glass microsphere into a 1-M urea solution stabilized by 2 wt. % agar by use of the -1005cm-1 peak.

Fig. 2
Fig. 2

(a) Three-dimensional (26×26×26 pixels; 13 µm×13 µm×15.6 µm) image of a DSPC MLV in water, showing xy, xz, and yz slices. Spectra were acquired at a rate of 20 ms/pixel and fitted with a three-peak line shape. In the fit, all parameters were optimized within constraints that avoid mutual compensation. Contrast is derived from the resonant amplitude at the -1129cm-1 peak. (b) A typical CARS spectrum of DSPC taken from the MLV image (filled circles) with its fit (continuous curve). (c) DSPC Raman spectrum in the corresponding frequency region.

Fig. 3
Fig. 3

(a) CARS spectra of a DSPC multilamellar vesicle in 0.5-M urea display three peaks assigned to DSPC, at Ω1=-1129, Ω2=-1102, and Ω3=-1062, and one peak for urea, Ω4=-1005 cm-1. By fitting all spectra to a four-peak line shape, we obtain chemical contrast by mapping both the DSPC R1 and the urea R4 fitted resonant amplitudes (in yellow and blue, respectively; scale bar indicates 2 µm). (a) The red lines indicate a line scan through the image, (b) displaying fit parameters R1 (continuous curve) and R4 (dashed curve). (c) The same line scan, now showing the measured intensity that comprises both the resonant and the nonresonant contributions at the DSPC peak position (continuous curve, -1129 cm-1) and the urea position (dashed curve -1005 cm-1). cps, cycles per second.

Equations (2)

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ICARSkNkχNR,k3+χR,k32,
χR3jRjδj-iΓj.

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