Rotating-polarization CARS microscopy: combining chemical and molecular orientation sensitivity

Giuseppe de Vito; Angelo Bifone; Vincenzo Piazza

doi:10.1364/OE.20.029369

Rotating-polarization CARS microscopy: combining chemical and molecular orientation sensitivity

Giuseppe de Vito, Angelo Bifone, and Vincenzo Piazza

Optics Express
Vol. 20,
Issue 28,
pp. 29369-29377
(2012)
•https://doi.org/10.1364/OE.20.029369

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Giuseppe de Vito, Angelo Bifone, and Vincenzo Piazza, "Rotating-polarization CARS microscopy: combining chemical and molecular orientation sensitivity," Opt. Express 20, 29369-29377 (2012)

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Related Topics
Table of Contents Category
- Microscopy
Optics & Photonics Topics
?

The topics in this list come from the OSA Optics and Photonics Topics applied to this article.
Previously assigned OCIS codes
- Three-dimensional microscopy (180.6900)
- Nonlinear microscopy (180.4315)
- Raman microscopy (180.5655)
- Nonlinear optics, four-wave mixing (190.4380)
- Spectroscopy, coherent anti-Stokes Raman scattering (300.6230)

About this Article
History
- Original Manuscript: October 26, 2012
- Revised Manuscript: November 30, 2012
- Manuscript Accepted: November 30, 2012
- Published: December 18, 2012
Virtual Issues
Virtual Journal for Biomedical Optics Vol. 8, Iss. 1

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Open Access

Abstract

Coherent Anti-Stokes Raman Spectroscopy (CARS) is a non-linear process in which the energy difference of a pair of incoming photons matches the energy of the vibrational mode of a molecular bond of interest. This phonon population is coherently probed by a third photon and anti-Stokes radiation is emitted. Here a novel approach to CARS microscopy is presented yielding the intensity of the anti-Stokes emission, the directionality the molecular bonds of interest, and their average orientation. Myelinated axons in fixed mouse-brain slices have been imaged by RP-CARS. We were able to detect the local average direction of the acylic chains of membrane phospholipids and their spatial anisotropy. This novel method may impact the study of healthy brain circuitry as well as demyelinating diseases or other pathological states associated with altered neural connectivity.

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References

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Year
|
Author
|
Publication

P. Maker and R. Terhune, “Study of optical effects due to an induced polarization third order in the electric field strength,” Phys. Rev. 137(3A), A801–A818 (1965).
[Crossref]
J. X. Cheng and X. S. Xie, “Coherent anti-Stokes Raman scattering microscopy: instrumentation, theory, and applications,” J. Phys. Chem. B 108(3), 827–840 (2004).
[Crossref]
C. L. Evans, E. O. Potma, M. Puoris’haag, D. Côté, C. P. Lin, and X. S. Xie, “Chemical imaging of tissue in vivo with video-rate coherent anti-Stokes Raman scattering microscopy,” Proc. Natl. Acad. Sci. U.S.A. 102(46), 16807–16812 (2005).
[Crossref] [PubMed]
C. L. Evans and X. S. Xie, “Coherent anti-Stokes Raman scattering microscopy: chemical imaging for biology and medicine,” Annu Rev Anal Chem (Palo Alto Calif) 1(1), 883–909 (2008).
[Crossref] [PubMed]
J. X. Cheng, S. Pautot, D. A. Weitz, and X. S. Xie, “Ordering of water molecules between phospholipid bilayers visualized by coherent anti-Stokes Raman scattering microscopy,” Proc. Natl. Acad. Sci. U.S.A. 100(17), 9826–9830 (2003).
[Crossref] [PubMed]
A. V. Kachynski, A. N. Kuzmin, P. N. Prasad, and I. I. Smalyukh, “Coherent anti-Stokes Raman scattering polarized microscopy of three-dimensional director structures in liquid crystals,” Appl. Phys. Lett. 91(15), 151905 (2007).
[Crossref]
F. Munhoz, H. Rigneault, and S. Brasselet, “Polarization-resolved four-wave mixing microscopy for structural imaging in thick tissues,” J. Opt. Soc. Am. B 29(6), 1541–1550 (2012).
[Crossref]
J. X. Cheng, L. D. Book, and X. S. Xie, “Polarization coherent anti-Stokes Raman scattering microscopy,” Opt. Lett. 26(17), 1341–1343 (2001).
[Crossref] [PubMed]
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(9), 1505–1507 (2002).
[Crossref]
Y. Fu, T. B. Huff, H.-W. Wang, J.-X. Cheng, and H. Wang, “Ex vivo and in vivo imaging of myelin fibers in mouse brain by coherent anti-Stokes Raman scattering microscopy,” Opt. Express 16(24), 19396–19409 (2008).
[Crossref] [PubMed]
E. Bélanger, S. Bégin, S. Laffray, Y. De Koninck, R. Vallée, and D. Côté, “Quantitative myelin imaging with coherent anti-Stokes Raman scattering microscopy: alleviating the excitation polarization dependence with circularly polarized laser beams,” Opt. Express 17(21), 18419–18432 (2009).
[Crossref] [PubMed]
H. Wang, Y. Fu, P. Zickmund, R. Shi, and J. X. Cheng, “Coherent anti-Stokes Raman scattering imaging of axonal myelin in live spinal tissues,” Biophys. J. 89(1), 581–591 (2005).
[Crossref] [PubMed]
C. Shang and H. Hsu, “The spatial symmetric forms of third-order nonlinear susceptibility,” IEEE J. Quantum Electron. 23(2), 177–179 (1987).
[Crossref]
R. W. Hellwarth, “Third-order optical susceptibilities of liquids and solids,” Prog. Quantum Electron. 5, 1–68 (1979).
[Crossref]
M. Axer, K. Amunts, D. Grässel, C. Palm, J. Dammers, H. Axer, U. Pietrzyk, and K. Zilles, “A novel approach to the human connectome: ultra-high resolution mapping of fiber tracts in the brain,” Neuroimage 54(2), 1091–1101 (2011).
[Crossref] [PubMed]
M. D. Budde and J. A. Frank, “Examining brain microstructure using structure tensor analysis of histological sections,” Neuroimage 63(1), 1–10 (2012).
[Crossref] [PubMed]

2012 (2)

F. Munhoz, H. Rigneault, and S. Brasselet, “Polarization-resolved four-wave mixing microscopy for structural imaging in thick tissues,” J. Opt. Soc. Am. B 29(6), 1541–1550 (2012).
[Crossref]

M. D. Budde and J. A. Frank, “Examining brain microstructure using structure tensor analysis of histological sections,” Neuroimage 63(1), 1–10 (2012).
[Crossref] [PubMed]

2011 (1)

M. Axer, K. Amunts, D. Grässel, C. Palm, J. Dammers, H. Axer, U. Pietrzyk, and K. Zilles, “A novel approach to the human connectome: ultra-high resolution mapping of fiber tracts in the brain,” Neuroimage 54(2), 1091–1101 (2011).
[Crossref] [PubMed]

2009 (1)

E. Bélanger, S. Bégin, S. Laffray, Y. De Koninck, R. Vallée, and D. Côté, “Quantitative myelin imaging with coherent anti-Stokes Raman scattering microscopy: alleviating the excitation polarization dependence with circularly polarized laser beams,” Opt. Express 17(21), 18419–18432 (2009).
[Crossref] [PubMed]

2008 (2)

Y. Fu, T. B. Huff, H.-W. Wang, J.-X. Cheng, and H. Wang, “Ex vivo and in vivo imaging of myelin fibers in mouse brain by coherent anti-Stokes Raman scattering microscopy,” Opt. Express 16(24), 19396–19409 (2008).
[Crossref] [PubMed]

C. L. Evans and X. S. Xie, “Coherent anti-Stokes Raman scattering microscopy: chemical imaging for biology and medicine,” Annu Rev Anal Chem (Palo Alto Calif) 1(1), 883–909 (2008).
[Crossref] [PubMed]

2007 (1)

A. V. Kachynski, A. N. Kuzmin, P. N. Prasad, and I. I. Smalyukh, “Coherent anti-Stokes Raman scattering polarized microscopy of three-dimensional director structures in liquid crystals,” Appl. Phys. Lett. 91(15), 151905 (2007).
[Crossref]

2005 (2)

C. L. Evans, E. O. Potma, M. Puoris’haag, D. Côté, C. P. Lin, and X. S. Xie, “Chemical imaging of tissue in vivo with video-rate coherent anti-Stokes Raman scattering microscopy,” Proc. Natl. Acad. Sci. U.S.A. 102(46), 16807–16812 (2005).
[Crossref] [PubMed]

H. Wang, Y. Fu, P. Zickmund, R. Shi, and J. X. Cheng, “Coherent anti-Stokes Raman scattering imaging of axonal myelin in live spinal tissues,” Biophys. J. 89(1), 581–591 (2005).
[Crossref] [PubMed]

2004 (1)

J. X. Cheng and X. S. Xie, “Coherent anti-Stokes Raman scattering microscopy: instrumentation, theory, and applications,” J. Phys. Chem. B 108(3), 827–840 (2004).
[Crossref]

2003 (1)

J. X. Cheng, S. Pautot, D. A. Weitz, and X. S. Xie, “Ordering of water molecules between phospholipid bilayers visualized by coherent anti-Stokes Raman scattering microscopy,” Proc. Natl. Acad. Sci. U.S.A. 100(17), 9826–9830 (2003).
[Crossref] [PubMed]

2002 (1)

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(9), 1505–1507 (2002).
[Crossref]

2001 (1)

J. X. Cheng, L. D. Book, and X. S. Xie, “Polarization coherent anti-Stokes Raman scattering microscopy,” Opt. Lett. 26(17), 1341–1343 (2001).
[Crossref] [PubMed]

1987 (1)

C. Shang and H. Hsu, “The spatial symmetric forms of third-order nonlinear susceptibility,” IEEE J. Quantum Electron. 23(2), 177–179 (1987).
[Crossref]

1979 (1)

R. W. Hellwarth, “Third-order optical susceptibilities of liquids and solids,” Prog. Quantum Electron. 5, 1–68 (1979).
[Crossref]

1965 (1)

P. Maker and R. Terhune, “Study of optical effects due to an induced polarization third order in the electric field strength,” Phys. Rev. 137(3A), A801–A818 (1965).
[Crossref]

Amunts, K.

Axer, H.

Axer, M.

Bégin, S.

Bélanger, E.

Book, L. D.

J. X. Cheng, L. D. Book, and X. S. Xie, “Polarization coherent anti-Stokes Raman scattering microscopy,” Opt. Lett. 26(17), 1341–1343 (2001).
[Crossref] [PubMed]

Brasselet, S.

F. Munhoz, H. Rigneault, and S. Brasselet, “Polarization-resolved four-wave mixing microscopy for structural imaging in thick tissues,” J. Opt. Soc. Am. B 29(6), 1541–1550 (2012).
[Crossref]

Budde, M. D.

M. D. Budde and J. A. Frank, “Examining brain microstructure using structure tensor analysis of histological sections,” Neuroimage 63(1), 1–10 (2012).
[Crossref] [PubMed]

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(3), 827–840 (2004).
[Crossref]

J. X. Cheng, L. D. Book, and X. S. Xie, “Polarization coherent anti-Stokes Raman scattering microscopy,” Opt. Lett. 26(17), 1341–1343 (2001).
[Crossref] [PubMed]

Cheng, J.-X.

Côté, D.

Dammers, J.

De Koninck, Y.

Evans, C. L.

Frank, J. A.

M. D. Budde and J. A. Frank, “Examining brain microstructure using structure tensor analysis of histological sections,” Neuroimage 63(1), 1–10 (2012).
[Crossref] [PubMed]

Fu, Y.

Grässel, D.

Hellwarth, R. W.

R. W. Hellwarth, “Third-order optical susceptibilities of liquids and solids,” Prog. Quantum Electron. 5, 1–68 (1979).
[Crossref]

Hsu, H.

C. Shang and H. Hsu, “The spatial symmetric forms of third-order nonlinear susceptibility,” IEEE J. Quantum Electron. 23(2), 177–179 (1987).
[Crossref]

Huff, T. B.

Kachynski, A. V.

Kuzmin, A. N.

Laffray, S.

Lin, C. P.

Maker, P.

P. Maker and R. Terhune, “Study of optical effects due to an induced polarization third order in the electric field strength,” Phys. Rev. 137(3A), A801–A818 (1965).
[Crossref]

Munhoz, F.

F. Munhoz, H. Rigneault, and S. Brasselet, “Polarization-resolved four-wave mixing microscopy for structural imaging in thick tissues,” J. Opt. Soc. Am. B 29(6), 1541–1550 (2012).
[Crossref]

Palm, C.

Pautot, S.

Pietrzyk, U.

Potma, E. O.

Prasad, P. N.

Puoris’haag, M.

Rigneault, H.

F. Munhoz, H. Rigneault, and S. Brasselet, “Polarization-resolved four-wave mixing microscopy for structural imaging in thick tissues,” J. Opt. Soc. Am. B 29(6), 1541–1550 (2012).
[Crossref]

Shang, C.

C. Shang and H. Hsu, “The spatial symmetric forms of third-order nonlinear susceptibility,” IEEE J. Quantum Electron. 23(2), 177–179 (1987).
[Crossref]

Shi, R.

Smalyukh, I. I.

Terhune, R.

P. Maker and R. Terhune, “Study of optical effects due to an induced polarization third order in the electric field strength,” Phys. Rev. 137(3A), A801–A818 (1965).
[Crossref]

Vallée, R.

Volkmer, A.

Wang, H.

Wang, H.-W.

Weitz, D. A.

Xie, X. S.

J. X. Cheng and X. S. Xie, “Coherent anti-Stokes Raman scattering microscopy: instrumentation, theory, and applications,” J. Phys. Chem. B 108(3), 827–840 (2004).
[Crossref]

J. X. Cheng, L. D. Book, and X. S. Xie, “Polarization coherent anti-Stokes Raman scattering microscopy,” Opt. Lett. 26(17), 1341–1343 (2001).
[Crossref] [PubMed]

Zickmund, P.

Zilles, K.

Annu Rev Anal Chem (Palo Alto Calif) (1)

Appl. Phys. Lett. (2)

Biophys. J. (1)

IEEE J. Quantum Electron. (1)

C. Shang and H. Hsu, “The spatial symmetric forms of third-order nonlinear susceptibility,” IEEE J. Quantum Electron. 23(2), 177–179 (1987).
[Crossref]

J. Opt. Soc. Am. B (1)

F. Munhoz, H. Rigneault, and S. Brasselet, “Polarization-resolved four-wave mixing microscopy for structural imaging in thick tissues,” J. Opt. Soc. Am. B 29(6), 1541–1550 (2012).
[Crossref]

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(3), 827–840 (2004).
[Crossref]

Neuroimage (2)

M. D. Budde and J. A. Frank, “Examining brain microstructure using structure tensor analysis of histological sections,” Neuroimage 63(1), 1–10 (2012).
[Crossref] [PubMed]

Opt. Express (2)

Opt. Lett. (1)

J. X. Cheng, L. D. Book, and X. S. Xie, “Polarization coherent anti-Stokes Raman scattering microscopy,” Opt. Lett. 26(17), 1341–1343 (2001).
[Crossref] [PubMed]

Phys. Rev. (1)

P. Maker and R. Terhune, “Study of optical effects due to an induced polarization third order in the electric field strength,” Phys. Rev. 137(3A), A801–A818 (1965).
[Crossref]

Proc. Natl. Acad. Sci. U.S.A. (2)

Prog. Quantum Electron. (1)

R. W. Hellwarth, “Third-order optical susceptibilities of liquids and solids,” Prog. Quantum Electron. 5, 1–68 (1979).
[Crossref]

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Fig. 1

Schematic representation of the RP-CARS setup. The 800-nm pulses from the laser (fs Laser), shown as red lines, are split by a beam splitter (BS) and routed to the supercontinuum generator (SCG) and, through two 3-nm bandpass filters (3-nm BP) centered at 800 nm, to the rotating λ/2 retarder (R-λ/2). The linearly-polarized broadband radiation (green lines) is delayed by a delay line (DL), transformed into circularly polarized light by a λ/4 retarder (λ/4), and recombined with the 800-nm radiation by means of a dichroic mirror (D). A second λ/4 retarder in the pump and probe path allows compensating for polarization distortions caused by D. The two are then routed to the high-numerical-aperture lens (Obj) through a pair of galvo-scanning mirrors, a scan lens and a tube lens. CARS signal is collected from the sample (S) by a condenser lens (Cond), band-pass filtered (BP), and routed to a photomultiplier tube (PMT). M0 to M2 are silver-coated mirrors. The output of the PMT is measured by phase-sensitive techniques by means of a lock-in amplifier. The reference phase and frequency for the lock-in amplifier is generated by a Hall sensor in close proximity to the rotor of the brushless motor that rotates R-λ/2.

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Fig. 2

Polarization-dependent artifact in a CARS image of a single nerve axon in transverse section (CH and CH₂ bonds). (a) The polarization-dependent artifact is evident when linearly-polarized light is used. In this case the CH and CH₂ chemical bonds aligned with the polarization plane of the incident light (vertical in the image) generate a stronger CARS signal. (b) Same as a) but using a perpendicular polarization plane of the incident light (horizontal in the image). The pump and probe beam and the Stokes beam power was respectively 30 mW and 12 mW. Scale bar: 0.5 μm.

Download Full Size | PPT Slide | PDF

Fig. 3

RP-CARS imaging of the same single nerve axon as in Fig. 2 (CH₂ bonds, transverse section). (a) Image constructed using $A_{d c} .$ Compared to the images in Fig. 2 it is free from the polarization artifacts. (b) Image constructed using $A_{2 ω}$ (bright pixels indicate that the signal value varies strongly as a function of the light polarization orientation). (c) Image of the phase of the signal collected from the same region (the colors indicate different light-polarization orientations). (d) RP-image constructed using the image in b) as brightness value and the one in c) as hue value. The myelin sheaths change color smoothly, covering all the possible orientations. The power levels were the same as in Fig. 2. The scale bar length is 0.5 μm.

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Fig. 4

Coronal section of the mouse brain anterior commissure imaged at different magnifications. This multi-scale acquisition shows the degree of local asymmetry in the direction of the acylic chains and their average spatial orientation (the color-coding scheme is described in the text). The white rectangles show the region magnified in the successive image of the series (indicated by an arrow). The colored lines in the top-left panel depict the color-to-orientation mapping. The last three images (*,§ and #) refer respectively to the three white rectangles in the preceding image. The pump and probe beam and the Stokes beam power was respectively 50 mW and 10 mW.

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Fig. 5

$A_{d c}$ and $A_{2 ω}$ signals of mouse brain slices (CH₂ bonds). (a) Conventional CARS image (linar polarizations) from a 100-μm-thick brain slice (bregma: 0.4 mm). (b) $A_{d c}$ signal from a 150-μm-thick brain slice (bregma: −0.1 mm). Note that in (a) white matter (e.g., corpus callosum or anterior commissure) appears bright, whereas in (b) it appears dark. This effect is due to the different slice thickness: the thicker the slice, the more its dense white matter diffuses the CARS signal. (c) Image from the same brain slice shown in (b), but constructed using the $A_{2 ω}$ signal (divided by the $A_{d c}$ signal): in this case the white matter appears bright as in (a). The medial longitudinal fissure, filled with aqueous solution, appears artifactually bright due to its low $A_{d c}$ signal. cc: corpus callosum. ac: anterior commissure, its shape appears different in (a) with respect to (b) and (c) because of the diverse positioning of the coronal slice (distance to bregma). ml: medial longitudinal fissure. The pump and probe beam and the Stokes beam power was respectively 30 mW and 18 mW. Scale bar: 0.25 mm.

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Equations (4)

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I_{C A R S} (t) = I_{p}^{2} I_{S} [A_{d c} + A_{2 ω} e^{(2 i ω t + i φ)} + A_{4 ω} e^{(4 i ω t + i φ)}],

\begin{array}{l} A_{d c} = 3 c_{11}^{2} + 2 c_{11} c_{16} + 14 c_{16}^{2} + 2 c_{33} c_{16} + 3 c_{33}^{2} \\ A_{2 ω} = 4 (c_{11}^{2} - c_{33}^{2}) \\ A_{4 w} = c_{11}^{2} - 2 c_{11} c_{16} - 6 c_{16}^{2} - 2 c_{33} c_{16} + c_{33}^{2} . \end{array}

χ_{i j k l} \propto δ_{i j} δ_{k l} + δ_{i k} δ_{j l} + δ_{i l} δ_{j k} .

I_{C A R S} (t) = I_{p}^{2} I_{S} [B_{d c} + B_{2 ω} e^{(2 i ω t + i φ)}],

Abstract

References

2012 (2)

2011 (1)

2009 (1)

2008 (2)

2007 (1)

2005 (2)

2004 (1)

2003 (1)

2002 (1)

2001 (1)

1987 (1)

1979 (1)

1965 (1)

Amunts, K.

Axer, H.

Axer, M.

Bégin, S.

Bélanger, E.

Book, L. D.

Brasselet, S.

Budde, M. D.

Cheng, J. X.

Cheng, J.-X.

Côté, D.

Dammers, J.

De Koninck, Y.

Evans, C. L.

Frank, J. A.

Fu, Y.

Grässel, D.

Hellwarth, R. W.

Hsu, H.

Huff, T. B.

Kachynski, A. V.

Kuzmin, A. N.

Laffray, S.

Lin, C. P.

Maker, P.

Munhoz, F.

Palm, C.

Pautot, S.

Pietrzyk, U.

Potma, E. O.

Prasad, P. N.

Puoris’haag, M.

Rigneault, H.

Shang, C.

Shi, R.

Smalyukh, I. I.

Terhune, R.

Vallée, R.

Volkmer, A.

Wang, H.

Wang, H.-W.

Weitz, D. A.

Xie, X. S.

Zickmund, P.

Zilles, K.

Annu Rev Anal Chem (Palo Alto Calif) (1)

Appl. Phys. Lett. (2)

Biophys. J. (1)

IEEE J. Quantum Electron. (1)

J. Opt. Soc. Am. B (1)

J. Phys. Chem. B (1)

Neuroimage (2)

Opt. Express (2)

Opt. Lett. (1)

Phys. Rev. (1)

Proc. Natl. Acad. Sci. U.S.A. (2)

Prog. Quantum Electron. (1)

Cited By

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