OSA's Digital Library

Optics Express

Optics Express

  • Editor: C. Martijn de Sterke
  • Vol. 17, Iss. 21 — Oct. 12, 2009
  • pp: 18419–18432

Quantitative myelin imaging with coherent anti-Stokes Raman scattering microscopy: alleviating the excitation polarization dependence with circularly polarized laser beams

E. Bélanger, S. Bégin, S. Laffray, Y. De Koninck, R. Vallée, and D. Côté  »View Author Affiliations

Optics Express, Vol. 17, Issue 21, pp. 18419-18432 (2009)

View Full Text Article

Enhanced HTML    Acrobat PDF (1998 KB)

Browse Journals / Lookup Meetings

Browse by Journal and Year


Lookup Conference Papers

Close Browse Journals / Lookup Meetings

Article Tools



The use of coherent anti-Stokes Raman scattering microscopy tuned to the lipid vibration for quantitative myelin imaging suffers from the excitation polarization dependence of this third-order nonlinear optical effect. The contrast obtained depends on the orientation of the myelin membrane, which in turn affects the morphometric parameters that can be extracted with image analysis. We show how circularly polarized laser beams can be used to avoid this complication, leading to images free of excitation polarization dependence. The technique promises to be optimal for in vivo imaging and the resulting images can be used for coherent anti-Stokes Raman scattering optical histology on native state tissue.

© 2009 Optical Society of America

OCIS Codes
(170.3880) Medical optics and biotechnology : Medical and biological imaging
(180.6900) Microscopy : Three-dimensional microscopy
(300.6230) Spectroscopy : Spectroscopy, coherent anti-Stokes Raman scattering
(180.4315) Microscopy : Nonlinear microscopy

ToC Category:
Medical Optics and Biotechnology

Original Manuscript: July 8, 2009
Revised Manuscript: September 1, 2009
Manuscript Accepted: September 12, 2009
Published: September 28, 2009

Virtual Issues
Vol. 4, Iss. 12 Virtual Journal for Biomedical Optics

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, 18419-18432 (2009)

Sort:  Author  |  Year  |  Journal  |  Reset  


  1. J. B. Pawley, ed., Handbook of biological confocal microscopy, 3rd ed. (Springer, 1995).
  2. N. C. Shaner, R. E. Campbell, P. A. Steinbach, B. N. G. Giepmans, A. E. Palmer, and R. Y. Tsien, "Improved monomeric red, orange and yellow fluorescent proteins derived from Discosoma sp. red fluorescent protein," Nat. Biotechnol. 22, 1567-1572 (2004). [CrossRef] [PubMed]
  3. F. Helmchen and W. Denk, "Deep tissue two-photon microscopy," Nat. Methods 2, 932-940 (2005). [CrossRef] [PubMed]
  4. N. C. Shaner, P. A. Steinbach, and R. Y. Tsien, "A guide to choosing fluorescent proteins," Nat. Methods 2, 905-909 (2005). [CrossRef] [PubMed]
  5. B. N. G. Giepmans, S. R. Adams, M. H. Ellisman, and R. Y. Tsien, "The fluorescent toolbox for assessing protein location and function," Science 312, 217-224 (2006). [CrossRef] [PubMed]
  6. I. Veilleux, J. A. Spencer, D. P. Biss, D. Côté, and C. P. Lin, "In vivo cell tracking with video rate multimodality laser scanning microscopy," IEEE J. Sel. Top. Quantum Electron. 14, 10-18 (2008). [CrossRef]
  7. W. R. Zipfel, R. M. Williams, R. Christie, A. Y. Nikitin, B. T. Hyman, and W. W. Webb, "Live tissue intrinsic emission microscopy using multiphoton-excited native fluorescence and second harmonic generation." Proc. Natl. Acad. Sci. U.S.A. 100, 7075-7080 (2003). [CrossRef] [PubMed]
  8. P. J. Campagnola and L. M. Loew, "Second-harmonic imaging microscopy for visualizing biomolecular arrays in cells, tissues and organisms," Nat. Biotechnol. 21, 1356-1360 (2003). [CrossRef] [PubMed]
  9. E. B. Hanlon, R. Manoharan, T. W. Koo, K. E. Shafer, J. T. Motz, M. Fitzmaurice, J. R. Kramer, I. Itzkan, R. R. Dasari, and M. S. Feld, "Prospects for in vivo Raman spectroscopy," Phys. Med. Biol. 45, R1-R59 (2000). [CrossRef] [PubMed]
  10. J.-X. Cheng and X. Xie, "Coherent Anti-Stokes Raman Scattering Microscopy: Instrumentation, Theory, and Applications," J. Phys. Chem. B 108, 827-840 (2004). [CrossRef]
  11. C. Evans and X. S. Xie, "Coherent Anti-Stokes Raman Scattering Microscopy: Chemical Imaging for Biology and Medicine," Annu. Rev. Anal. Chem. 1, 883-909 (2008). [CrossRef]
  12. 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, 16807-16812 (2005). [CrossRef] [PubMed]
  13. F. P. Henry, D. Côté, M. A. Randolph, E. A. Z. Rust, R. W. Redmond, I. E. Kochevar, C. P. Lin, and J. M. Winograd, "Real-time in vivo assessment of the nerve microenvironment with coherent anti-Stokes Raman scattering microscopy," Plast. Reconstr. Surg. 123(2S), 123S-130S (2009). [CrossRef]
  14. 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, 581-591 (2005). [CrossRef] [PubMed]
  15. E. O. Potma and X. S. Xie, "Detection of single lipid bilayers with coherent anti-Stokes Raman scattering (CARS) microscopy," J. Raman Spectrosc. 34, 642-650 (2003). [CrossRef]
  16. R. L. Friede, "Computer editing of morphometric data on nerve fibers. An improved computer program," Acta Neuropathol. 72, 74-81 (1986). [CrossRef] [PubMed]
  17. W. Beuche and R. L. Friede, "A quantitative assessment of myelin sheaths in the peripheral nerves of dystrophic, quaking, and trembler mutants." Acta Neuropathol. 66, 29-36 (1985). [CrossRef] [PubMed]
  18. R. L. Friede and W. Beuche, "A new approach toward analyzing peripheral nerve fiber populations. I. Variance in sheath thickness corresponds to different geometric proportions of the internodes." J. Neuropathol. Exp. Neurol. 44, 60-72 (1985). [CrossRef] [PubMed]
  19. C. Hildebrand and R. Hahn, "Relation between myelin sheath thickness and axon size in spinal cord white matter of some vertebrate species," J. Neurol. Sci. 38, 421-434 (1978). [CrossRef] [PubMed]
  20. T. L. Mazely and W. M. H.III, "Third-order susceptibility tensors of partially ordered systems," J. Chem. Phys. 87, 1962-1966 (1987). [CrossRef]
  21. C. C. Shang and H. Hsu, "The spatial symmetrical forms of third-order nonlinear susceptibility," IEEE J. Quant. Electron. 23, 177-179 (1987). [CrossRef]
  22. D. A. Kleinman, "Nonlinear dielectric polarization in optical media," Phys. Rev. 126, 1977-1979 (1962). [CrossRef]
  23. D. Chemla, R. F. Begley, and R. Byer, "Experimental and theoretical studies of third-harmonic generation in the chalcopyrite CdGeAs2," IEEE J. Quantum Electron. 10, 71- 81 (1974). [CrossRef]
  24. S.-W. Chu, S.-Y. Chen, G.-W. Chern, T.-H. Tsai, Y.-C. Chen, B.-L. Lin, and C.-K. Sun, "Studies of chi(2)/chi(3) tensors in submicron-scaled bio-tissues by polarization harmonics optical microscopy," Biophys. J. 86, 3914-3922 (2004). [CrossRef] [PubMed]
  25. C.-K. Chou, W.-L. Chen, P. T. Fwu, S.-J. Lin, H.-S. Lee, and C.-Y. Dong, "Polarization ellipticity compensation in polarization second-harmonic generation microscopy without specimen rotation," J. Biomed. Opt. 13, 014005 (2008). [CrossRef] [PubMed]
  26. Y. Fu, T. B. Huff, H.-W. Wang, H. Wang, and J.-X. Cheng, "Ex vivo and in vivo imaging of myelin fibers in mouse brain by coherent anti-Stokes Raman scattering microscopy," Opt. Express 16, 19396-19409 (2008). [CrossRef] [PubMed]
  27. P. Stoller, K. M. Reiser, P. M. Celliers, and A. M. Rubenchik, "Polarization-modulated second harmonic generation in collagen," Biophys. J. 82, 3330-3342 (2002). [CrossRef] [PubMed]
  28. W. Beuche and R. L. Friede, "A new approach toward analyzing peripheral nerve fiber populations. II. Foreshortening of regenerated internodes corresponds to reduced sheath thickness." J. Neuropathol. Exp. Neurol. 44, 73-84 (1985). [CrossRef] [PubMed]
  29. R. S. Smith and Z. J. Koles, "Myelinated nerve fibers: computed effect of myelin thickness on conduction velocity," Am. J. Physiol. 219, 1256-1258 (1970). [PubMed]
  30. I. Micu, A. Ridsdale, L. Zhang, J. Woulfe, J. McClintock, C. A. Brantner, S. B. Andrews, and P. K. Stys, "Realtime measurement of free Ca2+ changes in CNS myelin by two-photon microscopy," Nat. Med. 13, 874-879 (2007). [CrossRef] [PubMed]

Cited By

Alert me when this paper is cited

OSA is able to provide readers links to articles that cite this paper by participating in CrossRef's Cited-By Linking service. CrossRef includes content from more than 3000 publishers and societies. In addition to listing OSA journal articles that cite this paper, citing articles from other participating publishers will also be listed.

Supplementary Material

» Media 1: MOV (1263 KB)     

« Previous Article  |  Next Article »

OSA is a member of CrossRef.

CrossCheck Deposited