Chemical imaging of tissue in vivo with video-rate coherent anti-Stokes Raman scattering microscopy

doi:10.1073/pnas.0508282102

Chemical imaging of tissue in vivo with video-rate coherent anti-Stokes Raman scattering microscopy

^*Department of Chemistry and Chemical Biology, Harvard University, 12 Oxford Street, Cambridge, MA 02138; and ^‡Wellman Center for Photomedicine, Massachusetts General Hospital, 55 Fruit Street, Boston, MA 02114

Communicated by William Klemperer, Harvard University, Cambridge, MA, September 22, 2005 (received for review August 18, 2005)

Abstract

Imaging living organisms with molecular selectivity typically requires the introduction of specific labels. Many applications in biology and medicine, however, would significantly benefit from a noninvasive imaging technique that circumvents such exogenous probes. In vivo microscopy based on vibrational spectroscopic contrast offers a unique approach for visualizing tissue architecture with molecular specificity. We have developed a sensitive technique for vibrational imaging of tissues by combining coherent anti-Stokes Raman scattering (CARS) with video-rate microscopy. Backscattering of the intense forward-propagating CARS radiation in tissue gives rise to a strong epi-CARS signal that makes in vivo imaging possible. This substantially large signal allows for real-time monitoring of dynamic processes, such as the diffusion of chemical compounds, in tissues. By tuning into the CH₂ stretching vibrational band, we demonstrate CARS imaging and spectroscopy of lipid-rich tissue structures in the skin of a live mouse, including sebaceous glands, corneocytes, and adipocytes, with unprecedented contrast at subcellular resolution.

Footnotes

↵ § To whom correspondence may be addressed. E-mail: lin{at}helix.mgh.harvard.edu or xie{at}chemistry.harvard.edu.
↵ † Present address: Department of Chemistry, University of California, Irvine, CA 92697.
Author contributions: C.L.E., E.O.P., C.P.L., and X.S.X. designed research; C.L.E., E.O.P., and M.P. performed research; C.L.E., E.O.P., D.C., and X.S.X. analyzed data; and C.L.E., E.O.P., and X.S.X. wrote the paper.
Conflict of interest statement: No conflicts declared.
Abbreviations: CARS, coherent anti-Stokes Raman scattering; DiD, C₆₇H₁₀₃CIN₂O₃S.