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

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 realtime monitoring of dynamic processes, such as the diffusion of chemical compounds, in tissues. By tuning into the CH 2 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.nonlinear microscopy ͉ vibrational imaging ͉ back scattering

Virus trafficking – learning from single-virus tracking

“…It is therefore particularly exciting to track virus particles in live tissues and animals to see how viruses break host defence barriers to reach target cells for infection and how viral infection is transmitted from cell to cell. With the rapid development of in vivo imaging methods, such as non-linear optical microscopy 78,79 and endoscopy, this dream might be realized in the not-too-distant future.…”

Quantitative comparison of 3D third harmonic generation and fluorescence microscopy images

“…Like other multi-photon techniques [10][11][12][13][14][15][16][17][18], THG not only enables the recording of label-free images of unfixed 3D volumes of tissue [1][2][3][4][5][6][7][8][9][10] free from spatial distortion artifacts inherent to histopathology, but also potentially allows feedback on the nature of the tissue, i. e. whether it is healthy or tumorous, to the surgeon during surgery, as the relative speed of the imaging modalities approaches 'real' time, and no preparation steps of the tissue are required [5,6,[19][20][21][22][23]. THG was shown recently to yield label-free images of ex-vivo human tumor tissue of histopathological quality, in real-time [6].…”

Chemical Imaging with Frequency Modulation Coherent Anti-Stokes Raman Scattering Microscopy at the Vibrational Fingerprint Region

“…1 With its intrinsic vibrational contrast, great sensitivity, and three-dimensional sectioning ability, CARS microscopy has become an important labelfree chemical imaging tool in biology and material sciences. [1][2][3] Most CARS microscopy techniques demonstrated so far have employed two synchronized picosecond laser pulses. Since vibrational response of typical organic molecules lasts a few picoseconds, laser pulses with similar time duration are optimal in the consideration of both spectral resolution and signal sensitivity.…”