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Open Access Publications from the University of California

Polarization-sensitive sum-frequency generation microscopy of collagen fibers

  • Author(s): Han, Y
  • Hsu, J
  • Ge, NH
  • Potma, EO
  • et al.

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© 2015 American Chemical Society. Point-scanning sum-frequency generation (SFG) microscopy enables the generation of images of collagen I fibers in tissues by tuning into specific vibrational resonances of the polypeptide. It is shown that when collagen-rich tissues are visualized near the 2954 cm-1stretching vibration of methylene groups, the SFG image contrast is higher compared to the contrast seen in nonresonant second-harmonic generation (SHG) imaging. Polarization and spectrally resolved analysis of the SFG signal as a function of fiber orientation in the CH-stretching range of the vibrational spectrum enabled a comparative characterization of the achiral tensor elements of collagens second-order susceptibility. This analysis reveals that selected on-resonance tensor elements are enhanced over other elements, giving rise to a much stronger anisotropy ρ of the signal for SFG (ρ ≈ 15) compared to SHG (ρ ≈ 3). The improved anisotropy of the vibrationally resonant signal contributes to the higher contrast seen in the SFG tissue images.

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