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Phase-Sensitive Vibrationally Resonant Sum-Frequency Generation Microscopy in Multiplex Configuration at 80 MHz Repetition Rate

Abstract

Vibrationally resonant sum-frequency generation (VR SFG) microscopy is an advanced imaging technique that can map out the intensity contrast of infrared and Raman active vibrational modes with micron to submicron lateral resolution. To broaden its applications and to obtain a molecular level of understanding, further technical advancement is needed to enable high-speed measurements of VR SFG microspectra at every pixel. In this study, we demonstrate a new VR SFG hyperspectral imaging platform combined with an ultrafast laser system operated at a repetition rate of 80 MHz. The multiplex configuration with broadband mid-infrared pulses makes it possible to measure a single microspectrum of CH/CH2 stretching modes in biological samples, such as starch granules and type I collagen tissue, with an exposure time of hundreds of milliseconds. Switching from the homodyne- to heterodyne-detected VR SFG hyperspectral imaging can be achieved by inserting a pair of optics into the beam path for local oscillator generation and delay time adjustment, which enables self-phase-stabilized spectral interferometry. We investigate the relationship between phase images of several different C-H modes and the relative orientation of collagen triple-helix in fibril bundles. The results show that the new multiplex VR SFG microscope operated at a high repetition rate is a powerful approach to probe the structural features and spatial arrangements of biological systems in detail.

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