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Stimulated Raman Scattering Tomography Enables Label‐Free Volumetric Deep Tissue Imaging

A novel z‐scan‐free stimulated Raman scattering tomography (SRST) is presented which is enabled by using optical beating technique (OBT) associated with Bessel beams to achieve deeper penetration for label‐free volumetric chemical imaging with subcellular resolution. In SRST, the pump beam is modulated with a spatial light modulator to convert to Bessel beam with optical beating, which is overlapped with the Bessel Stokes beam in the sample. By electronically varying the beating frequency, the depth‐resolved SRS signals about the volumetric tissue are encoded in the spatial frequency domain and thus, SRST can be rapidly retrieved by implementing inverse fast Fourier transform without a need of mechanical depth‐scan. It is demonstrated that SRST imaging using Bessel beams with self‐reconstructing properties provides at least twofold improvement in imaging depth in highly scattering polymer beads phantom as compared to conventional SRS microscopy. The capability of SRST for label‐free volumetric deeper imaging on a variety of imaging targets (e.g., Raman‐active crystals, plant cells, and biological tissue) is also proved. The generality of optical sectioning ability of Bessel beam‐OBT in SRST can be readily extended to practically any other nonlinear optical imaging modalities for deep tissue 3D imaging in biological and biomedical systems.