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.

Automated summary

A novel z-scan-free stimulated Raman scattering tomography (SRST) utilizing optical beating technique (OBT) and Bessel beams enables deeper penetration for label-free volumetric chemical imaging with subcellular resolution. The technology encodes depth-resolved SRS signals in the spatial frequency domain, rapidly retrieving volumetric tissue information without mechanical depth-scanning, and demonstrates improved imaging depth in highly scattering samples compared to conventional SRS microscopy, with potential applications in various biological and biomedical systems.