While super-resolution in fluorescence microscopy is well established, achieving resolution beyond the diffraction limit in Raman-based methods remains challenging. In the article "Label-free super-resolution stimulated Raman scattering imaging of biomedical specimens", the authors propose and demonstrate a super-resolution Stimulated Raman Scattering (SRS) imaging method compatible with opaque biological specimens. Traditional super-resolution fluorescence techniques rely on fluorescent labels, which can complicate biological imaging and introduce artifacts. In contrast, Raman-based methods utilize intrinsic vibrational properties of molecules for label-free imaging, providing rich chemical information.

Principle of blind-S³

The authors develop a novel single-pixel imaging technique combined with structured illumination, termed single-pixel blind structured illumination microscopy (blind-S3). This method avoids the need for multi-pixel detectors and complex amplification systems by using a single-pixel detector and a spatial light modulator (SLM) to generate structured pump light patterns. These patterns, combined with scanning Stokes light and blind-SIM algorithms, achieve high-resolution imaging.

Proof-of-concept of blind- S³ capabilities to image beyond the diffraction-limit.

The proposed method enhances lateral resolution to 217 nanometers, surpassing the diffraction limit, and maintains high axial resolution, making it suitable for thick and opaque biological samples. Validation with synthetic and biological samples demonstrates its capability to resolve fine structural details without fluorescent labels. The reduced phototoxicity due to lower excitation energy makes it promising for live-cell imaging.

Overall, this work represents a significant advancement in super-resolution Raman imaging, combining SRS and structured illumination to achieve high-resolution, label-free imaging of opaque biological specimens.