Label-Free Biomedical Imaging with High Sensitivity by Stimulated Raman Scattering Microscopy

TLDR

Label‑free chemical contrast is highly desirable in biomedical imaging, yet spontaneous Raman microscopy is limited by low sensitivity despite providing specific vibrational signatures. The authors present a three‑dimensional multiphoton vibrational imaging technique based on stimulated Raman scattering (SRS). They demonstrate the technique by imaging lipid distributions in living cells, brain and skin tissues, and monitoring drug delivery through the epidermis. SRS imaging achieves markedly higher sensitivity than spontaneous Raman microscopy through high‑frequency phase‑sensitive detection and provides background‑free, readily interpretable chemical contrast compared to previous coherent Raman methods.

Abstract

Label-free chemical contrast is highly desirable in biomedical imaging. Spontaneous Raman microscopy provides specific vibrational signatures of chemical bonds, but is often hindered by low sensitivity. Here we report a three-dimensional multiphoton vibrational imaging technique based on stimulated Raman scattering (SRS). The sensitivity of SRS imaging is significantly greater than that of spontaneous Raman microscopy, which is achieved by implementing high-frequency (megahertz) phase-sensitive detection. SRS microscopy has a major advantage over previous coherent Raman techniques in that it offers background-free and readily interpretable chemical contrast. We show a variety of biomedical applications, such as differentiating distributions of omega-3 fatty acids and saturated lipids in living cells, imaging of brain and skin tissues based on intrinsic lipid contrast, and monitoring drug delivery through the epidermis.