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

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.