INSPIRE: Single-beam probed complementary vibrational bioimaging | Science Advances

Molecular spectroscopy provides intrinsic contrast for in situ chemical imaging, linking the physiochemical properties of biomolecules to the functions of living systems. While stimulated Raman imaging has found successes in deciphering biological machinery, many vibrational modes are Raman inactive or weak, limiting the broader impact of the technique. It can potentially be mitigated by the spectral complementarity from infrared (IR) spectroscopy. However, the vastly different optical windows make it challenging to develop such a platform. Here, we introduce in situ pump-probe IR and Raman excitation (INSPIRE) microscopy, a nascent cross-modality spectroscopic imaging approach by encoding the ultrafast Raman and the IR photothermal relaxation into a single probe beam for simultaneous detection. INSPIRE inherits the merits of complementary modalities and demonstrates high-content molecular imaging of chemicals, cells, tissues, and organisms. Furthermore, INSPIRE applies to label-free and molecular tag imaging, offering possibilities for optical sensing and imaging in biomedicine and materials science.