Project Abstract Optical microscopy has revolutionized our understanding of biological systems with fine subcellular resolution, fast temporal dynamics and superb target specificity. One of the recent technical breakthroughs in bioimaging is the super-resolution fluorescence microscopy, which breaks the diffraction limit and allows unprecedented optical investigation at the nanoscopic level. However, super-resolution fluorescence microscopy has been fundamentally limited in two aspects. First, it is not suitable for interrogating structures mainly composed of small biomolecules, due to the required large and perturbative fluorophore labeling. Second, it is ineffective for investigating small complex structures involving multiple components because the broad and featureless fluorescent spectra limits the colors that could be simultaneously imaged typically to only 2-5. Therefore, how to noninvasively super-resolve these two types of cellular structures has remained as grand challenges in bioimaging. In this proposal, we aim to develop two Super-resolution Chemical Imaging techniques to tackle these challenges by utilizing a single contrast mechanism, the Raman scattering, to study the aforementioned less-explored directions of imaging small biomolecules and simultaneous imaging a large number (>20) of targets in cells, respectively. To do so, we plan to bring the most advanced and biocompatible Raman imaging modality, the stimulated Raman scattering microscopy, into the super-resolution regime. We will first prove the working principles for the designed spectroscopy and then demonstrate the microscopy methods. We aim to achieve a resolution improvement of more than 3 times, thus pushing the achievable imaging resolution to below 100 nm. This resolution range would be highly suited for interrogating the target biological structures and the dynamics. If successfully developed, these two novel Super-resolution Chemical Imaging techniques could be transformative and bring optical bioimaging to the next frontier for studying unresolved biophysical and biochemical processes in the complex biological systems. All these fundamental understandings would ultimately help researchers to elucidate the key functional roles of these intricate structures in biomedical fields including neurobiology and cancer biology.