Enhancing Stimulated Raman Scattering Microscopy Capabilities for Biological Imaging

Abstract

Drug discovery and development remain a time- and cost-intensive task, over 90% of drugs that pass preclinical trials, ultimately fail to gain FDA approval. A factor in this high failure rate is that methods capable of monitoring drug systems and environments remain challenging. Traditional biochemical assays to understand drug properties provide insights into bulk cell-drug interactions and cannot provide spatially resolved information. Therefore, technologies capable of understanding drug responses at the cell-to-cell level may provide invaluable information to enhance drug discovery efforts. However, microscopy technologies that are informative of drug-cell interactions typically employ bulky fluorescent probes that are likely to interfere with drug behavior. Many label-free technologies promise to overcome this shortcoming but have limited capacity for high spatial resolution, rich chemical information, and/or live imaging. Stimulated Raman scattering (SRS) microscopy allows for such capability by acquiring vibrational signatures of all compounds while not requiring sample preparation or exogenous probes. As such, SRS has seen increasing adoption for label-free chemical imaging for diverse applications such as drug product stability, drug distribution in cells and tissues, drug-induced phenotypic responses, and Raman-based histology. While SRS microscopy offers a powerful, label-free solution for chemical imaging, its broader application has been constrained by inherent trade-offs between imaging speed, spectral coverage, and spatial resolution. This dissertation focuses on enhancing the technical and methodological capabilities of SRS microscopy to address these limitations and expand its utility in pharmaceutical research In this dissertation, I document the novel applications of SRS to advance the understanding of drug and cell interactions. Beyond typical SRS applications, this dissertation presents instrumental and computational enhancements to the SRS platform. Collectively, these advancements in SRS capabilities and applications establish SRS as a versatile tool in biological imaging.