Microscopy-based cell separation, selective propagation, and selective single cell sequencing

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The analysis of cells, or cytometry, is fundamentally important to many aspects of biological research. Flow cytometry has been the dominant cytometry technology since the 1960s because it is simple, high-throughput, and can be coupled with fluorescence activated cell sorting to separate specific cell populations. However, flow cytometry also has a number of limitations including (1) shear stress on the cell sample; (2) significant cell loss during processing; (3) difficulty in analyzing non-spherical cells, delicate cells, and cell clusters; (4) inability to assess sub-cellular structure and morphology. Many of these limitations could be addressed using fluorescence microscopy, but microscopy has the critical limitation of not being able to separate target cells, which in turn excludes many subsequent analyses, such as propagation to form clonal populations and single cell sequencing. Current technologies for isolating cells identified by microscopy involve physically transferring single cells using a micromanipulator, but this approach is difficult, low-throughput, and prone to contamination. The long-term vision of this research program is to develop microscopy-based technologies for cell separation, selective propagation, and selective single cell sequencing. Leveraging our recent work in this area, as well as advances in wide-field microscopy and image-processing, we believe our work will ultimately enable microscopy to compete with flow cytometry as a standard tool for cytometry, cell separation, and single cell analysis. This discovery grant will train HQPs at graduate, undergraduate, and post-doctoral levels. Trainees involved in the proposed research program will receive world-class training in an equitable, diverse, and inclusive environment that is dynamic and multi-disciplinary. Specific areas of research and training include optics, microscopy, instrumentation, image-processing, machine learning, polymer materials, cell biology, molecular biology, and genomics. Trainees will be able to leverage the high-level skills, accomplishments, and engineering acumen developed through their research projects into fantastic careers in industry and academia.