Microfluidic Tools for Studying Development in Embryos and Brain Organoids
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Charles, Seleipiri
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Abstract
Development in multicellular organisms is a complex process requiring multiple intracellular and extracellular signaling events. High content screening tools enable the cellular and subcellular assessment of developmental changes, which in turn have led to a variety of genetic, pharmacological, and therapeutic advancements involving multicellular organisms. Developing high-content screening tools for multicellular systems require high resolution imaging of protein and gene expression changes, relatively large number of samples to better characterize inter and intra-population differences, and multiplexed readouts using the same sample to obtain layered information about developmental changes. Microfluidics can address these challenges by enabling high magnification imaging, parallelization, rapid reagent delivery and exchange and lower reagent consumption. Hence, this thesis sought to address high content screening challenges that affect the study of development in two multicellular systems: C. elegans embryos and brain organoids. This thesis will demonstrate this through three specific aims that involve developing and improving microfluidic-based technology and assays for large-scale imaging and characterization. As a result, I developed a microfluidic-based assay for conducting high temporal resolution measurements of gene expression changes during C. elegans embryogenesis using single molecule fluorescence in situ hybridization (smFISH). Next, I developed an integrated platform to enable robust and long-term culturing of brain organoids. I designed a mesofluidic bioreactor device based on a unique diffusion-reaction scaling theory, which achieves convective media exchange for sufficient nutrient delivery in long-term culture. Finally, I modified several aspects of the integrated platform to enable live and longitudinal imaging of organoids during culture for in situ characterization of cell quality and differentiation. Combining high throughput microfluidic technology with high content imaging tools will improve the characterization of factors affecting development in these two biological systems.
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Date
2022-05-03
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Dissertation