BME PhD Preliminary Exam Announcement for Hao-Cheng Gao (F. Huang, advisor) Everyone is invited to attend the public presentation beginning at 2PM. Research title: Ultrahigh-Resolution Imaging and Quantitative Analysis Through Brain Tissue with high accuracy Date: October 11, 2023 Time: 2PM Location: MJIS 2001 Thesis Committee members: Fang Huang, Chair; Alexander A. Chubykin; Daniel M. Suter; Kevin J. Webb Abstract: 4Pi single-molecule localization microscopy (4Pi-SMLM) enables the revealing of ultrastructure by combining interferometric coherent fluorescence imaging and single-molecule localization. Current methods model the interferometric point spread function (PSF) by scanning in-vitro beads on coverslips to infer in-situ 3D location of single-molecule fluorophores in specimens. Although a resolution of sub-20-nm throughout the whole cell has been achieved, imaging tissue specimens was impeded by the heterogeneous aberration and scattering in tissue environment. Here, we introduce 4Pi in-situ PSF retrieval method (4Pi-INSPR), which allows 12-nm-resolution visualization and quantitative analysis through brain tissue with high accuracy. 4Pi-INSPR incorporates sample-induced aberrations, previously unattainable by conventional 4Pi-SMLM. 4Pi-INSPR also minimizes reconstruction artifacts from instrumental imperfections such as objective and imaging-quadrant misalignments. We demonstrate 4Pi-INSPR capabilities by revealing the ultrastructure of dendrites and dendritic spines in 50-μm mouse brain sections and quantifying their morphology changes and nanoscale molecular distribution. Together, 4Pi-INSPR provides potential in examining synaptic functions for brain development.