A New Approach to 3-Dimensional Super-Resolution Microscopy

Thesis (Ph.D.)--University of Washington, 2019Super-Resolution microscopy has transformed our ability to image biological specimens at the nanoscale with high contrast and molecular specificity. However, acquiring 3-dimensional images over large volumes remains a challenge because the imaging process causes fluorophores outside the focal volume to photobleach before they can be imaged. Additionally, acquiring highly multiplexed images is challenging due to a lack of spectrally distinct fluorophores which possess the required photophysical properties for super-resolution imaging. Here I present my work which specifically addresses these issues. In chapter 2 I first discuss the importance of sample preparation for super-resolution imaging. In chapter 3 I introduce a novel labeling scheme called probe-refresh STORM (prSTORM) which enables practitioners obtain multiplexed, extended-depth super-resolution images with a single dye, and in chapter 4 I show the importance of hardware for super-resolution microscop