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- 저자 : Croxford, Matthew (검색결과 1 건)
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Croxford, Matthew ProQuest Dissertations & Theses University of Cali 2022 해외박사(DDOD)
The field of cell biology originated with, and has always been inextricably tied to, the microscope. Direct visualization has always been a key technique for understanding cell organization and behavior. Today, cell biologists have microscopes many orders of magnitude more powerful than their progenitors, enabling the direct visualization of cells and their constituent parts down to sub-nanometer resolution. Cryo-focused ion beam milling combined with cryo-elecyron tomography (cryo-FIB-ET) is a technique in which cells are rapidly frozen to preserve them in their native state, followed by focused ion beam (FIB) thinning of the frozen cells, which allows for the collection of high resolution transmission electron microscopy data. When these electron micrographs are collected over a range of different tilt angles, a full three-dimensional reconstruction of the cellular interior can be made and analyzed. However, despite the unprecedented level of detail available to cell biologists through cryo-FIB-ET, several challenges hinder its applicability to cell biology. On the experimental side, commonly used cryo-FIB-ET compatible substrates were not designed with cell biology in mind, and have features that make them sub-optimal for this purpose. Furthermore, limitations in the range of tilts at which data may be collected lead to missing information in the three-dimensional reconstructions that cause distortions and make the resulting volumes more difficult to interpret. The work presented in this dissertation focuses on both of these issues in an attempt to increase the utility of cryo-FIB-ET for cell biologists and expand the range of questions that may be addressed with the technique.
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