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3D Cryo-Imaging System For Whole Mouse
Roy, Debashish

2009, Doctor of Philosophy, Case Western Reserve University, Biomedical Engineering.
We developed a novel 3D Cryo-imaging system that provides information rich, very high resolution, color brightfield, and molecular fluorescence images of a whole mouse using a section-and-image block face imaging technology. The system consists of a mouse-sized, motorized cryo-microtome with special features for imaging, a modified, brightfield/fluorescence microscope, and a robotic xyz imaging system positioner, all of which is fully automated by a control system. The system acquires microscopic tiled image volumes (limiting system resolution ≈3µm, nominal whole mouse pixel size ≈15µm, 40 µm sections), giving >60 GB of image data per mouse. Software is fully automated with programmable imaging/sectioning protocols, and email/cell-phone notifications for unattended imaging sessions. We developed a specialized, multi-scale visualization system which provides color rich volume rendering and multi-planar reformatting enabling one to visualize the entire mouse and zoom in to organ, tissue, and microscopic scales. A definable region-of-interest multi-sample array imaging scheme allows multi-sample studies of smaller specimens such as excised organs, mice and chick embryos. Autofluorescence images of excised chick hearts show cardiac details without administering any stains. Enhanced 3D visualization of mouse embryos with lacZ/β-gal whole mount staining shows sub-ectodermal cell expression patterns along arbitrary planes. In toxicity studies, transgenic mouse embryos expressing GFP in smooth muscles show developmental abnormalities due to toxin dosing during embryogenesis. We have anatomically characterized, in 3D from whole animal to tissue level, a transgenic mice that overexpress phosphoenolpyruvate carboxykinase (PEPCK-C) in skeletal muscle with astonishing metabolical and physiological consequences. We semi-automatically visualized and analyzed selected organs and found enlarged heart, much less visceral and subcutaneous adipose tissue but with increased vascularization, less pericardial adipose tissue, and higher tibia-to-femur ratio in the transgenic. Microscopically, individual skeletal muscle fibers, fine mesenteric blood vessels, intestinal villi, among others, were clearly seen. With a unique combination of field-of-view, depth of field, color contrast and very high resolution, the Case cryo-imaging system fills the gap between whole animal in vivo imaging and histology and is ideally suited for applications such as morphological phenotyping, stem cell imaging, developmental biology and multi-modality correlative studies with in vivo imaging techniques.
David Wilson, PhD (Committee Chair)
Michiko Watanabe, PhD (Committee Member)
Andrew Rollins, PhD (Committee Member)
Zhenghong Lee, PhD (Committee Member)

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Roy, Debashish. "3D Cryo-Imaging System For Whole Mouse." Electronic Thesis or Dissertation. Case Western Reserve University, 2009. OhioLINK Electronic Theses and Dissertations Center. 17 Oct 2017.

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Roy, Debashish "3D Cryo-Imaging System For Whole Mouse." Electronic Thesis or Dissertation. Case Western Reserve University, 2009.


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