The field of cellular biology is undergoing a revolution due to recent advancements in serial block-face imaging scanning electron microscopy (SBF-SEM). This technology combines automated sectioning of plastic-embedded tissue blocks with imaging by SEM, and does so in a fully-automated fashion. SBF-SEM enables users to generate and image many hundreds (>1000) of serial sections at a rate of 30-40 seconds per section. This Z stack of sections is then computationally assembled into a 3D image, creating a very high-resolution volume. The most recent advance in SBF-SEM has been the development of automated multi-energy deconvolution (MED) technology by the company FEI. MED enables users to obtain resolution in the z dimension of less than 10 nm. Previously, z resolution was limited to the minimum thickness of microtome-generated sections (~30 nm). FEI has now incorporated MED technology into their SBF-SEM system, which they have named the Teneo LoVac VolumeScope (Teneo VS). This microscope enables users to generate 3D isotropic data at 10 nm resolution, and to do so over a very large volume (up to ~1000 m x 1000 m x 1000 m). With this combination of volume and resolution, cell biologists can now generate ?Google Earth? representations of complex tissues such as brains and liver. Such datasets will enable a researcher to quickly scan their specimens for features of interest, and then zoom in 100,000 fold to examine these features in exquisite detail. Equally important, these datasets will enable researchers to visualize how complex tissues are assembled from their cellular components, and even how the internal components of cells are structured in 3D. This proposal seeks funds to purchase an FEI Teneo VS, which will be installed in the Indiana University Bloomington-Electron Microscopy Center (IUB-EMC). The IUB- EMC is a multi-user facility that serves the entire IU system (8 campuses), and is a service core of the NIH- funded Indiana Clinical and Translation Sciences Institute (CTSI), which includes Purdue University and Notre Dame University. To date, no EM center in Indiana has installed an SBF-SEM. Indeed, the Teneo VS is so new that only two have been installed in the USA, thus the IUB-EMC seeks to become a regional center for SBF-SEM imaging. To demonstrate the broad impact of this technology on NIH-funded research at IUB, projects from seven major users are described. These users are Dan Tracey, who is investigating the neural circuits underlying pain perception, Andy Zelhof, who is investigating eye development, James Glazier, who is investigating how acetaminophen damages liver, Mike Lynch, who is investigating the evolution of cellular structures, Ke Hu, who is investigating the cell biology of Toxoplasma gondii, an important human pathogen, Tuli Mukhopadhyay, who is investigating arbovirus assembly inside mammalian cells, and Roger Innes, who is investigating the role of extracellular vesicles in the immune system of plants. Although extremely diverse, these projects will all be greatly accelerated by access to SBF-SEM.
We propose to acquire a Serial Block-face Imaging Scanning Electron Microscope (SBF-SEM). SBF-SEM technology enables users to obtain three-dimensional images of the interiors of cells and tissues at unprecedented resolution (<10 nm in x, y, and z dimensions), and to do so over a much larger area (1000 m x 1000 m) than competing technologies. This microscope will become part of the Indiana University Bloomington Electron Microscopy Center and will be available to IU researchers throughout the IU system. Access to SBF-SEM technology will greatly enhance the ability of Indiana University scientists to visualize the interior structures of cells and tissues, and thus answer fundamental biological questions previously out of reach.
