The National Center for Microscopy and Imaging Research (NCMIR) is centered around computer-aided intermediate voltage transmission electron microscopy (IVEM). These facilities are available to investigators working on biological problems that could benefit from the use of thicker specimens and 3D image analysis. Our plan for continuation of the core technological research is directed towards the development of technologies that will contribute to structural neurobiology, cell and molecular biology. Our goals for collaboration, service, training and dissemination are all related to expanding the use of these technologies to maximize their value to the biomedical community. Our core biological projects include development of new selective staining methods for IVEM and for correlated light and IVEM analyses of specimens to examine the architecture of neuronal systems and the dynamics of subcellular processes, new IVEM-compatible macromolecular probes for protein localization, methods to improve penetration of probes and improved methods for high resolution nucleic acid detection. Core research is tightly linked to collaborative projects, in areas such as neurodegenerative diseases, stroke, heart disease, tumor biology and HIV. The capabilities to be developed and research accomplishments anticipated in all of the Core specimen preparation areas will be augmented by available and planned features of the Resource image processing facilities. We emphasize the use of computer-aided methods for the enhancement of image contrast, extraction of accurate 3D information from within single thick specimens and the 3D reconstruction of larger structural complexes using tomographic and serial thick section analysis. We plan to implement or develop improve methods for tomographic reconstruction, 3D data analysis and visualization. Ongoing projects will expand our capability to analyze large 3D data sets by distributing computation to supercomputers linked to the Resource via high-speed networks. We will continue to develop a system allowing a remote user to control and acquire data from the IVEM and distribute computational tasks to supercomputers via the network; thereby providing researchers greater access to the Resource. Plans are proposed to extend this technology to other microscopes at the NCMIR and other microscope facilities. To provide enhanced imaging of thick sections, we plan to acquire a next generation, energy filtered 300 keV IVEM and equip it with new imaging modes developed at the NCMIR. We plan to expand outreach efforts via a web-based repository for tutorials, software and sample data related to structural biology with links to other biological databases.
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