Application): The Brookhaven STEM (Scanning Transmission Electron Microscopy) Resource provides a unique service for mass mapping and heavy atom labeling of individual biological molecules. The STEM achieves mass resolution of 0.5 to 5% for proteins or complexes as large as viruses or cell organelles, imaged at low dose. We have developed heavy atom labels to mark specific sites or subunits within the overall contour of molecular complexes. The Core R&D program seeks to advance the state-of-the-art in four areas: 1) new heavy atom cluster labeling reagents for important biological applications, 2) development of mass mapping using frozen-hydrated specimens in the Cryo TEM/STEM, 3) mass/image analysis improvements to provide additional features, and 4) STEM instrumentation development to improve performance and add new capabilities. All four should enhance the quality & quantity of users' results. We propose to continue to expand our productive user/collaborator program as we develop new labeling methods and improve specimen preparation, imaging, and image analysis. Currently 40 to 50 projects are active in the areas of: proteins and protein complexes, filaments and filamentous proteins, membranes and membrane proteins, DNA, RNA and nucleic acid/protein complexes, viral proteins, virus structure, cell organelles, and in particular the fields of filament and virus assembly. An average of 25 new projects are added each year. All projects are expected to lead to publications in the highest quality journals. The STEM Advisory Committee plays an integral role in planning, quality control and development of new projects. Training and dissemination will continue using scientific publications, meetings, workshops, brochures, our web site, and expanded to include a new FTP site for training and additional workshops. The Brookhaven STEM has proven to be a valuable resource to many investigators, and we look forward to providing an even stronger and more useful Facility to the scientific community, which is not available elsewhere in the U.S.
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