We propose to acquire the Carl Zeiss Cell Observer spinning disk confocal system, to support a large cadre of NIH-funded investigators at the George Washington University (GW) and its partner institutions. This system will primarily be dedicated to short and long-term live-cell imaging experiments, as well as other current applications, including high-throughput/high-content image acquisition. The acquisition of the instrument will surmount the large array of current barriers for live-cell imaging, a result of our existing point scanning confocal microscopes. For cutting-edge, live-cell imaging microscopy, these are essential technological requirements that must be addressed for time-lapse recording of fluorescence signals to study cell motility, organelle trafficking, free living nematodes (c. elegans) and murine genetically engineered embryos. The proposed spinning disk microscopic technology is uniquely tailored to rapid sampling of fluorescent images from living specimens, while providing high detection sensitivity and limited light radiation. Minimizing light radiation s critical for most live-cell functional assays in order to reduce potential phototoxicity and generation of reactive oxidative species. Spinning disk microscopes subject specimens to much lower light levels than the presently used point scan confocal microscopes and therefore ideally suited for live-cell imaging. Numerous projects will benefit from acquiring the instrument, including research in molecular mechanisms of neural development at the cellular and organelle levels, dysregulation of cell cycles associated with tumor cells, molecular parasitology investigating signaling pathways controlling re-initiation of hookworm development, host-pathogen interactions related to HIV and transgenic helminth parasites, and hepatobiology and molecular basis of liver diseases and fibrosis. The instrument will be integrated into the infrastructure of GW's well-established Center for Microscopy and Image Analysis (CMIA), and made available to members of NIH-funded Centers, including the Intellectual and Developmental Disabilities Research Center (IDDRC) at the GW-affiliated Children's National Medical Center and the District of Columbia Developmental Center for AIDS Research (DC D-CFAR). In addition, CMIA will manage and regulate maintenance of the instrument, design and implement appropriate training programs, methodology development and dissemination to the research community. To conclude, the proposed instrument acquisition will address existing deficits and positively impact NIH-funded research programs and create new collaborative venues for complementary Centers and Institutions. As such, we anticipate the user base to expand substantially with the acquisition of this new technology and implementation of training. Members of the advisory committee, the principal investigator on this proposal, and the broader user group will be involved in collectively managing future alterations to the user base and prioritization of access;the Director of the CMIA will have sole responsibility for training and compliance management.

Public Health Relevance

The requested spinning disk confocal microscope allows the visualization of cellular processes in living cells in biologically relevant time-scale. This microscope will advance numerous research programs, including understanding of developmental neural disorders associated with intellectual deficits, the molecular basis of cancer transformation, and the molecular basis for parasitic and HIV infections. The overarching goal of these projects is to improve our understanding of the mechanisms underlying those diseased states and to design novel strategies to eradicate them.

National Institute of Health (NIH)
Office of The Director, National Institutes of Health (OD)
Biomedical Research Support Shared Instrumentation Grants (S10)
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Special Emphasis Panel (ZRG1-CB-N (30))
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Levy, Abraham
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George Washington University
Other Domestic Higher Education
United States
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Brichacek, Beda; Darwish, Christina; Popratiloff, Anastas et al. (2014) HIV-1 infection of macrophages induces retention of cholesterol transporter ABCA1 in the endoplasmic reticulum. AIDS Res Hum Retroviruses 30:947-8
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