CCR VB FACS CORE

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Project Summary The Vaccine Branch FACS Core Facility has been created to meet an unmet need on the Bethesda Campus of NCI for a Flow Cytometry Core Facility with BL2/BL3 containment necessary to safely sort live cells infected with retroviruses, such as HIV/SIV, and other infectious agents known or suspected in blood and tissue samples. In addition, this facility has analysis capabilities from four flow cytometry instruments that were donated to the facility from three Investigators in the CCR. Laboratory space renovations were completed in May 2008, and the donated flow cytometers were relocated to the facility in June 2008. The facility currently maintains two investigator- run 2 laser analog flow cytometers, one investigator-run 2 laser digital flow cytometer, and one investigator-run 3 laser digital high end flow cytometer. The 3 laser digital high end flow cytometer has undergone extensive hardware and software upgrades to add additional fluorescent and sample processing abilities, and one of the 2 laser analog flow cytometers has been upgraded to a new computer system. The facility has been approved to purchase a 4 laser high end analyzer and a 3 laser high speed cell sorter. These will be purchased at the beginning of FY09. The facility currently supports more than 25 Investigators from the Vaccine Branch, the Laboratory of Receptor Biology and Gene Expression, and the Laboratory of Cellular Oncology. Since the facility has only been open for 4 months, the number of investigators is expected to increase significantly. Experiments currently conducted in the facility involve multi-parameter single cell analysis of cells from human, non-human primate and mouse primary sources. Additionally analysis is also conducted on in vitro cultured cells. Reagents for these experiments include monoclonal and polyclonal antibodies with fluorescent labels and fluorescent dyes used for staining a variety of cellular components. Current applications being supported by the facility include multi-color phenotypic analysis, cell cycle analysis, apoptosis analysis, proliferation analysis with BrdU and CFSE, intracellular cytokine analysis, rare event analysis and calcium flux analysis. Currently supported projects include, but are not limited to, immune monitoring for HIV, SIV, HTLV-1 and HTLV-2 vaccine studies on non-human primates, and monitoring the results of gene expression on cell proliferation and apoptosis.