Funds for the purchase of a Meridian Instruments Corporation ACAS 570 Interactive Laser Cytometer/Cell Sorter are requested. This instrument is an integrated fluorescence analysis and sorting system for the interactive investigation of morphologically and functionally defined rapid biological processes at the single cell and subcellular level. It is the only commercially available instrument using a laser to both interrogate and analyze individual viable cells, and interacting groups of cells on the stage of an inverted fluorescence microscope. The specific configuration requested includes a 100 mW argon laser, acousto-optic modulator for millisecond control of laser pulse duration and intensity, dual sensitive photomultiplier detectors, computer controlled stage with selectable 0.25 micron resolution stepping, 80386 based microcomputer hardware, 37o mini- incubator, and Olympus IMT-2 inverted microscope. The system allows the investigation, in a controlled atmosphere, of cell-cell communication via gap junctions, lateral diffusion of membrane proteins and lipids by photobleaching, cytoplasmic tracer diffusion, time-resolved cytoplasmic free calcium concentration, pH, H2O2 production, as well as quantitative measurement of structures that can be identified with fluorescent substrates. The latter includes the ability to measure beta-galactosidase activity, a prokaryotic enzyme that can be transfected into eukaryotic cells for the purposes of studying gene promoter function and cell lineage. The instrument's laser can also be used to select viable cells of interest from a mixed population. Investigators from six major departments at three institutions, the Yale Medical School, Yale School of Public Health and Harvard Medical School, all sharing an interest in the molecular mechanisms of cell-cell interaction, signal transduction, gene regulation, have been identified as the major users for this instrument. The acquisition of the instrument would extend ongoing work by these investigators into the forefront of interactive single cell research, lead to new collaborative projects by the involved investigators, and likely also lead to new applications of the technology. The P.I. has extensive experience in the organization of fluorescence core facilities and there is an institutional committment for space and operating funds.
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