Deltavision RT Image Restoration System for Established Research Core Facility

Oregon Health & Science University, Portland OR

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Abstract

[unreadable] DESCRIPTION (provided by applicant): The MMI Research Core Facility at the Oregon Health & Science University is a well-established shared resource facility providing high-resolution fluorescence microscopy to more than 80 labs in 26 departments at OHSU and Portland metro area. This application requests funds to replace an outdated Applied Precision DeltaVision image restoration system with a new API DeltaVision RT system (DVRT) with the accessories needed for live-cell and laser enhanced applications. The DVRT will be used to complement imaging with laser scanning confocal microscopes (LSCM) available to our core facility. The requested DVRT will be used for imaging cell culture samples where the adherent cells are less than ~50 [unreadable]m thick and have relatively weak fluorescence. For these specimens, the DVRT's iterative constrained deconvolution image restoration surpasses LSCM imaging in resolution and signal to noise. The requested DVRT has shown itself superior for cells susceptible to photodamage or where the fluorescent molecules to be imaged are susceptible to photobleaching or dim, in experiments in which two or more colors of fluorescently labeled molecules are interacting, for acquiring a differential interference contrast (DIC) along with fluorescence and for highly accurate point-visiting for multiple data sets and for 3D and 4D reconstruction. New capabilities provided by the requested DVRT include 3-4 X faster color switching than our current system; 10-20x greater signal-to-noise (S/N) than we can achieve currently with low level fluorescence samples provided by our clients; a controlled environment chamber; an improved extremely accurate XYZ stage with auto-focus to maintain focus within each z-stack (set of optical sections along the Z-axis). This is important for time-lapse studies where stage drift and other mechanical or biological sources cause loss of focus during extended times (hours to days). The requested accessory diode lasers provide the focused illumination needed to perform fluorescence resonance energy transfer (FRET), fluorescence recovery after (laser induced) photobleaching (FRAP) and fluorescence loss in photobleaching (FLIP), to study dynamic interactions of specific proteins and subcellular components in living cells. In particular, the DVRT system has proven far superior to anything else available for extremely large files (>2GB) that require 3D or 4D reconstruction in order to properly understand the data. RELEVANCE - The >80 clients in >26 departments of our imaging facility do both basic biomedical and translational medical research. Examples include studies of the survival mechanisms used by disease causing organisms to evade detection in their human hosts, intercellular communication and control in the nervous system, formation of the immunological synapse between lymphocytes and antigen presenting cells, and cell to cell spread by human viruses, and pathogenesis and host-cell survival of Salmonella typhimurium, Francisella tularensis, Neisseria gonorrheae and N. meningitides. [unreadable] [unreadable] [unreadable]