ShEEP Request for Laser Microdissection Instrument

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We are requesting funds to purchase a Zeiss PALM Microbeam laser capture microdissections system. This instrument allows investigators to dissect and collect, down to the single cell level, specific fields, structures, and cells from tissue sections. Currently, neither the research facility at the Oklahoma City VA Medical Center (OKC VAMC) or the affiliated University of Oklahoma Health Sciences Center have the ability to perform this type of microscopic sample collection. It is well known that a principle limitation to analyses of bulk tissue is that there is a heterogeneity of cell types and any resultant molecular or biochemical analyses reflect the cellular status across this mixture of cells. Being able to analyze each cell type or structure separately has great advantages for understanding cellular function in health and disease and how different changes across multiple cell types in a tissue work in a cell non-autonomous fashion to cause tissue dysfunction and disease. Traditionally, the approach to resolving this challenge is to use cell sorting techniques. While cell sorting approaches are invaluable, in many cases they must be performed on fresh tissue, have appropriate cell surface markers or transgenic reporters, and require extensive technical optimization to ensure that cells remain unchanged during this process. An alternative approach is to microscopically cut specific cells out of tissue sections. In this approach tissues are collected in an intact form, either fresh frozen or fixed, sectioned and then imaged on the laser capture microscope. The cells or structures of interest and then excised with a computer controlled laser. These cells can then be used for a wide variety molecular and biochemical analyses. The only requirement is that there be some form of structure, stain, or immunohistochemical marker to identify the cell type(s) of interest. With this instrument the microdissected samples then are collected in a contactless manner ensuring sample purity and ability to use for many types of downstream molecular and biochemical analyses. The Zeiss PAM Microbeam instrument was selected based on the versatility of the instrument to meet the various needs of OKC VAMC investigators. The ability of this system to collect fields/cell layers is important to neuroscience and cancer investigators wanting to analyze microscopic structures within organs that cannot be dissected by hand. All of the investigators can make use of the ability to detect specific cell types with stains or antibodies and then collect only these cells. A future potential use is the ability to couple cell cultures with this instrument and collect specific types of lives cells for sub-culturing. Also central to instrument selection was the ability to ‘teach’ the robotic software to recognize a specific cell types based on the staining, size and shape to collect cell automatically. A survey of all VA researchers at OKC VAMC showed a wide variety of researcher will utilize the system. This instrument will be used by five major users and a number of future users who have six VA Merit grants funded, three senior career scientist awards, and one career development award funded.