Each of the projects of this PPG application utilize mouse models to assess innate immune cell function in infectious, inflammatory and autoimmune disease. As such, each project will rely on histologic analysis of tissues from experimental animals to determine innate immune cell numbers, location, and function (such as cytokine secretion). Histologic analysis of lymphoid architectures in relationship to the disease models being studied will also be a key method for each of the projects. Since the PPG envisions extensive sharing of animal resources, ideas and technologies, it is important than standardized methods of tissue preparation, immune cell staining and immunofluorescent detection be utilized by all four projects in the PPG. This will allow rational comparison of results between the projects. The Histology Core will be run through Dr. Lowell's group (Project #3) and will perform the following analyses: 1) standard tissue fixation, paraffin embedding, sectioning and histochemical staining;2) frozen tissue embedding sectioning and immunohistochemical staining;3) frozen tissue embedding with immunofluorescent staining and fluorescent cell localization;4) Multiplex cytokine analysis on serum, tissue and cell supernatant samples using Luminex technologies. Core personnel have extensive experience with all of the above methods, as described in the Preliminary data. The core is equipped with an OTF5000 cryostat for frozen tissue sectioning and a standard Leica microtomb for paraffin sections. Routine microscopy is done with an Eclipse TS100F epi-fluorescent inverted microscope with Cool-Scan 3.0 mega pixel digital imagining system. Access to larger upright Zeiss epi-fluorescent microscopes is available through the UCSF Immunology Imaging Core. This PPG core will also contain a staining station for routine histologic stains and will maintain a battery of mAbs for immunostaining. Multiplex bead based analysis of both cytokines, intracellular signaling molecules and nucleic acids can be performed on the new BioPlex200 analyzer that will be available to PPG personnel.
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