CORE--CELL MORPHOLOGY

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The Cell Morphology Core (CMC) will support the grant members with all aspects of histology-related work necessary to accomplish their research goals. Key services will be transgene detection in tissues (immunohistochemistry, in situ hybridization, reporter gene detection such as beta-galactosidase and GFP), viral vector evaluation by electron microscopy, and standard histology for pathological evaluation. Specifically, The CMC prepares sections from paraffin, frozen and plastic-embedded tissues. Routine and special stains, as well as apoptosis and cell proliferation assays, are performed regularly. Immunohistochemistry is performed with fluorescent or enzyme-labeled antibodies using all the appropriate positive and negative controls. In situ hybridization for identification of transduced cells or the detection of viruses uses labeled DNA or RNA probes. Light microscopic images are taken with a digital camera and can be sent to investigators electronically or on a disc. Image analysis can be performed, quantifying the degree of staining for any target. Transmission electron microscopy (TEM) is performed to evaluate tissue samples at an ultrastructural level. This technique can be combined with immunostaining (immuno-electron microscopy) for the specific subcellular localization of transgenic proteins with gold-labeled antibodies. For vector analysis, negative staining of viral preparations is applied for EM analysis. Virus vectors are assessed for average density, purity, uniformity and viral particle integrity. Contaminations in adenovirus preparations with AAV (and vice versa) are detectable due to the difference in size of both viruses. Photographs are taken, developed and printed within the CMC. Investigators can receive either the hard copy photographs or computer files including written evaluations.