Comprehensive investigations at the cellular level to study the expression and fate of specific cellular products requires a combination of biochemical and morphologic techniques. These complementary tools can help to formulate a complete image of the interrelationship between the various stages and events that occur during the lifetime of a cellular product. To provide appropriate morphologic evaluation of gene products expressed along the gastrointestinal tract we will use a classical approach combining light and electron microscopy coupled with immunocytochemistry. Initial light microscopic evaluations provide the framework for screening and selecting materials for immunocytochemical and ultrastructural studies. With these techniques we will survey tissues of interest and screen and test our antibody preparations. Immunocytochemistry and electron microscopy allow for precise localization of specific cellular products and provide needed controls for subcellular fractionations, protein electrophoretic separations, and immunoprecipitations aimed at studying alterations in the cellular expression and processing of gene products of interest. Characterization and localization of gastrointestinal tract (GIT) products at the submicroscopic level will be accomplished utilizing a variety of electron microscopic techniques. Many studies will only require standard transmission electron microscopy. Quantitative evaluation of both light and electron microscopic studies will be accomplished using stereologic techniques and automated image analysis on a personal computer-based system. This methodology will provide valuable qualitative as well as quantitative data when used in concert with biochemical studies. Immunoelectron microscopy will be extensively utilized to study the secretory pathway of a variety of GIT products and to verify altered gene expression in transfected epithelial cell lines and transgenic mice.
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