The Single Nephron and Metabolomics Core will provide an important national resource for investigators who wish to define the expression, localization and functional characteristics of transport and other relevant proteins in single nephron tubules or defined renal epithelial cells. Moreover, metabolomics services will be available to measure changes in levels of small molecules involved in regulating nephron function. It is expected that the data generated from this Core will be complemented by analyses performed in other Center Cores, such as the Cellular Physiology and Kidney Imaging Cores. The Single Nephron and Metabolomics Core aims to offer an integrated approach including functional (including in vitro microperfusion of isolated segments, measurements of transepithelial ion/solute fluxes, fluorescence functional imaging of single tubular cells), biochemical (microassays of enzyme/transporter activity), molecular (single tubule quantitative PCR and immunoblotting), and analytical (renal metabolomics) strategies applied to microdissected tubules to address relevant questions proposed by users. Furthermore the Core will provide expertise in design and implementation of single nephron/cell studies and instruction in the technical aspects of all services offered by the Core. The specific objectives of the Core are to: (1) provide microdissected tubules for quantification of mRNA abundance (real time PCR) and protein expression (immunoblotting), immunolocalization, and enzyme/transporter microassays;(2) perform functional fluorescence assays of channel/transporter function in isolated tubules microperfused in vitro;(3) perform measurements of transepithelial ion/solute fluxes across isolated tubules microperfused in vitro;(4) quantify mRNA and protein abundance in urinary exosomes;(5) perform metabolomics analyses of microdissected tubules and perfusate;and (6) provide training in all of the above.
This Core offers a functional, biochemical, molecular, and analytical approach applied to microdissected tubules and defined renal epithelial cells to address relevant questions proposed by users. Core B's expertise in measuring transport of ions, solutes and other molecules across renal epithelial cell membranes, available in few labs nationally, promises to provide novel insight into our understanding of kidney disease.
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