The objectives of this proposal are to purchase a state-of-the-art multiplex assay system for the detection of soluble molecules (cytokines, growth factors, metabolic markers, hormones, mediators of inflammation, other clinical markers) in order to support, enhance and fulfill the specialized technologic needs of a group of multiple NIH-funded researchers at UT Southwestern. The major user group conducts research in varied areas of immunology, hormone production, studies of cell signaling and differentiation, and soluble factor production. All investigators require the proposed instrument in order to enhance scientific insight into the questions being addressed in ongoing projects. As our understanding of the immunologic basis of various disease situations grows, it is becoming increasingly clear that populations of immune cells are not homogeneous and produce cytokines in a complex array of combinations. These cytokines need to be studied in an increasingly multiplexed and in depth fashion in order to understand their precise role in intercellular modulation of function as well as in therapeutic settings. In the study of growth factor and hormone receptors, several recent reports indicate that these receptors serve as sensors that can coordinately regulate the expression of genes controlling the intracellular levels of their ligands and induce cell differentiation. As with cytokines, the array of molecules present in a single sample (body fluid, cell culture supernatant, cell lysate) can be extraordinarily complex. While these molecules can be detected by conventional ELISA methods and, more recently, by cytometric bead array systems, robust simultaneous detection of multiple molecules in a single sample has not been possible, especially where small quantities may be available for evaluation, as is often the case in clinical/translational research. Availability of the unique electrochemiluminescence detection methodology in this novel system will allow such multiplexed analyses and add substantially to our understanding of the interplay between various cell types and a host of soluble molecules and their receptors. In addition, powerful utilization and analysis of multi-parametric data can be attained through bioinformatics. We present here the justification and rationale for our proposal, cite various specific examples of how the instrument would provide greater insight into ongoing NIH-funded research and outline the strong institutional commitment that will enable the smooth implementation of the NCRR SIG program. PROJECT NARRATIVE: Analysis of soluble molecules is a critical component of contemporary basic, translational, and clinical research. However, the ability to dissect complex biological fluids is hampered by inadequate methods. We propose to purchase the Meso Scale Discovery SECTOR(r)Imager 6000 reader. The instrument has multiplexing, wide dynamic range, sensitivity, and high throughput capabilities that are not present on currently available instrumentation at UT Southwestern or on other instruments in the market. The instrument is unique in its ability to simultaneously analyze multiple soluble molecules present in a wide variety of samples
