Enhanced Dynamic Range Proteomic Analysis: Phase II

DESCRIPTION (provided by applicant): The long-term goal of this effort is to develop an optical module that will reduce stray light within imaging systems, thereby providing more accurate measurements from digital images and increased dynamic range of detection to enable analysis of objects not currently measurable. The present Phase II proposal is aimed at greatly enhancing the analysis of multicolor spectral imaging of fluorescent dyes in proteomics to detect changes in protein levels and protein posttranslational modifications in gel electrophoresis. Potential future applications of the optical module will improve multicolor fluorescence detection in immunohistochemistry and analysis of fluorescent proteins in cells and tissues, microplate reading and microfluidic analysis for new methods of multiplex diagnostics. The system to be developed will be usable on nearly all optical imaging systems so as to broaden the scope of applications and ultimately reduce the cost. The specific aims are to: (1) Develop and characterize a Noise Reduction Module (NoRM). This system will record an initial image, then utilize a feedback loop to turn off the bright pixels and associated stray-light to more accurately measure regions within an image; (2) Demonstrate the NoRM in proteomics applications, which will validate the technology, and enable detection of proteins and patterns of proteins in 2-D gels that are currently too weak to observe (the enhanced protein patterns are expected to have diagnostic value), and guide enhanced electroelution/microfluidic digestion/integrated mass spectral analysis ; and (3) Prepare a production prototype NoRM for a limited scale release by the end of the Phase II effort. This Phase II effort builds on a successful Phase I effort that demonstrated greater than factor of 10 improvement in dynamic range as compared to currently used image bracketing technology. The proposed effort is cross-disciplinary, with expertise required in optical and mechanical design, software development, productionengineering, biochemistry, proteomics, and systems biology. The resulting technology will be useful for proteomics, microscopy, and many other technologies that utilize digital cameras. PUBLIC HEALTH RELEVANCE: The proposed technology will reduce the stray light noise for digital imaging systems, thereby expanding capabilities for proteomics, glycomics, cell biology, diagnostics and any biomedical application that utilizes digital cameras. During this effort, patterns of weakly-expressed proteins andchanges in these proteins, whose signals are currently too weak to be identified in electrophoresis gels will be measured and identified to better understand biological mechanisms, improve development of more specific drugs, and enhance regenerative and preventative medicine.