Shared UPLC-MS/MS for Absolute Quantitative Proteomics

[unreadable] DESCRIPTION (provided by applicant): A shared instrument, nanoUPLC-ESI-MS/MS, is requested for a focused group of NIH supported researchers in the School of Pharmacy at the University of North Carolina at Chapel Hill. The experienced group using the instrument will employ it primarily for absolute quantitative analysis of targeted proteins, especially those proteins involved in drug and metabolite disposition, such as enzymes of metabolism and transport proteins. Other Users will employ targeted absolute quantitative proteomic methods to follow human proteins delivered via lipoplexes in animal models, and to assess the expression of multiple sulfotransferases that modulate heparan sulfation in viral recognition. The targeted absolute quantitative proteomics approach is far superior to presently employed semi-quantitative Western blotting, since it has a large dynamic range, equal or greater sensitivity, better accuracy and precision, and is less time consuming. These methods, now sometimes employed in dedicated proteomics labs, can now be employed more widely, with high throughput, in labs commonly familiar with small molecule quantitative LC-MS/MS where stable isotope standards are routinely employed. Thus, the method will have a greater impact when it is used routinely by biomedical scientists with access to a shared facility. Targeted absolute quantitative proteomics will allow experimental approaches previously unavailable to our NIH supported research labs, thus advancing their science by being able to measure and quantify protein expression in tissue, or the organism directly, for almost any protein rather than measuring indirect or poorly correlated measures such as mRNA. Moreover, using stable isotope labeled peptides as standards will permit the methods to be compared between labs more easily. Targeted absolute quantitative proteomics is an exciting method that will quickly become mainstream, and the shared instrument requested herein will drive this effort at UNC Chapel Hill and establish methods that can be broadly disseminated and employed within the University and the general scientific community. [unreadable] [unreadable] Relevance (for the lay public) Current methods to detect major proteins that eliminate drugs from the body, or deliver drugs to targeted sites in the body, are time-consuming and often unreliable. The requested instrument will detect and quantify several of these proteins in a much more time-efficient and reproducible manner. The researchers using the instrument will explore the expression and delivery of proteins associated with drug metabolism and gene delivery, thus improving our understanding of drug-drug interactions and our ability to treat diseases. [unreadable] [unreadable] [unreadable]