Preparation and Immunoaffinity Depletion of Fresh Frozen Tissue Homogenates for Mass Spectrometry-Based Proteomics in the Context of Drug Target/Biomarker Discovery.

摘要: The discovery of novel drug targets and biomarkers via mass spectrometry (MS)-based proteomic analysis clinical specimens has proven to be challenging. wide dynamic range protein concentration in the high background/noise originating from highly abundant proteins tissue homogenates serum/plasma encompass two major analytical obstacles. Immunoaffinity depletion blood-derived is a well-established approach adopted by numerous researchers; however, utilization this technique for immunodepletion obtained fresh frozen lacking. We first developed immunoaffinity homogenates, using renal cell carcinoma as model disease, followed study applying it different types. Tissue homogenate may equally important recognized need serum/plasma, enabling more sensitive MS-based targets, and/or complex samples. Provided detailed protocol designed guide researcher through preparation two-dimensional liquid chromatography, tandem (2D-LC-MS/MS)-based molecular profiling context target biomarker discovery.