Abstract

Functional Protein Profiling in Tissue Samples for Next Generation Molecular Diagnostics

Karl-Friedrich Becker, Head, Technical University of Munich

Proteomics methods are very slowly or not at all translated into current clinical workflows for tissue analysis. This is in sharp contrast to proteomics applications in basic sciences. Reverse phase protein arrays (RPPA) represent a very promising sensitive and precise high-throughput technology for the quantitative measurement of hundreds of signalling proteins in biological and clinical samples. This array format allows quantification of one protein or phosphoprotein in multiple samples under the same experimental conditions at the same time. Moreover, it is suited for signal transduction profiling of small numbers of cultured cells or cells isolated from human biopsies, including formalin fixed and paraffin embedded (FFPE) tissues. After protein extraction, each patient sample is arrayed in duplicates on nitrocellulose-coated slides in a miniature dilution curve. Validated antibodies are used to detect the proteins of interest. Thus, each analyte/antibody combination can be analysed in the linear dynamic range. The aim of this presentation is to highlight the progress of RPPA studies with human tissues for functional protein profiling and to discuss the challenges that must be understood and addressed for successful integration of RPPA to clinical practice. A unique tumour tissue sample collection for quality control and analysis of protein/phosphoprotein biomarker stability as determined by RPPA is introduced.