Abstract

Mass spectrometry based proteomics has advanced tremendously in the last few years. We describe a shotgun proteomics workflow that allows us to detect and quantify the large majority of the proteins expressed in a biological system such as cancer cell lines and even formalin-fixed, paraffin embedded material. This included streamlined and highly efficient sample preparation, analysis with very high sequencing speed using modern mass spectrometers and bioinformatic analysis using the MaxQuant and Perseus platforms. Efforts in our group have focused on ‘single shot’ analysis and we demonstrate very high coverage in this mode (Mann et al., Mol. Cell, 2013). We have also extended this concept to the analysis of interactomes (Hein et al. Cell 2015) and phosphorylation, where the ‘EasyPhos’ method now allows acquiring large numbers of phosphoproteomes, for instance for the analysis of in vivo signaling (Humphrey et al. Nat. Biotech, 2015). Together such developments make proteomics increasingly relevant to translational research as I will discuss in this talk.