BIH-Technologieplattform Proteomik

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Mass spectrometry-based proteomics offers the possibility of measuring thousands of proteins in a single run and has fundamentally changed the way that biological systems are analyzed. Latest developments in liquid chromatography-tandem mass spectrometry (LC-MS/MS) have reduced the effort and instrument time necessary for the determination of global proteomes down to several hours.

To accurately and precisely quantify proteins in complex samples, we employ and further develop, in a fit for purpose manner, all commonly used approaches, such as label-free techniques and techniques encompassing different types of isotopic labels. For proteome-wide discovery applications we use label-free, metabolic labeling (SILAC) and isobaric tagging (TMT, iTRAQ) approaches and for targeted SRM/PRM-applications we use synthetic spike-in reference peptides for quantification.

Our collaborations with basic and clinical research groups span all areas of quantitative proteomics, from profiling different biological states in cells, tissues and body fluids, to measuring protein interactions with other proteins, drugs and nucleic acids, to determining the spatial information of proteins and also the identification of their post-translational modifications, such as phosphorylation, ubiquitination and lysine-acetylation.

Data analysis and interpretation are facilitated with well-established software packages, such as MaxQuant, Perseus, Spectrum Mill and Skyline, and also in-house developed software tools that enable protein-, post-translational modification (PTM)- and pathway-based analyses. To integrate proteomics data with other –omics datasets, we collaborate across the BIH technology facilities, to provide a holistic molecular annotation of biological and clinical samples.

Our approach enables a highly accurate and precise protein characterization of biological and clinical samples and will contribute to the identification of new candidates for disease-associated biomarkers and new therapeutic targets in the future.

Team

Dr. Philipp Mertins, Head (philipp.mertins@mdc-berlin.de)
Dr. Marieluise Kirchner, Senior Postdoc/Staff Scientist (marieluise.kirchner@mdc-berlin.de)
Colleagues at the integrated MDC Proteomics platform

Equipment

  • 1 Q Exactive Plus Hybrid Quadrupole-Orbitrap Mass Spectrometer (Thermo Scientific)
  • 1 Orbitrap Fusion Tribrid Mass Spectrometer (Thermo Scientific)
  • 1 TSQ Quantiva Triple Quadrupole Mass Spectrometer (Thermo Scientific)
  • 1 Triple-Quadrupole Mass Spectrometer, 6500 (ABSciex)
  • 4 nanoHPLC systems, Eksigent nanoLC 415 (ABSciex)
  • 1 sample processing roboter RTC-PAL

Analysis methods

  • Global proteome and post-translational modification (phosphorylation, ubiquitination, lysine-acetylation) profiling
  • Customized proteome-wide quantification techniques, including label-free, SILAC, stable isotope dimethyl labeling, iTRAQ and TMT
  • Targeted LC-MS/MS assays using SRM/PRM for highest accuracy and target verification
  • Automated sample preparation using RTC-PAL robotics
  • Data processing for primary peptide-, protein-, and PTM-site datasets in MaxQuant, Spectrum Mill and Skyline
  • Data analysis in Perseus and R-packages with Shiny user interfaces
  • Pathway- and signature-based analysis using single sample GSEA
  • Feature identification and significance testing for differential, correlation, class-marker, enrichment, clustering, and outlier analysis
  • Proteogenomic data integration by identification of sample-specific proteins with amino acid variants and cross-correlation analysis of CNA, RNA, and protein datasets
  • Data quality control by routinely monitoring LC-MS/MS performance, quantification and interference analysis, annotation of MS2 spectra, and cross-correlation of multiple datasets