Platform 1: Integrated Proteomics and Lipidomics
Proteomics: Prof. Dr. Stefan Tenzer (University Medical Center Mainz)
Lipidomics: Dr. Laura Bindila (University Medical Center Mainz)
The integrated proteomics and lipidomics platform provides latest generation quantitative mass spectrometry techniques to the SPP 2225 to decipher the underlying molecular mechanisms of host cell egress by intracellular pathogens and to identify the mediators, regulators and effectors involved.
The proteomics platform headed by Prof. Dr. Stefan Tenzer specializes in label-free, quantitative proteomic analyses. To support the individual projects within the consortium, sample preparation and subsequent analysis is optimized in close consultation with the participating scientists to achieve ideal coverage of post-translationally modified peptides. The generated peptides are separated by high-resolution liquid chromatography (nanoUPLC) and analyzed using data-dependent and data-independent measurement methods on our Thermo Orbitrap Exploris 480 and Bruker TIMS-TOF Pro instrument platforms. The collected raw data are processed using an appropriate reference proteomic database. Label-free quantification of the peptides is performed using an in-house developed software pipeline ISOQuant, which includes all necessary post-processing steps, including retention time alignment, clustering, normalization and TOP3-based absolute quantification of the detected proteins. This process provides both high proteome coverage and high reproducibility. Furthermore, only small amounts of starting material are required for complete proteome analyses (3-20 µg total protein, corresponding to approximately 30-200,000 cells).
The lipidomics platform headed by Dr. Laura Bindila focuses on absolute quantification and qualitative lipid profiling on latest generation instrument platforms for the investigation of lipidome dynamic in various biospecimens under physiological and pathophysiological conditions. Using LC/MS profiling on QTRAP MS, targeted discovery as well as absolut quantification of selected lipids is afforded, which is particularly beneficial for low abundant lipids such as signal lipids. A general, deep lipidome analysis for biomarker discovery and profiling as well as structural identification of lipid species is afforded by modern high-end mass spectrometry platforms such as TIMS TOF and QTRAP- MS. In addition, the group has developed and implemented a semi-automated sample processing solution which ensures high sample throughput and reproducible results. Dedicated software solutions as well as the necessary expertise for lipid identification and quantification are available. In close consultation with the researchers, samples are processed and lipids extracted using standardized work instructions, and new and/or tailored analytical workflows will be developed to allow identification and quantification of relevant lipid species. A biosafety level SII laboratory is available.