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Analytischer Service

Proteomics

Our proteome service includes optimization of sample extractionprotein quantification and sample clean-up.

1- and 2-dimensional gelelectrophoresis in small, medium and large scale. The service includes staining (Coomassie, silver, Sypro-Ruby, ProQ Diamond, DIGE etc.) of gels, scanning, electroblotting, and western-blott analyses with ECL.

For image analysis we can compare 2D-gels with the software packages ProteomWeaver and PDQuest.

For spot-picking data can be directly transfered to the ExQuest Spot-Cutter, which picks up to 600 spots/h. Fluorescently labelled proteins can be directly visualized. Furthermore, the ExQuest is also capable to cut membranes (NC and PVDF) for subsequent on-membrane proteolytic digest.

Proteolytic in-gel digest and subsequent MALDI-TOF-MS peptide  fingerprints and MS/MS analyses combined with database searches can be done for protein identification.

The equipment comprises an isoelectric-focusing system from Amersham Bioscience, two electrophoresis systems (Hoefer Dalt and Hoefer 600), an ExQuest Spot-Cutter (BioRad), a Hamilton liquid handling system for automatic proteolytic in-gel digest and a Bruker Ultraflex TOF/TOF MALDI instrument.

Full data integration with ProteinScape (Bruker Daltonics). ProteinScape is the central bioinformatics platform for storage and processing of MS data. It organizes all relevant data for all types of proteomics projects – including LC-data, gel data, mass spectra, process parameters, search results and quantitative data. ProteinScape provides researchers with sophisticated tools for the analysis and evaluation of proteomics and glycomics data. Collaboration partners have direct IP-based access to all primary data from their desk after installing a client program.

 

N-Terminal Edman sequencing

The amino acid sequence of proteins and peptides can be determined by Edman sequencing. If the N-terminus is not blocked, this is still the method of choice for identification of the N-terminus.

We perform Edman sequencing with the Procise from Applied Biosysstems. Samples should be as pure as possible. We can analyze either electroblotted proteins (on PVDF) or dissolved samples. For optimal results we nedd about 10 pmol.

 

HPLC separation techniques

We offer HPLC methods for isolation of proteins, peptides and other biomolecules.

The methods include gelpermeation-, ion-exchange-, normal- and reverse-phase-chromatography.

 

LC-MALDI coupling

For LC-MALDI analyses we have a nano-LC systems (with a Probot fraction collector which allows direct spotting of complete LC-runs on a MALDI-TOF target to obtain higher sequence coverages compared to total digest MS analysis.

 

Analysis of posttranslational modifications

We have methods to localize and analyze phosphorylation sites, O- and N-glycans and disulfide linkages.