The Proteomics and Metabolomics Core Facility of the Redox Biology Center (PMC-RBC), Department of Biochemistry, supports Faculty, scientists, clinicians and students not only within the RBC, but also the University of Nebraska system and other universities and industries. The Core provides all the tools of modern functional proteomics and metabolomics. The facility is equipped with chromatography and mass spectrometry-based state-of-the-art technologies for proteomics and metabolomics (Clinical as well as non clinical), personalized experimental design consultation and individualized bioinformatics support.
We are interested in collaborations with faculty and scientists in who are interested in studying problems of biomedical importance such as clinical proteomics and metabolomics, biomarker discovery and validation. We also have the capability of analyzing microbial and plant samples for proteomics and metabolomics analysis. Please contact Javier Seravalli for consultation and services provided.
General Services Offered by the RBC Proteomic and Metabolomic Core Facility:
- Small molecule exact mass determination or quantitation using positive or negative ion mode ionization via LC-MS/MS
- Proteomics: Protein identification via standard bottom-up LC-MS/MS analysis coupled with MASCOT and SEQUEST database searches
- Shot gun proteome analysis of biological samples.
- Biomarker discovery from biological fluid
- Targeted Metabolomic analysis of up to ~256 compounds utilizing LC-MRM-MS methodologies.
- Protein complex isolation and identifying interacting partners and its quantitation.
- Global PTM analysis and quantitation
- Protein ID from gels stained with Coomassie Blue or Silver Stains.
- De novo peptide sequencing by tandem mass spectrometry.
- Confirmation of mutations in protein via LC-MS/MS and Intact Mass analysis
- Customized sequence search of in-house produced proteins, not yet available in database
- Post translational modifications (phosphorylation, sumoylation, ubiquitination, oxidation, etc.)
- Determination of oxidation state of cysteine (disulfide bonds) by means of blocking tags.
- Intact proteins and peptides mass determination.
Proteomics
- Redox proteomics
- High throughput targeted SRM/MRM peptide/proteomic assay development
- Multiplex assays for biomarker research and clinical application
- Organelle proteomics
- Membrane proteomics
- Signaling pathway analysis
- Protein complex analysis
- Peptide/nucleotide/protein-ligand interactions
- Protein sequencing and identification
- Characterization of protein modifications such as phosphorylation, oxidation, methylation
- Antibody-based multiplexed assay (coming soon)
Metabolomics:
Discovery metabolomics: UHPLC/MS based routine shot gun quantitative metabolite profiling from complex biological samples (clinical or non clinical).
Targeted metabolomics: HPLC/MS Multiple reaction monitoring (MRM or SRM) analysis of metabolite, small molecules, lipids, relative and absolute quantitation (labeled and label free) of targeted group of metabolites from plasma, serum, urine, CSF, tissue and other clinical and non clinical biological samples.
Instruments:
This mass spectrometer can operate in 12 different scanning modes for structural and quantitative workflows. The first and third quadrupoles have a resolution of > 0.4 Da with scan rates up to 4000 m/z units per second (Trap mode). The instrument is equipped with the standard ESI, APCI and nanoSpray nanosources. It is interfaced with 2 HPLC separate stacks from Agilent: an LC-1200 series with column thermostated compartment, thermostated capillary Autosampler with 96-well plate capability and capillary quad-pump; an LC-1200 series HPLC with standard binary pump, thermostated autosampler and diode-array detector.
This instrument can be used for bottom-up proteomics, intact protein mass analysis, peptide analysis, small molecule characterization and quantification, targeted MS/MS analysis of peptides (proteomics), protein PTM characterization. The Velos Pro is a low- and medium-resolution linear ion-trap instrument that features a dual chamber, dual pressure ion analyzer that is suitable for ion-trap scanning and fragmentation without the requirement of time consuming ion selection in the ion trap. Scan rates of up to 60,000 amu per sec. at 0.7 Da resolution are routinely achieved in datasets from this instrument. The Velos Pro is interfaced to a Dionex Ultimate nanoRSLC system, capable of flow rates from 0.1 to 2500 uL/min and equipped with a pump with upper pressure limit of 800 bars. The ETD accessory is employed in the characterization of post-translational modifications.