UAMS Proteomics Core (proteomics@uams.edu)

Overview of Services

The UAMS Proteomics Core provides a full range of services required for protein characterization by mass spectrometry, including protein identification, mapping of post-translational modifications, global proteomics, and quantitative comparison of proteins in biological samples. Please contact core personnel at proteomics@uams.edu to discuss how the core can support your research.

 

 

Proteomics Core Equipment

 

Orbitrap Fusion Mass Spectrometers

The Thermo Orbitrap Fusion Lumos system is the latest generation of the state-of-the-art Orbitrap mass spectrometry technology installed at UAMS in July 2017. The Proteomics Core also operates two Orbitrap Fusion Tribrid instruments installed in 2015. The Orbitrap Fusion systems combine a quadrupole mass filter with Orbitrap and linear ion trap mass analyzers in a novel configuration that maximizes ion transfer efficiency, allowing flexible combinations of different fragmentation techniques, scanning modes, and mass analyzers. The quadrupole mass filter allows precursor ion selection, enabling parallel use of the mass analyzers and synchronous precursor selection for MS3 analysis. An ion-routing multipole serves as a higher-energy collisional dissociation (HCD) cell and also transfers ions efficiently between the Orbitrap and linear ion trap mass analyzers, allowing use of either mass analyzer in combination with any ion fragmentation technique. The ultra-high-field Orbitrap mass analyzer has a resolving power of up to 450,000 FWHM and 15 Hz scan rate. A linear ion trap performs collision-induced dissociation (CID) and electron-transfer dissociation (ETD).

 

Orbitrap Velos Mass Spectrometer

The Thermo LTQ Orbitrap Velos is an earlier generation of the Orbitrap technology that has been maintained and operated by the Proteomics Core since 2011. This instrument system combines the rapid data acquisition capability of the LTQ linear ion trap with the high resolution of the Orbitrap. This configuration allows extremely rapid in-depth proteomic analysis at high mass accuracy and supports quantitative isotope-based proteomic profiling. The Velos configuration is also capable of HCD and ETD complementary to CID, which is available with earlier LTQ-based instruments.

 

Nanoflow Ultra Performance Liquid Chromatography

Each of the mass spectrometers described above is interfaced with a Waters nanoAcquity or Thermo Easy-nLC 1200 nanoflow UPLC system, including a binary solvent manager and a sample manager. Precise chromatography is achieved by these systems through low detector cell volume, minimized tubing volumes and specialized fittings. The binary solvent manager and injector can sustain pressures up to 15,000 psi and can generate high-pressure gradients at nanoliter flow rates with minimal delay. The sample manager can accommodate two 96-well plates in a microtiter plate format.

 

UltiMate 3000 Binary Analytical Liquid Chromatography System

The Thermo UltiMate 3000 System is capable of precise and efficient offline fractionation of complex protein digests. This system is used in the Proteomics Core workflow to fractionate peptides at basic pH in order to complement the acidic pH separation performed during liquid chromatography-mass spectrometry/mass spectrometry (LC-MS/MS) analysis. The UltiMate 3000 can sustain pressures as high as 9000 psi and flow rates as low as 50 nL/min. The system includes an analytical autosampler and fraction collector to allow efficient processing of large numbers of samples, as well as a UV detector to monitor peptide elution and prevent cross-contamination.

 

Computers and Software

The core maintains eight Dell Precision workstations, each with solid state hard drives and 128 GB RAM, supporting a full range of data analysis software, including an in-house Mascot Server (Matrix Science), several MaxQuant (Max Planck Institute) installations, Scaffold Q+S, PTM and perSPECtives (Proteome Software), and PEAKS Studio (Bioinformatics Solutions). The core has 150 TB of dedicated data storage space to maintain secure backups of all data files.

 

 

Proteomics Core Services

 

The UAMS Proteomics Core offers comprehensive support from sample preparation through data analysis at reasonable cost with rapid turnaround. Please contact core personnel before starting a new experiment to ensure that sample prep is MS compatible. Services and fees can be adapted to fit the requirements of specific projects. All fees below include data analysis. Turnaround times are estimates and may vary for specific projects. Data are provided in Scaffold files. A free viewer version of Scaffold is available (http://www.proteomesoftware.com/products/scaffold/download/).

 

Protein Identification – Single Gel Band

A protein band excised from a standard SDS-PAGE gel is identified by tandem mass spectrometry. Any band visible by Coomassie staining can be identified. Please consult core personnel before using other gel stains.

Service includes: in-gel trypsin digestion, MS/MS analysis, Mascot database search

Cost: $15 per gel band

Estimated Turnaround: <2 weeks

Data file format: Scaffold

 

PTM Identification – Single Gel Band

Protein modifications are identified and localized by tandem mass spectrometry from an SDS-PAGE gel band. A minimum of 100 ng protein is required. Success depends on protein sequence coverage and context, stoichiometry of modification, and PTM stability.

Service includes: in-gel trypsin digestion, MS/MS analysis, Mascot database search, manual validation

Cost: $30 per gel band

Estimated Turnaround: <3 weeks

Data file format: Scaffold or Scaffold PTM

 

GeLC Analysis – Global Proteomics

An entire SDS-PAGE gel lane is analyzed by tandem mass spectrometry to identify hundreds to thousands of proteins. For cell lysates or tissue homogenates, 25 ug total protein per lane is recommended. For immunoprecipitations or affinity purifications, the entire sample is usually loaded. The core offers an SDS-PAGE gel service (NuPage 4-12% Bis-Tris minigel), or external gels can be submitted (except for Lumos analysis). Gels with significant streaking, overloading, or other resolution problems may not be accepted. Please consult core personnel regarding recommended gels and protocols.

Service includes: SDS-PAGE separation, in-gel trypsin digestion, MS/MS analysis, Mascot spectral counting

Cost: $360-$720 per gel lane (plus $60 per gel for SDS-PAGE service)

Estimated Turnaround: 2-4 weeks

Data file format: Scaffold

 

Basic pH Peptide Chromatography – Global Proteomics

Peptides from complex samples are fractionated prior to tandem mass spectrometry to identify and quantify thousands of proteins. This technique is more sensitive than GeLC (described above), but sample prep requirements are more stringent. Cell pellets or lysates, tissue homogenates, or other protein fractions or preparations can be submitted. Please consult core personnel regarding sample prep guidelines before starting an experiment.

Service includes: cell lysis, trypsin digestion, basic pH peptide separation, MS/MS analysis, MaxQuant label-free quantification

Cost: starting at $525 per sample

Estimated Turnaround: 2-4 weeks

Data file format: Scaffold or Scaffold Q+S

 

Serum Profiling – Global Proteomics

Serum or plasma samples are analyzed directly to identify and quantify hundreds of proteins by tandem mass spectrometry. As little as 25 ul of serum/plasma is required. Results are strongly dependent on initial collection and handling protocol, so please consult core personnel beforehand if possible.

Service includes: albumin depletion, trypsin digestion, basic pH peptide separation, MS/MS analysis, MaxQuant label-free quantification

Cost: starting at $420 per sample

Estimated Turnaround: 2-4 weeks

Data file format: Scaffold or Scaffold Q+S

 

Phosphorylation Profiling – Global Proteomics

Phosphorylated peptides are enriched from complex samples to identify thousands of phosphorylation sites by tandem mass spectrometry. Cell pellets or lysates, tissue homogenates, or other protein fractions or preparations can be submitted. Please consult core personnel regarding sample prep guidelines before starting an experiment.

Service includes: cell lysis, trypsin digestion, phosphopeptide enrichment, MS/MS analysis, MaxQuant label-free quantification

Cost: starting at $345 per sample

Estimated Turnaround: 2-4 weeks

Data file format: Scaffold or Scaffold Q+S

 

SILAC – Quantitative Global Proteomics

Metabolic labeling of cultured cells is used to quantify thousands of proteins identified by tandem mass spectrometry. Cell culture and SILAC labeling must be done prior to submission of cell pellets. Please consult core personnel regarding sample prep guidelines before starting an experiment.

Service includes: cell lysis, trypsin digestion, basic pH peptide separation, MS/MS analysis, MaxQuant SILAC quantification

Cost: starting at $540 per H/L sample pair

Estimated Turnaround: 2-4 weeks

Data file format: Scaffold Q+S

 

TMT – Quantitative Global Proteomics

Isobaric tandem mass tags are incorporated at the peptide level to quantify thousands of proteins identified by tandem mass spectrometry. TMT reporter ions are generated by synchronous precursor selection (MultiNotch) and subsequent MS3 fragmentation of MS/MS ions. TMT labeling is done by the core, so unlabeled cell pellets or lysates, tissue homogenates, or other protein fractions or preparations can be submitted for analysis. Please consult core personnel regarding sample prep guidelines before starting an experiment.

Service includes: cell lysis, trypsin digestion, TMT labeling, basic pH peptide separation, MultiNotch MS3 analysis, MaxQuant TMT quantification

Cost: starting at $1500 for 6 samples, $1920 for 10 samples

Estimated Turnaround: 4-6 weeks

Data file format: Scaffold Q+S

 

Shipping Address

UAMS Proteomics Core

Biomedical Research Center, Bldg 1, Rm B421

325 Jack Stephens Dr

Little Rock, AR  72205

 

Contacts

Name Role Phone Email Location
Alan Tackett
Director, IDeA National Resource for Proteomics
 
(501) 686-8152
 
ajtackett@uams.edu
 
Biomed I, B405E
 
Rick Edmondson
Co-Director, UAMS Proteomics Core
 
(501) 766-3665
 
rdedmondson@uams.edu
 
WPRCI, 9121
 
Sam Mackintosh
Co-Director, UAMS Proteomics Core
 
(501) 526-6165
 
sgmackintosh@uams.edu
 
WPRCI, 9111
 
Lisa Orr
Research Assistant
 
(501) 686-5954
 
orrlisam@uams.edu
 
Biomed I, B406
 
Renny Lan
Data Analyst
 
(501) 526-5238
 
slan@uams.edu
 
WPRCI, 9110
 

Services

Name Description