Proteomics






Investigators using INBRE facilities, services or resources are required to cite NIH Grant Number P20 RR-016464 from the INBRE Program of the National Center for Research Resources in all publications and presentations. © Nevada INBRE 2006.
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Mass Spectrometry Services

 

The Mass Spectrometry Lab of the Nevada Proteomics Center offers a variety of services.  These include enzymatic protein digestion, MALDI/MS, LC/MS, and GC/MS.

 

Mass_Spec_Lab

 

Enzymatic Protein Digestion

Digest proteins (in-gel or in-solution) using a Genomic Solutions ProPrep workstation using either trypsin or a protease of your choice or combination of proteases.

 

Requirements:  visible spots by CBB staining (>100 fmol) or in appropriate buffer

 

MALDI/MS Analysis (Matrix Assisted Laser Desorption Ionization/Mass Spectrometry)

MALDI/MS analysis is performed using an Applied Biosystems 4700 Proteomics Analyzer with TOF/TOF Optics.  MALDI services include PMF, tandem MS, and accurate mass.

  • Acquire a PMF (Peptide Mass Fingerprint) from a digested protein that will be submitted to MASCOT for searching against a protein database for identification.
  • Perform tandem MS to obtain partial amino acid sequence using either protein database searches or a variety of De Novo software programs.
  • Obtain an accurate mass of a peptide or an average mass of an intact protein

Requirements: PMF and Tandem MS visible spots by CBB staining (>100 fmol)

Accurate mass >1 fmol, average mass >1 pmol protein and in appropriate solution

 

LC/MS (Liquid Chromatography/Mass Spectrometry)

LC/MS analysis is performed using a Thermo Finnigan LCQ Deca XP Plus that can be coupled to a Michrom Paradigm HPLC and a Dionex Probot.  LC/MS services include LC ESI-Ion Trap (Liquid Chromatography Electrospray Ionisation), LC/MALDI, Multi-Dimensional LC/MS, and direct infusion LC/MS.

  • Perform LC ESI of a digested protein and analyze using SEQUEST to find an identification, sequence information, or a PTM(post translational modification) such as phosphorylation. 
  • perform LC/MALDI (or 2D LC MALDI) of a digested protein or mixture of proteins and analyze by MALDI and MASCOT searching of protein databases for protein identification.
  • Multi-Dimensional LC/MS can be performed for either of the above techniques for better fractionation of peptides.

Direct Infusion ESI/MS obtain mass information on samples that do not require separation. 

 

Requirements: LC ESI >200 fmol on column; sample in an aqueous solution of  <5% acetonitrile, <0.01% (w/v) detergent concentration, <1mM salt

LC MALDI >100 fmol on column; sample in an aqueous solution of  <5% acetonitrile, <0.01% (w/v) detergent concentration, <10 mM salt

Direct Infusion ESI >200 fmol/ul (minimum 10 ul) in an aqueous solution

 

GC/MS (Gas Chromatography/Mass Spectrometry)

GC/MS analysis is performed using a Thermo Finnigan Polaris Q.  The GC/MS is used primarily for metabolomic studies of small nonpolar volatiles.

 

Requirements: sample in nonpolar solvent, >10 ng