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Mass Spectrometry Lab

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

Enzymatic Protein Digestion

Before being analyzed by mass spec, most proteins must be broken into smaller fragments. This is accomplished by digestion with a protease, usually trypsin, but may be another proteases of your choice.

Requirements: >100 fmol protein, may be a protein in solution, a gel slice or spot that is visible by coomassie staining.

LC/MS (Liquid Chromatography/Mass Spectrometry)

• LC/MS analysis is performed using a Thermo LTQ Orbitrap XL with ETD coupled to a Michrom Paradigm MDLC and Michrom CaptiveSpray.
• Protein Identification from excised 2D PAGE or DIGE gel plugs are performed by manual injection and using a quick LC gradient of 15 minutes. Data analysis is performed on our SORCERER™ server using SEQUEST®. Data output is provided to the user as a Scaffold output file.
• Protein Identification from excised 1D PAGE gel plugs are performed by manual injection and using an LC gradient of 60- 90 minutes. Data analysis is performed on our SORCERER™ server using SEQUEST®. Data output is provided to the user as a Scaffold output file.
• Protein Identification from complex samples are performed using multi-dimensional LC. The first dimension is typically strong cation exchange (SCX) followed by reversed phase LC. The first dimension is carried out on a Michrom Paradigm MDLC and fractions are collected using a Dionex Probot. SCX fractions are then injected by autosampler onto the Orbitrap using an LC gradient of 3-4 hours. Data analysis is performed on our SORCERER™ server using SEQUEST®. Data output is provided to the user as a Scaffold output file.

Requirements: >50 fmol of tryptic digest on column; sample in an aqueous solution of <5% acetonitrile, <0.01% (w/v) detergent concentration.

MALDI/MS Analysis (Matrix Assisted Laser Desorption Ionization)

MALDI analysis is performed using an Applied Biosystems 4700 Proteomics Analyzer with TOF/TOF Optics. MALDI services include Protein Identification and accurate mass determination.

• Protein data from a digested protein are submitted to MASCOT® for searching against a protein database for identification.
• Accurate mass of a peptide or an average mass of an intact protein can be obtained

Requirements: Protein Identification - visible spots by CBB staining (>100 fmol); accurate mass - >10 fmol; average mass >1 pmol protein and in appropriate solution.

GC/MS (Gas Chromatography/Mass Spectrometry)

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

Requirements: sample in nonpolar solvent, >10 ng.