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The facility has four mass Spectrometers, including three Thermo Orbitraps (Elite, Q-Exactive Plus and Q-Exactive HF) and a AB Sciex Voyager DE-STR MALDI-TOF (matrix assisted laser desorption ionization with a time-of-flight).  Ass mass spectrometers for peptides have high sensitivity (atmol to fmol), high resolution (7,500 to 240,000) and high mass accuracy (sub ppm to 10 ppm error).  The electrospray instruments are interfaced with two dimensional nanoflow liquid chromatographic systems (Eksigent Technologies, Thermo or Waters Corp.) for on-line separation and fragmentation for identifying peptides.  Off-line chromatography is also performed for fractionating complex samples and for specialized applications (e.g. quantifying proteins using iTRAQ, TMT, SILAC, AQUA, PSAQ or enrichment for post-translational modifications) on the Core's Agilent 1100/1200 capillary chromatography systems or BioRad Mini-Prep Cell gel electrophoresis systems.

Enzymatic digestions are performed in a laminar flow clean hood.  All proteomic data is stored on a mirrored 14TB NAS server in the Core's office suite were the director and staff consult and review data with Hopkins investigators.  Basic Sciences Network Office houses and maintains the Core's 10 core Xeon Mascot server for searching protein databases, 4TB NAS server for password protected transferring of large datasets and results files to investigators, and the backup mirrored 14TB NAS server with investigators proteomic data.

The Core also has two-dimensional (2D) gel electrophoresis and gel imaging equipment to support many sample preparation techniques.  Electrophoresis equipment include: IPGphors (3 units) and slab gel electrophoresis units (12 mini-gel) for protein separation by pI and size; a Typhoon laser scanner and Epson flatbed densitometer for image analysis; a pot picker for automated coring protein spots.  The laminar flow clean hood is also used for manual cutting out gel bands or spots.  The gel electrophoresis equipment as well as the MALDI-TOF are available to investigators for self-service work after successfully completing a training workshop.  Investigators can reserve Core self-service equipment via the Facility's iLab website Mass Spectrometry and Proteomics Core Facility

Mass Spectrometers

Orbitrap Fusion with ETD

The Orbitrap Fusion Tribrid mass spectrometer combines the quadrupole, Orbitrap, and ion trap mass analysis in a Tribrid architecture that delivers allows in depth analysis of protein and peptide samples. The high sensitivity aids in analyzing low-abundance, high-complexity, or difficult samples to accurately identify, quantify, and elucidate peptide and protein structures. The ETD capability facilitates mapping PTM modifaction sites (O-GlcNAc, phosphorylation, and other) by retaining the PTM on the modified amino acid during fragmentation.

Q-Exactive HF

The Q-Exactive HF is a hybrid quadrupole-Orbitrap high resolution mass spectrometer with an ultra-high-field Orbitrap analyzer.  This high mass accuracy (sub ppm), high resolution (up to 240,000 resolution at 200 m/z) mass spectrometer has scan speeds up to 18 Hz that allow monitoring up to 20 precursor masses per second.  Although all of our Orbitrap mass spectrometers have the speed, sensitivity and resolution for label free and multi-plex quantitative proteomics, the QE-HF is our primary instrument for analysis of complex protein extracts, discovery proteomics and quantification.

Q-Exactive Plus

The Q Exactive Plus is a hybrid quadrupole-Orbitrap high resolution mass spectrometer with high mass accuracy (sub ppm), high resolution (up to 140,000 resolution at 200 m/z) and scan speeds up to 12 Hz.  This mass spectrometer is our workhorse for routine protein identification, quantification and high resolution intact mass analysis of proteins (<60kDa).

 

Voyager DE-STR MALDI-TOF

The Voyager is a matrix assisted laser desorption ionization with a time-of-flight (MALDI-TOF) mass spectrometer capable of analyzing a wide variety of organic and inorganic compounds with masses ranging from 50 to 150,000 Daltons with high resolution (>10,000 at 2000 m/z).  This mass spectrometer is available to investigators after successfully completing a training workshop provided by the core and may reserve via the reservation tab.

HPLC Systems

The electrospray instruments are interfaced with nanoflow liquid high performance chromatographic systems, Waters NanoAcquity UPLC, Thermo Easy-nLC 1000, and Eksigent 2D NanLC, for on-line separation and fragmentation for identifying peptides.

The Core's Agilent 100/1200 capillary chromatography systems are used for offline fractionating of proteins and peptides for specialized applications such as affinity depletion of high abundance proteins body fluids, enrichment for post-translational modifications or quantifying proteins using iTRAQ, TMT, SILAC, AQUA, PSAQ technology is performed) or HPLVC systems (Agilent Technologies) for offline fractionation of using various LC based techniques such as strong cation exchange chromatography (SCX), anion exchange chromatography and macroporous reversed phase chromatography.  We are also equip to use antibody depletion columns for human samples (serum and urine).

 

Gel Equipment

Typhoon Imager

Our Typhoon imager 9400 is available for client use after a brief training. It has three laser options, red, blue and green. It is most useful for the detection of fluorescent or chemiluminescent dyes or antibodies.

Epson Professional Scanner

Our Epson Professional Scanner 750 is available to use after a short training session. It is able to scan gels (One and two dimensional SDS-PAGE) at high resolution.

Gel Electrophoresis Equipment

We use and maintain three iso-electric focusing units (Ge Healthcare) and 10 mini  gel electrophoresis units (Invitrogen). We also routinely use the Mini Prep Cell (Bio-Rad) for off-gel fractionation, while coupled to our fraction collector (Agilent) for protein separation and purification.

Location

The Johns Hopkins Mass Spectrometry and Proteomic Facility

725 N. Wolfe Street
Wood Basic Science Building, Room 504
Baltimore, MD 21205

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