About the Reseach Resource
Resource instrumentation includes a comprehensive range of separation and mass spectrometry capabilities for conducting leading-edge bioanalytical research. Separation capabilities are unparalleled with the ability to rapidly implement nearly any desired methodologies in a matter of days. Mass spectrometry assets include the latest and most powerful mass spectrometers available on the market today. Many PNNL-developed technologies such as the electrodynamic ion funnel, devices for external ion accumulation, and ultra-high-sensitivity electrospray ionization (ESI) sources take these resources beyond what is available on the market resulting in capabilities truly unique to this facility. Instrumentation is described in detail below.
A diverse collection of 40 HPLC systems is available to meet the separation needs of any project based on sample type, quantity, and complexity. Included are ten commercial nano-flow LC systems for more routine types of work and 30 custom systems built around ISCO pumps, Agilent nano-pumps, or a combination of both. These custom LC systems provide unique and sophisticated advanced biological separation capabilities through custom software control, operating pressures up to 20k psi, easily modified valve configurations, and custom packed LC columns of any separation type (reversed-phase, ion-exchange, size-exclusion, etc.) and dimension (down to 10 um i.d.). The flexibility of the control software and valve configurations allows for rapid reconfiguration and automation to take immediate advantage of new separation techniques rapidly evolving in the proteomics field.
Four Thermo Scientific LTQ mass spectrometers. These instruments are based around a linear ion trap with a dual electron multiplier based detection system, providing excellent sensitivity and scan speed. A single CID MS/MS spectra is acquired approximately every 0.3 seconds when configured for data dependent analysis of the top 10 ions from a precursor scan, our typical operating mode for this instrument. Dynamic exclusion in combination with data dependant m/z sampling provides for a significantly better survey of species present in a sample by discriminating against previously observed ions.
One Thermo Scientific LTQ-ETD mass spectrometer. This instrument is identical to the LTQ mass spectrometer except that an Electron Transfer Dissociation (ETD) source is included for introduction of a reagent ion into the linear ion trap, providing an alternative to CID fragmentation. ETD has been shown to be a superior fragmentation mechanism for sequencing modified peptides and proteins (i.e., post-translational modifications).
Three Thermo Scientific TSQ triple quadrupole mass spectrometers (two Quantum Ultra's and one Vantage) Three research grade triple quadrupole instruments are available, and are primarily used for high throughput quantitative proteomics and metabolomics measurements applying multiple-reaction-monitoring (MRM) strategies.
Three Thermo Scientific LTQ-Orbitrap mass spectrometers. An LTQ mass spectrometer coupled with an Orbitrap mass analyzer that combines all the features of the LTQ with the high resolution and mass accuracy detection capabilities of an Orbitrap. All ion manipulation takes place in the linear ion trap but detection can take place in either the LTQ or the Orbitrap. This combination allows for highly accurate mass determination in the precursor scan using the Orbitrap and true parallel MS/MS data acquisition in the LTQ. In our standard operating mode, a single Orbitrap precursor scan from m/z 400-2000 and resolution 100k and the top 6 MS/MS spectra from the LTQ are acquired approximately every 2 to 2.5 seconds (variability due to ion trap fill time). This provides sufficient sampling speed for defining chromatographic peaks when doing quantification work and a MS/MS sampling rate approaching what can be achieved on a stand-alone LTQ.
Three Thermo Scientific Exactive Orbitrap mass spectrometers. These new instruments are used for high resolution, high-throughput LC-MS measurements applying the PNNL-developed accurate mass and time (AMT) tag approach. While these instruments have no MS/MS capability, they do provide amongst the best sensitivity for high accuracy LC-MS work.
Four Thermo Scientific LTQ-Orbitrap Velos mass spectrometers. These new instruments combine the proven mass accuracy and ultra-high resolution of the Orbitrap mass analyzer, with the increased sensitivity and improved cycle time of the LTQ Velos ion trap. Each instrument has dual-pressure linear trap, HCD collision cell, and ETD ion source providing multiple fragmentation techniques for optimal structural characterization.
One 7-tesla Thermo Scientific LTQ-FTICR mass spectrometer.
One 12-tesla Bruker solariX FTICR mass spectrometer.
One 15-tesla Bruker solariX FTICR mass spectrometer with dual ESI/MALDI source.
Two ESI-IMS-TOF mass spectrometers. The instruments incorporate an ion mobility spectrometer (IMS) with an Agilent orthogonal time of flight (TOF) mass spectrometer. With the inherent high speed of gas phase separation in IMS and the well designed interface between IMS and TOF MS using PNNL’s electrodynamic ion funnel technology, the instruments can perform sample analysis with extremely high throughput and sensitivity.
One Agilent Triple Quadrupole mass spectrometer. This is a prototype triple quadrupole mass spectrometer equipped with latest LC and ESI interface technologies including Chip cube nano flow LC system and multicapillary inlet/iFunnel interface for high sensitivity. The instrument is especially suitable for quantitative sample analysis with high sensitivity and broad dynamic range.
Two Agilent Single Quadrupole mass spectrometers. These instruments are used for development of ESI interface technologies to improve analytical sensitivity and limits of detection. Currently, a low pressure ESI source and interface, utilizing electrodynamic ion funnels, is being refined and tested for use in routine proteomic analyses.
One Thermo Scientific TSQ Quantum Ultra GC/LC-MS triple quadrupole mass spectrometer. This research grade triple quadrupole instrument is capable of interfacing with both gas and liquid chromatographs. The GC-MS mode is well suited for targeted analyses of volatile small molecules, while the LC-MS mode is suitable for high throughput quantitative proteomics and metabolomics measurements.
Instrument support and maintenance. All of the above-listed spectrometers are on a common computer network with direct high-speed access to the EMSL data storage and archive facility, a multi terabyte storage system that allows large numbers of data files to be archived indefinitely. Researchers have full access to both the resources and support personnel in the EMSL Instrument Design Laboratory (IDL) to support instrument development and repair.
Our instrumental resources also include three home-built planar and one cylindrical commercial (Thermo Fisher, San Jose, CA) FAIMS units.
All of the aforementioned spectrometers are on a common computer network with direct high-speed access to the EMSL data storage and archive facility, a multi-terabyte storage system that allows large numbers of data files to be archived indefinitely. Collaborators are able to access and download their data via the Internet and have access to other data at the discretion of the Resource staff and other collaborators.
The EMSL laboratories also include facilities for sample preparation and storage, chemical synthesis and derivatization, one-dimensional (1-D) and 2-D polyacrylamide gel electrophoresis, capillary electrophoresis (CE) and ESI-MS, and related instrumentation development and experimentation, purifications, synthesis, and biological research activities. Four CE systems are available, including three Crystal systems that are optimal for MS interfacing. The CE systems have electrical isolation and interlock and equipped with robotic injectors and a Pharmacia PhastSystem integrated planar electrophoresis system.
Other equipment includes an Applied Biosystems 270A integrated instrument and data system for CE with UV-Vis/absorbance detection and a Beckman P/ACE System 1000 CE instrument extensively modified for application with an ESI interface for mass spectrometry.