MALDI Produced Ions Inspected with a Linear Ion Trap - Orbitrap Mass Analyzer
Kerstin Strupat, Thomas Moehring1
1Thermo Fisher Scientific, Bremen, Germany
Introduction
Matrix-Assisted Laser Desorption Ionization (MALDI) has being used as ionization technique for a large variety of thermo-labile (bio) molecules since the late 1980s and has been typically coupled to Time of Flight (ToF) instruments. Relative to MALDI ToF and MALDI ToF-ToF there is a significantly fewer number of publications in which MALDI is coupled to mass analyzers such as (q)QTof, ion trap or FTICRbased systems. While pulsed lasers are most straight forwardly coupled to ToF systems, as the laser pulse can be used as start signal for the ToF measure, the coupling to scanning mass analyzers is based upon different strategies(1). This presentation demonstrates coupling of a MALDI source to a hybrid linear ion trap - orbitrap mass analyzer.
Experimental
MALDI is coupled to a state of the art linear ion trap - orbitrap hybrid mass analyzer. The MALDI source is equipped with a Nitrogen Laser (Lasertechnologie, Berlin, Germany) which operates at 337.1 nm wavelength, 3 ns pulse duration and 60 Hz repetition rate. The beam diameter is about 80 * 100 μm2 on the sample plate. A xy-stage sample plate holder based on micro titer format can be moved with a reproducibility of 10 μm. Ions produced in the source are collected in a quadrupole and subsequently transferred to the linear ion trap and orbitrap analyzers, respectively. All linear ion trap as well as all orbitrap devices can be calibrated straight forward with the kit “MSCal4” provided by Sigma-Aldrich. Analytical scans are achieved under the conditions of Automatic Gain Control (AGC). Single (non-enzymatic) peptides were purchased from Sigma-Aldrich. Enzymatic digests were purchased from Michrom Bioresources Inc. USA. Enzymatically digested peptides were labeled with Tandem Mass Tags (TMT ®) following isobaric labeling approaches for quantitation. HCCA matrix as well as DHB matrix can be used for the analyses on the instrument.
Results
This presentation gives insight into the MALDI source design and shows results of the MALDI source coupled to a high resolution, accurate mass detector. Ions or fragment ions produced can be analyzed with orbitrap detection at a maximum mass resolution of > 100,000 @ m/z 400 with high mass accuracy. An accuracy of ≤ 2 ppm is achieved by internal mass calibration using lock mass functionality; using external mass calibration an accuracy of ≤ 3 ppm is obtained routinely. External mass calibration of the instrument is performed following the protocol of MSCal4 purchased from Sigma-Aldrich; the calibration kit provides a variety of different peptides as well as matrix molecules used for calibration of all devices of the hybrid mass spectrometer. The instrument equipped with a collision cell following the C-trap – allows performing Higher Energy Collisional Dissociation (HCD) besides Collision Induced Dissociation (CID) in the linear ion trap. The instrument and its capabilities are presented by demonstrating examples such as Protein ID approaches using Peptide Mass Fingerprinting (PMF) and MSMS analyses based upon data dependent decisions from sequence information, HCD is also used to produce sequence specific information plus diagnostic immonium and immonium-related ions. Data obtained form glyco-conjugates are interpreted which show the instrument’s potential to perform post-translational modification analysis. Stability of mass accuracy and signal-to-noise ratio for low samples loads on plates are demonstrated. Also demonstrated is the capability of using hot (such as HCCA) as well as cool (such as DHB) matrices, which opens the path for a variety of different demanding MALDI applications with accurate, highly mass resolved mass measurements.
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