MALDI TOF analysis
1) Molecular mass determination by MALDI TOF
2) Protein identification
The proteins previously purified or resolved on a gel can be identified by peptide mass fingerprinting (PMF). The process includes: protein digestion by tripsin, peptide extraction when required, sample preparation, analysis by MALDI TOF, data processing and database search by Mascot search engine.
The sample can also be analyzed by peptide fragment fingerprinting using MALDI TOF/TOF.
Liquid chromatography on-line coupled to tandem mass spectrometry platforms.
Protein bands or purified proteins can be digested by tripsin either in gel or in solution and resulting peptides will be resolved by a nano-flow liquid chromatography system, coupled to a tandem mass spectrometer (nLC MS/MS). The Proteomics Core Facility at CIC bioGUNE works with two high-end nLC MS/MS systems:
- A Nano-Aquity Ultra Performance liquid chromatography system (UPLC, Waters) is coupled to a Synapt G2S, which combines revolutionary high performance StepWave™ ion optics, Quantitative Tof (QuanTof™) and High Definition MS™ technologies.
- A Nano-Aquity Ultra Performance liquid chromatography system (UPLC, Waters) is coupled to a LTQ-Orbitrap XL, which combines a linear ion trap mass spectrometer and the Orbitrap mass analyzer.
Since the performance of these instruments is comparable, samples will be run in either of the two systems.
Liquid chromatography off-line coupled to MALDI TOF/TOF analysis
Differential in-gel-electrophoresis (DIGE)
- The Agilent 1200 Series Nanoflow LC system is coupled to a Probot that will deposit the sample onto a 384 MALDI target. Autoflex III smartbeam is the instrument to perform the MALDI TOF/TOF analysis.
For DIGE type of experiments, please contact the head of the unit.
Note: for 1D bands or 2DE gel spots we highly recommend providing an image of the gel; this would supply additional information for correct protein identification.