timsTOF Pro

The timsTOF Pro instrument from Bruker is now installed at the NAPI core facilities at the Department of Biosciences, University of Oslo, and the Norwegian University of Life Sciences (NMBU).

Video: Bruker Daltonics

The timsTOF Pro enables enhanced speed, specificity and sensitivity, delivering superior quality data and facilitating the detection of more proteins at high confidence, even when starting sample amounts are low.

The timsTOF Pro with nanoElute. Image: Bruker Daltonics

Ion mobility: a new dimension for mass spectrometry

In addition to the three dimensions that have been used to identify peptide ions by mass spectrometry for many years (LC retention time, mass-to-charge ratio and MS/MS fingerprint) the timsTOF Pro incorporates an extra dimension of peptide separation – ion mobility. This is a powerful extension to mass spectrometry that delivers information about the three dimensional structure of an ion, and increases peak capacity and confidence in compound characterization. More specifically, the timsTOF Pro features ‘trapped ion mobility spectrometry’ (TIMS), in which ions are separated according to their mobility prior to entering the QTOF mass analyser.

Peptides ions elute sequentially from the TIMS tunnel, with more mobile ions released first (larger blue circles in illustration below), followed by lower mobility ions (orange and grey circles). This extra dimension of separation enables more comprehensive profiling of the protein content of samples.

Dual TIMS tunnel design of the timsTOF Pro instrument, with ion accumulation in the first tunnel and serial elution of ions in the second. Image: Bruker Daltonics

The mobility offset mass aligned (MOMA) feature of the TIMS technology means isobaric peptides (different peptides with the exact same mass) that cannot be distinguished by m/z are separated based on their different mobility. Furthermore, modifications such as phosphorylation can alter peptide mobility, so the separation of modified vs unmodified peptides is improved by TIMS. PTM-positional isomers – a common occurrence in MS – are not only detected but can be accurately quantified.

PASEF allows nearly 100% all peptides to be selected in each run

The dual TIMS tunnel design of the timsTOF Pro allows parallel accumulation of ions in the first tunnel and release of ions (dependent on their mobility) from the second tunnel (see illustration above).

Unlike a standard TIMS-MS/MS experiment in which just one precursor ion is selected in a single TIMS scan, the timsTOF Pro allows multiple precursor ions to be selected per scan. This process is called parallel accumulation serial fragmentation (PASEF), and allows nearly 100% usage of all peptide ions that enter the mass spectrometer. PASEF delivers effective MS/MS acquisition rates and sensitivity by interfacing the time-focusing power of the TIMS dimension with high-resolution QTOF technology.

The timsTOF Pro in action at the NMBU core facility. Photo: Morten Skaugen

The timsTOF platform measures ion mobility to assign collisional cross section (CCS) values for all compounds. Subsequently, CCS-aware analysis software improves the confidence of identifications and quantitation.

New analytical possibilities

In addition to the more commonly used data-dependent mode of MS data acquisition, the timsTOF Pro can perform data-independent acquisition with PASEF, called dia-PASEF. This allows the use of a pattern of m/z isolation windows within consecutive TIMS events, which increases the percentage of ions utilized and therefore facilitates deeper coverage of proteins within a sample and/or shorter run times.

The timsTOF Pro system is also compatible with cutting-edge accessories such as PaSER (Parallel database Search Engine in Real-time; see video below) which removes time-consuming data processing steps after completion of the MS run. With PaSER, as soon as the LC-MS run is complete, results files are searched and ready for downstream analysis.

Video: Bruker Daltonics

Further information on the timsTOF Pro and its potential for advanced proteomic analysis can be found on the Bruker website.

If you are interested in using the timsTOF Pro for your research project, contact the core facilities at UiO or NMBU directly, or alternatively email the NAPI Administrative Manager. Contact details are available on our contacts page.

Published Feb. 11, 2021 3:02 PM - Last modified Feb. 15, 2021 10:52 AM