Parallel Accumulation Serial Fragmentation (PASEF) Acquisition Method with the timsTOF Pro

Mass spectrometry (MS)-based proteomics has become a powerful technology that allows researchers to dig deeper and faster into the proteome. It can be utilized for the identification and quantification of thousands of proteins; however, the coverage of complete proteomes is still limited because of the speed, sensitivity and resolution of current mass spectrometers.

The timsTOF Pro uses the Parallel Accumulation Serial Fragmentation (PASEF) acquisition method to deliver tremendously high speed and sensitivity with the ability of reaching new depths in shotgun proteomics and phosphoproteomics, using low sample amounts.

The dual TIMS technology achieves a near 100% duty cycle for high sensitivity and high-speed shotgun proteomics. The innovative design allows for ions to be accumulated in the front section, while ions in the rear section are released in sequence depending on their ion mobility.

The timsTOF Pro is powered by PASEF, which is seen as the perfect fit for shotgun proteomics. It offers a sequencing speed higher than 100 Hz, without losing sensitivity or resolution. This is accomplished by synchronizing the quadrupole isolation mass window with the elution time of the specific peptide packages from the TIMS funnel.

The new orthogonal design of the ion optics gives it superior robustness. Its former unachievable level of robustness has been eliminated and shotgun proteomics performance is now uncompromised over large sample cohorts, as required for proteomics in clinical research.

MaxQuant/Perseus and PEAKS Studio enabled data processing now allows for digging into the 4th dimension. An open-file data format means that researchers can work directly with the raw data and use industry leading software. MaxQuant has been modified to manage 4-dimensional features in the space spanned by retention time, ion mobility, mass, and signal intensity.

This benefits the identification and quantification of peptides, proteins, and posttranslational modifications. PEAKS Studio combines de novo sequencing with traditional database searches and is enhanced to process timsTOF raw data.

We now know that the peptide mixtures are still extremely complex when analyzing them in two dimensions (retention time and m/z).

Adding one more dimension should in principle get us a long way ahead. In addition to the additional dimension of separation, the timsTOF Pro gives us extremely high speed and sensitivity to get deeper into the proteome, using less sample material.

Prof. Dr. Matthias Mann, Director Department of Proteomics and Signal Transduction, Max-Planck-Institute of Biochemistry, Germany.

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