Omics in 4D
TIMS adds 4th dimension of separation
Mass spectrometry (MS)-based proteomics has become a powerful technology for the identification and quantification of thousands of proteins. However, the coverage of complete proteomes is still very challenging due to the limited speed, sensitivity and resolution of current mass spectrometers.
The timsTOF Pro uses the Parallel Accumulation Serial Fragmentation (PASEF®) acquisition method to provide extremely high speed and sensitivity to reach new depths in shotgun proteomics and phosphoproteomics, using low sample amounts.
Trapped Ion Mobility Spectrometry (TIMS) is first and foremost a separation technique in gas phase, which resolves sample complexity with an added dimension of separation in addition to HPLC and mass spectrometry, increasing peak capacity and confidence in compound characterization. Equally as important, the TIMS device also serves to accumulate and concentrate ions of a given mass and mobility, enabling a unique increase in sensitivity and speed along with the additional dimension of separation.
Achieve near 100% duty cycle for shotgun proteomics
A near 100% duty cycle for high sensitivity, high speed shotgun proteomics can now be achieved with the dual TIMS technology. The novel design allows for ions to be accumulated in the front section, while ions in the rear section are sequentially released depending on their ion mobility. This process is called Parallel Accumulation Serial Fragmentation, or PASEF®.
The second generation dual TIMS analyzer is optimized for shotgun proteomics needs. Due to its unique geometry ions are released dependent on their mobility from the second section of the TIMS analyzer while the further incoming ions can be accumulated in parallel in the first part of the TIMS analyzer. With the parallel accumulation technology a duty cycle of nearly 100% is achieved resulting in nearly no ion loss.
In the timsTOF Pro an advanced segmented quadrupole mass filter is used for high ion transmission and isolation efficiency. The quadrupole mass position is synchronized with the elution times of the specific ions from the TIMS analyzer. Due to its ultra-high mass position switching time (< 1 ms) it enables the best performance for the PASEF® method.
With speed in mind, Bruker experts redesigned MS/MS technology to meet the requirements of shotgun proteomics. Peptide ions are separated using trapped ion mobility spectrometry, eluted (~ 100 ms) and detected in the QTOF, generating the TIMS MS heat map. In the PASEF® method the same TIMS separation is used with the quadrupole isolating a certain ion species during its elution and immediately shifting to the next precursor. Parent and fragment spectra are aligned by mobility values. With the Parallel Accumulation Serial Fragmentation (PASEF®) Technology >100 Hz sequencing speed can be achieved. With the PASEF® method the MS/MS spectra quality of the low abundant peptides can be increased by selecting them several times.
PASEF®: the perfect fit for shotgun proteomics
The timsTOF Pro powered by PASEF® offers sequencing speed >100 Hz without losing sensitivity or resolution. This is achieved by synchronizing the quadrupole isolation mass window with the elution time of the specific peptide packages from the TIMS funnel.
The only PASEF®-enabled mass spectrometer
Bruker has introduced the TIMS (Trapped Ion Mobility Spectrometry) technology in 2016 as a revolutionary ion mobility technology, achieving unprecedented levels of ion mobility resolution in an extremely compact device. The second generation of dual TIMS analyzer now enables PASEF® (Parallel Accumulation Serial Fragmentation) that is setting new standards of uncompromised speed, sensitivity and resolution for shotgun proteomics.
Superior robustness, no cleaning required
Many MS instruments used for shotgun proteomics require biweekly or monthly cleaning when run 24 hours a day on large sample cohorts, and performance degradation is noticeable over even shorter time periods. The superior robustness of the timsTOF Pro means that the instrument can be run 24/7 over many days or weeks without noticeable loss of sensitivity or other performance metrics.
The timsTOF Pro allows the analysis of hundreds of samples with minimal sample load (<200 ng) at sequencing speeds exceeding 100 Hz. All of this with uncompromised proteomic depth. This has changed the way proteomics is done, but it also unleashes a new constraint - data analysis. Modern researchers must process hundreds of samples of information dense content with the timsTOF Pro. For most pipelines this creates a bottleneck at the data analysis step. Bruker is innovating this step by introducing real-time database search capabilities called PaSER (Parallel database Search Engine in Real-time) which removes the data analysis hurdle. Using PaSER, as soon as a LC-MS run is completed, results are in hand or Run & Done.
MaxQuant/Perseus and PEAKS Studio data processing
An open-file data format allows researchers to work directly with the raw data and use industry leading software. MaxQuant has been adapted to manage 4-dimensional (4D) features in the space spanned by retention time, ion mobility,
mass, and signal intensity which benefit the identification and quantification of peptides, proteins, and posttranslational modifications. PEAKS Studio combines de novo sequencing with traditional database searches and is optimized for processing timsTOF raw data.
prm-PASEF®: enabling high-throughput, high sensitivity targeted proteomics
Enabling high-throughput, high sensitivity targeted proteomics. The prm-PASEF® acquisition method has been developed to translate the advantages of the parallel accumulation serial fragmentation (PASEF®) acquisition strategy to the targeted proteomics field.
Proteomic Interrogation of Primary Insulin-secreting Pancreatic β-cells
The combination of integrated ion mobility with warp-speed MS/MS acquisition on the timsTOF Pro paves the way for deep, molecular level understanding of β-cell physiology and potential new targetable pathways for diabetes.
High throughput 4D-Proteomics™ – Application of dia-PASEF® and the Evosep One for short gradients
The timsTOF Pro offers a combination of two unique technologies, namely a 4th dimension provided by Trapped Ion Mobility Spectrometry (TIMS) to enhance ion separation and sensitivity and Parallel Accumulation Serial Fragmentation (PASEF®)
to improve ion utilization efficiency and data acquisition speed.
"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 and using less sample material.”
For Research Use Only. Not for use in clinical diagnostic procedures.