Real-time results = efficiency
Break the data analysis bottleneck by integrating real-time database search and smart acquisition provided by PaSER
Massive parallelization with GPU power
PaSER Data Review
TIMScore provides the greatest number of PSM, peptide and protein identifications by enabling the CCS dimension for true 4D-Proteomics.
The PaSER search algorithm is run as normal, combined with comparison of the predicted and measured CCS values and calculating a TIMScore for the top 5 peptide candidates for each spectra.
The TIMScore benefit is realized during the peptide-validation and False Discovery Rate (FDR) estimation steps. In a non CCS-enabled algorithm, only two dimensions can be utilized to estimate the FDR rate, and so a discriminate line is fit to a 1% error (Panel A) to distinguish forward and reverse peptide candidates. With TIMScore, and the extra CCS dimension, the peptide-candidates can be vectorized in 3 dimensions (Panel B) allowing a discriminate contoured plane to be applied to achieve the same 1% error.
Applying a discriminate plane provides increased accuracy and precision, helping to validate formerly poorly scoring PSMs in the standard two dimensions.
PaSER 2023 is capable of processing dia-PASEF workflows utilizing a customized version of the popular DIA-NN (DIA by Neural Networks) software from the Lilley, Rasler and Demichev labs. TIMS DIA-NN enables reliable, robust, and quantitatively accurate large-scale experiments. PaSER utilizes the same stream mechanism for dia-PASEF workflows, streaming in real-time MS and DIA frames from the acquisition PC to the PaSER box. These dia-PASEF data are processed and stored on the PaSER box. At the end of the acquisition a spectral library search is triggered, and the results recorded. TIMScore powered DDA search results can be easily utilized to build spectral libraries or spectral libraries can be imported from other popular tools. At the end of the project the users can trigger match-between-runs analysis to view a quantitative profile across projects. This provides users an integrated environment for PASEF and dia-PASEF data analysis.
"Our ongoing collaboration with Bruker to tailor DIA-NN to a streamlined
processing tool for dia-PASEF data with a CCS focus has been really
rewarding. It simplifies and accelerates identifying and quantifying
thousands of proteins in even very short gradients. We are pleased that
within our close collaboration with Bruker, the vendor-integrated version
of DIA-NN called TIMS DIA-NN now becomes part of the PaSER bioinformatics
Blazing search speed with uncompromised search results.
A) Protein ID from replicate DDA runs of a human cell lysate using PaSER for real-time results and benchmarked against MaxQuant
B) Peptide ID from replicate DDA runs of a human cell lysate using PaSER for real-time results and benchmarked against MaxQuant
C) PaSER and MaxQuant identify 97% of the same proteins and nearly 90% of the same peptides.
PaSER provides consistent results obtained from the real-time search, identical to offline search of the same data. Since the database search algorithm is run on the GPU, the search times are negligible compared to CPU based searches.
In addition to real-time search the opportunity to search a larger space for specific applications, including including peptidome studies, becomes reality. PaSER benchmarked against the gold standard in DDA searches performs as good or better.
Take advantage of the real-time identification to drastically reduce the required time for dda-PASEF label-free quantitation (LFQ) with match-between-runs (MBR).
Easily identify poor samples and re-acquire them with Run & Done on PaSER before starting the quantitative portion of your workflow. Our IM filtered XIC based quantitative workflow is powered by the Census algorithm (https://doi.org/10.1002/0471250953.bi1312s29).
|Project||# of Samples||MS acquisition Length (min)||Total Acquisition Time (min)||LFQ Processing Time (min)||Ratio vs Acquisition|
|Dense HE Data||6||40||240||26.2||1/9|
|Tenzer style HYE124||6||120||720||24.5||1/30|
|Human Ecoli spike||18||40||720||116.8||1/6|
|Plasma App Note||212||14.4||3052.8||59.6||1/50|
TIMS Viz provides the powerful ability to view 4-D proteomics data like never seen before. The mobility separation and focusing upstream from the quadrapole and TOF result in the separation of isobaric and near isobaric species. Using other technologies and in the case of PTM enrichments and the analysis of complex samples it is common that co-eluting species that are isobaric or near isobaric in mass co-fragment. The timsTOF Pro/flex obviates this by first separating ions by their collisional cross section. The ability to visualize, search and sort on such events has never been available before. We term isobaric masses with different mobility values mobility offset mass aligned (MOMA), and TIMS Viz can tease such events out within complex runs. TIMS Viz allows you to set tolerances to your discretion while simultaneously overlaying both features and PSMs that were observed post database search. TIMS Viz enables you to view what you never knew was there before.
“Real-time results delivered by the PaSER box on our timsTOF Pro has been a huge time saver. This allows us to develop methods in real-time, inform on LC and instrument performance and overall a major gain in efficiency.“
“Innovative software tools are a necessity to address unanswered biological questions with mass spectrometry. The trapped ion mobility functionality and the robustness of the timsTOF Pro offer unique bottom-up proteomics capabilities that can be effectively used to study many diseases.”
For Research Use Only. Not for use in clinical diagnostic procedures.