Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Ion Mobility-Assisted Free Radical Initiated Peptide Sequencing

ABSTRACT: Free radical-initiated peptide sequencing (FRIPS), a TEMPO-assisted mass spectrometry, has shown potential as a peptide tandem mass spectrometry (MS/MS) tool that can be an alternative to electron-based mass spectrometry and the traditional collision-based methods. However, a couple of instrumentation barriers need to be overcome before this approach can be a practical tool. One of the barriers is the limitation of the FRIPS-based technique to ion trap mass spectrometry or an orbitrap due to the MS3 workflow. To overcome this challenge, the work introduces a recently developed activation within the trapped ion mobility mass spectrometry (TIMS) device that was found to initiate the radical species. More importantly, the generated product ion was mobility separated, which increased the sequence coverage. Overall, coupling the activation within the TIMS cell with the para-TEMPO-Bz-based FRIPS mass spectrometry offers another practical approach that can be utilized for peptide MS/MS analysis

INSTRUMENT(S): timsTOF

ORGANISM(S): Homo Sapiens (ncbitaxon:9606)

SUBMITTER: Nicholas Borotto

PROVIDER: MSV000095935 | MassIVE |

REPOSITORIES: MassIVE

ACCESS DATA

Json Xml

Dataset's files

Source:

			Action	DRS
		Other

Items per page:

1 - 1 of 1

Similar Datasets

Ion Mobility-Assisted Free Radical Initiated Peptide Sequencing v2

Project description:Free radical-initiated peptide sequencing (FRIPS) is a tandem mass spectrometry technique (MS/MS) that enables radical-based dissociation on instruments only capable of collisional activation. In FRIPS, peptides are chemically-derivatized with a compound that undergoes homolytic cleavage and generates radicals upon collisional activation. These radicals then propagate through the peptide backbone enabling the sequencing of peptide ions. This MS/MS technique has shown promise in sequencing post-translationally modified peptides, but it is typically performed in an MS3 workflow and single-step MS/MS approaches result in the generation of both collisional- and radical-driven dissociation products and highly complex spectra. Recently, our group developed a method to dissociate peptide ions prior to ion mobility analysis within a trapped-ion mobility spectrometry (TIMS) device. In this work, we examine if this CIDtims technique is capable of initiating the homolytic cleavage of the FRIPS precursor. We then examine if the resultant ion mobility separation results in additional assignments of product ions and improved sequence coverage. We demonstrate that activation within the TIMS device does indeed promote robust radical initiation and fragmentation and that the generated product ions are mobility separated enabling facile assignment and increased sequence coverage.

2024-10-15 | MSV000096102 | MassIVE

Ion mobility fractionation coupled to PASEF for enhanced proteome coverage

Project description:Data provided report the use of trapped ion mobility spectrometry (TIMS) to fractionate ions in the gas phase based on their ion mobility (V⋅s/cm2) followed by parallel accumulation serial fragmentation (PASEF) in a quadrupole time-of-flight instrument. TIMS fractionation coupled to DDA-PASEF allowed for the detection of approximately 7,000 proteins and over 70,000 peptides per overall run from 200ng of human (HeLa) cell lysate per injection using a commercial UPLC column with a 90-minute gradient. This project also explored the utility of TIMS fractionation to generate a DDA library for downstream DIA analysis using shorter LC gradients (20 minutes) as well as lower sample input. Using a 20min gradient, we identified 4,092 and 6,654 proteins on average per run, respectively, from 10ng and 200ng of human (HeLa) cell lysate input based on a TIMS-fractionated library consisting of 82,214 peptides derived from 7,615 proteins.

2022-08-12 | PXD033129 | Pride

Extending the Separation Space with Trapped Ion Mobility Spectrometry Improves the Accuracy of Isobaric Tag-based Quantitation in Proteomic LC/MS/MS

Project description:Two-dimensional separation by nano-LC and trapped ion mobility spectrometry (TIMS) prior to Q-TOF tandem mass spectrometry significantly improves the accuracy of isobaric tag-based quantitation in proteome analysis without the need for additional measurement time for TIMS insertion. The obtained peak capacity of up to 3,300 per hour in LC/TIMS reduced the co-isolation of precursor ions at the quadrupole analyzer, resulting in more accurate ratios of reporter ions derived from isobaric tags in product ion spectra obtained at the TOF analyzer. We also found that TIMS with a narrow quadrupole isolation window could reduce the ratio compression effect at least as effectively as the synchronous precursor selection method using MS3 scans without compromising sensitivity or coverage. Our results suggest that the short-gradient LC/TIMS/Q/TOF system is an excellent platform for high-throughput proteomics studies.

2020-05-22 | PXD018650 | JPOST Repository

Machine learning-based noninvasive assays explore the use of volatile organic compounds for clinical diagnosis and evaluation of gastrointestinal tumors

Project description:Interventions: Gold Standard:pathological diagnosis ;Index test:Volatile organic compounds detected by gas chromatography-mass spectrometer and gas chromatography-ion migration spectrometry Primary outcome(s): Volatile organic compounds;sensitivity;specificity Study Design: Diagnostic test for accuracy

| 89221 | ecrin-mdr-crc

Optimizing single cell proteomics on a trapped ion mobility spectrometry for label-free experiments

Project description:Although single cell RNA-seq has had a tremendous impact on biological research, a corresponding technology for unbiased mass spectrometric analysis of single cells has only recently become available. Significant technological breakthroughs including miniaturized sample handling have enabled proteome profiling of single cells. Further, a trapped ion mobility spectrometry (TIMS) in combination with parallel accumulation-serial fragmentation operated with data-dependant acquisition mode has allowed improved proteome coverage from low-input samples. It has been demonstrated that modulating ion flux in TIMS affects overall performance of proteome profiling. However, the effect of TIMS settings for analyzing low-input samples has been less investigated. Thus, we sought to optimize conditions of TIMS in regards of ion accumulation/ramp times and ion mobility range for low-input samples. We observed that ion accumulation times of 180 ms and monitoring a narrower ion mobility range from 0.7 to 1.3 Vs cm-2 resulted in a substantial gain in the depth of proteome coverage and in detecting proteins with low abundance. We applied these optimized conditions for proteome profiling of sorted human primary T cells, which yielded an average of 365, 804, 1,116, and 1,651 proteins from single, five, ten, and forty T cells, respectively. Notably, we demonstrated that the depth of proteome coverage from low number of cells was sufficient to delineate several essential metabolic pathways and T cell receptor signaling pathway. Finally, we showed the feasibility of detecting post-translational modifications including phosphorylation and acetylation from single cells. We believe that these parameters could be applied to label-free analysis of single cells obtained from clinically relevant samples.

2024-08-02 | PXD039066 | Pride

Synchro-PASEF slices peptide precursors enabling specific fragment extraction and interference removal in data independent acquisition

Project description:Data independent acquisition methods have become increasingly popular in mass spectrometry (MS)-based proteomics because they enable parallel and continuous acquisition of fragment spectra. However, these advantages come with the challenge of correctly reconstructing the precursor-fragment relationship in these highly multiplexed spectra for reliable identification and quantification. Here we introduce a scan mode for the combination of trapped ion mobility spectrometry (TIMS) with parallel accumulation – serial fragmentation (PASEF) that naturally and continuously follows the ion cloud in ion mobility and peptide mass dimensions as opposed to the step functions we employed before. Termed synchro-PASEF, it increases fragment yield several fold at sub-second cycle times. The quadrupole selection window moves synchronously through the mass and ion mobility range, deconvoluting the DIA spectra and defining precursor-quadrupole relationships. In this process, the quadrupole slices through the peptide precursors, which separates fragment ion signals of each precursor into adjacent synchro-PASEF scans. This precisely defines precursor – fragment relationship in ion mobility and mass dimensions. Importantly, the two parts of the fragment ion transitions in synchro-PASEF provide a further dimension of specificity via a lock and key mechanism. This is especially advantageous for quantification, where signals from interfering precursors in the dia selection window only affect the top or bottom part of the fragment ion, allowing them to be filtered out. In this paper, we establish the defining features of synchro-PASEF and explore its potential for proteomics analyses.

2023-03-10 | PXD037766 | Pride

AlphaTims: Indexing unprocessed trapped ion mobility spectrometry - time of flight data for fast and easy accession and visualization

Project description:High resolution mass spectrometry-based proteomics generates large amounts of data, even in the standard liquid chromatography (LC) Ã¢â‚¬â€œ tandem mass spectrometry configuration. Adding an ion mobility dimension vastly increases the acquired data volume, challenging both analytical processing pipelines and especially data exploration by scientists. This has necessitated data aggregation, effectively discarding much of the information present in these rich data sets. Taking trapped ion mobility spectrometry (TIMS) on the quadrupole time of flight platform (Q-TOF) as an example, we developed an efficient indexing scheme that represents all data points as detector arrival times on scales of seconds (LC), milliseconds (TIMS) and microseconds (TOF). In our open source AlphaTims package, data are indexed, accessed and visualized by a combination of tools of the scientific Python ecosystem. We interpret unprocessed data as a sparse 4D matrix and use just in time compilation to machine code with Numba, accelerating our computational procedures by several orders of magnitude while keeping to familiar indexing and slicing notations. For samples with more than six billion detector events a modern laptop can load and index raw data in about a minute. Loading is even faster when AlphaTims has already saved indexed data in a HDF5 file, a portable scientific standard used in extremely large-scale data acquisition. Subsequently, data accession along any dimension and interactive visualization happen in milliseconds. We have found AlphaTims to be a key enabling tool to explore high dimensional LC-TIMS-Q-TOF data and have made it freely available as an open-source Python package with a stand-alone graphical user interface at https://github.com/MannLabs/alphatims or as part of the AlphaPept framework.

2021-07-30 | PXD027359 | Pride

MaxQuant software for ion mobility enhanced shotgun proteomics

Project description:Ion mobility can add a dimension to LC-MS based shotgun proteomics which has the potential to boost proteome coverage, quantification accuracy and dynamic range. Required for this is suitable software that extracts the information contained in the four-dimensional (4D) data space spanned by m/z, retention time, ion mobility and signal intensity. Here we describe the ion mobility enhanced MaxQuant software, which utilizes the added data dimension. It offers an end to end computational workflow for the identification and quantification of peptides, proteins and posttranslational modification sites in LC-IMS-MS/MS shotgun proteomics data. We apply it to trapped ion mobility spectrometry (TIMS) coupled to a quadrupole time-of-flight (QTOF) analyzer. A highly parallelizable 4D feature detection algorithm extracts peaks which are assembled to isotope patterns. Masses are recalibrated with a non-linear m/z, retention time, ion mobility and signal intensity dependent model, based on peptides from the sample. A new matching between runs (MBR) algorithm that utilizes collisional cross section (CCS) values of MS1 features in the matching process significantly gains specificity from the extra dimension. Prerequisite for using CCS values in MBR is a relative alignment of the ion mobility values between the runs. The missing value problem in protein quantification over many samples is greatly reduced by CCS aware MBR.MS1 level label-free quantification is also implemented which proves to be highly precise and accurate on a benchmark dataset with known ground truth. MaxQuant for LC-IMS-MS/MS is part of the basic MaxQuant release and can be downloaded from http://maxquant.org.

2020-04-23 | PXD014777 | Pride

Spatial Proteomics by Trapped Ion Mobility supported MALDI MS/MS Imaging: A First Glance into Multiplexed and Spatial Peptide Identification

Project description:RATIONALE In spatial proteomics, matrix-assisted laser desorption/ionization (MALDI) imaging enables rapid and cost-effective peptide measurement. Yet, in situ peptide identification remains challenging. Therefore, this study aims to integrate the trapped ion mobility spectrometry (TIMS)-based parallel accumulation-serial fragmentation (PASEF) into MALDI imaging of peptides to enable multiplexed MS/MS imaging. METHODS An initial MALDI TIMS MS1 survey measurement is performed, followed by a manual generation of a precursor list containing mass over charge values and ion mobility windows. Inside the dual TIMS system, submitted precursors are trapped, separately eluted by their ion mobility and analyzed in a quadrupole time-of-flight device, thereby enabling multiplexed MALDI MS/MS imaging. Finally, precursors are identified by peptide to spectrum matching. RESULTS This study presents the first multiplexed MALDI TIMS MS/MS imaging (iprm-PASEF) of tryptic peptides. Its applicability is showcased on two histomorphologically distinct tissue specimens in a 4-plex and 5-plex setup. Precursors were successfully identified by the search engine MASCOT in one single MALDI imaging experiment for each respective tissue. Peptide identifications were corroborated by liquid-chromatography tandem mass spectrometry experiments and fragment colocalization analyses. CONCLUSIONS In the present study, we demonstrate the feasibility of TIMS-based MALDI MS/MS imaging for the multiplexed and rapid identification of tryptic peptides in a spatial manner. Hence, it represents a first step towards the integration of MALDI imaging into the emerging field of spatial proteomics. The datasets will be uploaded separately. This data set contains the PDX MS1 data. There are two tissues vissible on the.tif image. The relevant one is on the left side of the slide (away from the barcode).

2024-11-11 | MSV000096374 | MassIVE

Spatial Proteomics by Trapped Ion Mobility supported MALDI MS/MS Imaging: A First Glance into Multiplexed and Spatial Peptide Identification

Project description:RATIONALE In spatial proteomics, matrix-assisted laser desorption/ionization (MALDI) imaging enables rapid and cost-effective peptide measurement. Yet, in situ peptide identification remains challenging. Therefore, this study aims to integrate the trapped ion mobility spectrometry (TIMS)-based parallel accumulation-serial fragmentation (PASEF) into MALDI imaging of peptides to enable multiplexed MS/MS imaging. METHODS An initial MALDI TIMS MS1 survey measurement is performed, followed by a manual generation of a precursor list containing mass over charge values and ion mobility windows. Inside the dual TIMS system, submitted precursors are trapped, separately eluted by their ion mobility and analyzed in a quadrupole time-of-flight device, thereby enabling multiplexed MALDI MS/MS imaging. Finally, precursors are identified by peptide to spectrum matching. RESULTS This study presents the first multiplexed MALDI TIMS MS/MS imaging (iprm-PASEF) of tryptic peptides. Its applicability is showcased on two histomorphologically distinct tissue specimens in a 4-plex and 5-plex setup. Precursors were successfully identified by the search engine MASCOT in one single MALDI imaging experiment for each respective tissue. Peptide identifications were corroborated by liquid-chromatography tandem mass spectrometry experiments and fragment colocalization analyses. CONCLUSIONS In the present study, we demonstrate the feasibility of TIMS-based MALDI MS/MS imaging for the multiplexed and rapid identification of tryptic peptides in a spatial manner. Hence, it represents a first step towards the integration of MALDI imaging into the emerging field of spatial proteomics. All Datasets are uploaded separateley. This file contains the dataset for mouse kidney MS1 data. On the same slice a mouse liver was measured for a different experiment.

2024-11-11 | MSV000096373 | MassIVE

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data