Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Human HeLa cells LC MS/MSLC-MSMS

ABSTRACT: We developed a reagent of 10-plex isobaric tags (IBT) with high stability and low cost. The labeled peptides were sensitively detected on Orbitrap Q Exactive MS with MS2 resolution of 35000 at 30% NCE, while the peptides were efficiently labelled over 97% by IBT at ratio of 20:1 of reagent/peptide (w/w) in 200mM TEAB buffer within 2h. The IBT labeling was demonstrated in a wide dynamic range of 50 folds but without obvious matrix effect on quantification. Importantly, there was little quantification bias found among the individual IBT tags, indicating that the peptides labeled by different tags were quantitatively comparable. The IBT 10-plex reagents were applied for dynamically monitoring the quantitative responses of phosphoproteome ignited by EGF treatment in Hela cells. Total 5361 unique phosphopeptides were identified, which reached a similar conclusion as other reported. The IBT reagents were therefore experimentally proven as a new type of isobaric labeling peptides and useful for a large quantity peptides of quantitative proteomics.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Hela Cell

SUBMITTER: He Yanbin

LAB HEAD: Siqi Liu

PROVIDER: PXD008681 | Pride | 2022-03-04

REPOSITORIES: Pride

ACCESS DATA

Json Xml

Dataset's files

Source:

			Action	DRS
	Fraction_1.mgf	Mgf
	Fraction_1.raw	Raw
	Fraction_2.mgf	Mgf
	Fraction_2.raw	Raw
	Fraction_3.mgf	Mgf

Items per page:

1 - 5 of 12

Similar Datasets

A Tribrid Hyperplexing approach for Large-scale Quantitative Proteomic Analysis

Project description:Isobaric labeling has emerged as an indispensable quantitative proteomic approach for its unprecedented multiplexing capacity in a single analysis. Currently, different hyperplexing approaches have been developed to meet the need for increasing sample numbers in large-scale cohort analysis. In this report, we present a tribrid hyperplexing approach by the combinational use of three types of isobaric reagents, a novel isobaric tag 16-plex (IBT16) reagent, and the widely used TMT (TMT11) and TMTpro (TMT18) reagents. After the determination of the labeling efficiency and the optimization testing conditions, we systematically evaluated the identification and quantification performance of the three labeling methods in both independent and combinatorial means using the mixtures of E. coli and HeLa peptides with different ratios. Our results reveal that the three reagents are quite similar in all testing aspects although with some differences, and the combination use of three reagents to expand the multiplexing capacity is feasible. Furthermore, we provide practical recommendations for the choice of isobaric reagent combinations according to different plexes.

2023-03-17 | PXD037498 | Pride

27-plex Tandem Mass Tag Mass Spectrometry for Profiling Brain Proteome in Alzheimer's Disease

Project description:Systematically optimized the protocol for the newly developed 16-plex TMT Reported a novel 27-plex method to enhance the multiplexity for large-scale proteomic profiling, by combining two sets of distinct isobaric tags (11-plex and 16-plex TMT) based on their disparate mass shifts on modified peptides.

2021-04-09 | PXD025261 | Pride

NHS-Ester Tandem Labeling in One Pot for 48-Plex Proteomics

Project description:Stable-isotope labeling strategies are extensively used for multiplex quantitative proteomics. Hybrid isotope labeling strate-gies that combine the use of isotopic mass difference labeling and isobaric tags can greatly increase sample multiplexity. In this work, we present a novel hybrid isotope labeling approach that we termed NHS-ester tandem labeling in one pot (NETLOP). We first optimized 16-plex isobaric TMTpro labeling of lysine residues followed by 2-plex or 3-plex isotopic mTRAQ labeling of peptide N-termini, both of which with commercially available NHS-ester reactive reagents. We then demonstrated the utility of the NETLOP approach by labeling HeLa cell samples and performing proof-of-principle quanti-tative 32-plex and 48-plex proteomic analyses, each in a single LC-MS/MS experiment. Compared to current hybrid isotope labeling methods, our NETLOP approach requires no sample cleanup between different labeling steps to minimize sample losses, induces no retention time shifts that compromise quantification accuracy, can be adapted to other NHS-ester isotop-ic labeling reagents to further increase multiplexity, and is compatible with samples from any origin in a wide array of bio-logical and clinical proteomics applications.

2022-02-17 | PXD024780 | Pride

Mouse,GC-1,TMT（Tandem Mass Tag）tag quantification

Project description:Tandem Mass Tag (TMT) technology is an in vitro peptide labeling technique developed by Thermo Fisher Scientific, a peptide labeling technology developed in vitro. The technology uses six (TMTsixplex™ Isobaric Label Reagent Set) or 10 (TMT10plex™ Isobaric Label Reagent Set) isotope labels by labeling peptide specific amino acid sites. In TMT experiments, the relative and absolute amounts of proteins in up to 10 different samples can be compared flexibly by tandem mass spectrometry analysis. In TMT experiments, whole proteins from tissue cells are obtained by protein extraction, and the whole proteins are enzymatically cleaved into peptides by trypsin, and the TMT reagents are used to label all the obtained peptides. The TMT reagent consists of three main components: a reporter group, an equilibrium group and a peptide reaction group.

2022-01-19 | PXD030967 | Pride

Identification of targets of the protein disulfide reductase thioredoxin

Project description:Identification of targets of the protein disulfide reductase thioredoxin using liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) and thiol specific differential labeling with isotope-coded affinity tags (ICAT). Reduction of specific target disulfides is quantified by measuring ratios of cysteine residues labeled with the heavy (13C) and light (12C) ICAT reagents in peptides derived from tryptic digests of Trx-treated and non-treated samples. Keywords: protein, LC-MS/MS, ICAT

2008-11-20 | GSE11650 | GEO

102-Plex Approach for Accurate and Multiplexed Proteome Quantification

Project description:Numerous hyperplexing approaches have been developed to meet the demand for large-scale basic and clinical analysis. Currently, the analysis capacity has expanded to up to 54 samples within a single experiment by utilizing different isotopic and isobaric reagent combinations. In this report, we propose a super multiplexed approach to enable the analysis of up-to 102-plex samples within a single experiment. by the combination of our recently developed TAG-TMTpro and TAG-IBT16 labeling. We systematically investigated the identification and quantification performance of the 102-plex approach using the mixtures of E. coli and HeLa peptides. Our results revealed that all analyses demonstrated accurate and reliable quanti-fication performance. The combination of TAG-TMTpro and TAG-IBT16 approaches expands the multiplexing capacity to 102-plex, providing a more multiplexed quantification method for even larger-scale proteomic analysis.

2024-01-15 | PXD042398 | Pride

iTRAQ-based phospho-proteomics identifies a critical role of ATF2 in pseudorabies virus replication

Project description:Pseudorabies virus (PRV), the etiological agent of Aujeszky’s disease, is a swine pathogen of the alphaherpesvirinae subfamily resulting in devastating neurological and respiratory disorders in piglets, or abortion in pregnant sows.Here, we performed isobaric tags for relative and absolute quantitation (iTRAQ) quantitative phosphoproteomics on PRV-infected PK-15 cells, which identified 5723 phospho-peptides, corresponding to 2180 proteins, including 810 proteins showing significant changes in phosphorylation level during early PRV infection.

2022-10-13 | PXD014685 | Pride

TMTpro-18plex: The Expanded and Complete Set of TMTpro Reagents for Sample Multiplexing

Project description:The development of the TMTpro-16plex series expanded the breadth of commercial isobaric tagging reagents by nearly 50% over classic TMT-11plex. In addition to the described 16plex reagents, the proline-based TMTpro molecule can accommodate two additional combinations of heavy carbon and nitrogen isotopes. Here, we introduce the final two labeling reagents, TMTpro-134C and TMTpro-135N, which permit the simultaneous global protein profiling of 18 samples with no missing values. For example, six conditions with three biological replicates can now be perfectly accommodated. We showcase the 18plex reagent set by profiling the proteome and phosphoproteome of a pair of isogenic breast cancer cell lines under three conditions in triplicate. We compare the depth and quantitative performance of this dataset with a TMTpro-16plex experiment in which two samples were omitted. Our analysis revealed similar numbers of quantified peptides and proteins, with high quantitative correlation. We interrogated further the TMTpro-18plex dataset by highlighting changes in protein abundance profiles under different conditions in the isogenic cell lines. We conclude that TMTpro-18plex further expands the sample multiplexing landscape, allowing for complex and innovative experimental designs.

2021-04-28 | PXD024275 | Pride

TMTpro16plex Isobaric Labeling Reagents: Attaining New Sample Multiplexing Heights and New Cellular Proteome Depths in Quantitative Proteomics

Project description:Isobaric labeling empowers proteome-wide expression measurements simultaneously across multiple samples. Here, an expanded set of 16 isobaric reagents based on a proline backbone (TMTpro) is presented. These reagents have similar characteristics to existing TMT reagents but with increased fragmentation efficiency and signal. In a proteome-scale example dataset, we compared eight common cell lines with and without Torin1 treatment with three replicates, quantifying more than 8,800 proteins (mean of 7.5 peptides/protein) per replicate with an analysis time of only 67 min per proteome examined. Finally, we modified the powerful thermal stability assay to examine proteome-wide melting shifts after treatment with DMSO, 1 or 20 ��M staurosporine with five replicates. This new assay identified and dose-stratified the staurosporine binding to 228 cellular kinases in just one, 18-hr experiment. TMTpro reagents allow unprecedented complexity in experimental designs��all with basically no missing values across the 16 samples and no loss in quantitative integrity.

2020-03-19 | PXD016940 | Pride

TMTpro16plex Isobaric Labeling Reagents: Attaining New Sample Multiplexing Heights and New Cellular Proteome Depths in Quantitative Proteomics

Project description:Isobaric labeling empowers proteome-wide expression measurements simultaneously across multiple samples. Here, an expanded set of 16 isobaric reagents based on a proline backbone (TMTpro) is presented. These reagents have similar characteristics to existing TMT reagents but with increased fragmentation efficiency and signal. In a proteome-scale example dataset, we compared eight common cell lines with and without Torin1 treatment with three replicates, quantifying more than 8,800 proteins (mean of 7.5 peptides/protein) per replicate with an analysis time of only 67 min per proteome examined. Finally, we modified the powerful thermal stability assay to examine proteome-wide melting shifts after treatment with DMSO, 1 or 20 µM staurosporine with five replicates. This new assay identified and dose-stratified the staurosporine binding to 228 cellular kinases in just one, 18-hr experiment. TMTpro reagents allow unprecedented complexity in experimental designs—all with basically no missing values across the 16 samples and no loss in quantitative integrity.

2020-03-19 | PXD016491 | Pride

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