Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

A quick guideline for PRM method development applied to target proteomics in biological fluids

ABSTRACT: Combining in-silico analysis, serum high abundant protein depletion, and DDA and PRM acquisitions, we established a simple workflow to select PRM high response peptides for protein quantification in serum. The workflow enables evaluating peptide intensity, MS2 match with the spectral library, peptide stability, and linearity of response to different injected masses. This workflow was successfully applied to twelve human complement system proteins. Besides, we employed synthetic labelled peptides to compensate inter LC-MS run variability, improving the quantification. We could demonstrate this method is useful and applicable for quantification of diverse targets with time optimization and accuracy insurance by quantitative proteomics technology.

INSTRUMENT(S): Orbitrap Fusion Lumos, Orbitrap Exploris 120

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Blood Serum

SUBMITTER: Michel Batista

LAB HEAD: Fabiano Borges Figueiredo

PROVIDER: PXD045089 | Pride | 2024-02-05

REPOSITORIES: Pride

ACCESS DATA

Json Xml

Dataset's files

Source:

			Action	DRS
	20191205_52819_Fabiano_SoroDepletadoHumano.raw	Raw
	20191205_52919_Fabiano_SoroTotalHumano.raw	Raw
	20210526_24221_Fabiano_soroDPTD_700ng_T311.raw	Raw
	20210706_24221_Fabiano_FCN_C1qA_MBL_soroDPTD.raw	Raw
	20210708_27321_Fabiano_soroTT_T150.raw	Raw

Items per page:

1 - 5 of 53

Similar Datasets

Haptenation of Macrophage Migration Inhibitory Factor

Project description:Firstly, the adductome of the two major mouse blood proteins, as well as of the protein macrophage migration inhibitory factor (MIF) with the model hapten tetramethylrhodamine isothiocyanate (TRITC)was invastigated. After incubation at different molar ratios of TRITC, an untargeted Full MS/dd-MS2 experimental approach was used with a QExactive Quadrupole-Orbitrap mass spectrometer. The data were processed with Proteome Discoverer and the suggested sites of adductions were confirmed in parallel reaction monitoring mode (PRM) on the same instruments. Blood and lymph node samples of mise treated topically with the same hapten were screened for the peptides containing the most reactive sites of the aformentioned proteins. Only the adducted N-terminal peptide of the protein MIF could be detected. Thereafter, a quantification approach in PRM mode using standards was implemented to quantify this peptide, both modified and unmodified. Quantification was based in characteristic b and y ions.

2022-04-08 | PXD031143 | Pride

TARGETED PROTEOMICS FOR THE DETECTION OF SARS-COV-2 PROTEINS

Project description:TARGETED PROTEOMICS FOR THE DETECTION OF SARS-COV-2 PROTEINS: SARS-CoV-2, COVID-19, proteomics, targeted mass spectrometry, LC-MS, parallel reaction monitoring (PRM), limit of detection

2020-04-24 | PXD018760 | Pride

Picky - a simple online method designer for targeted proteomics

Project description:Picky is an online method designer for targeted proteomics. To assess the performance of PRM methods designed by Picky we carried out a benchmark experiment. As reference samples we used different amounts of human proteins spiked into 1.4 µg yeast lysate. These sample where analyzed in PRM and DDA mode. All methods inlcuded a shotgun method (top10 for DDA and top3 for PRM) and can therefore all be analyzed by MaxQuant.

2017-08-04 | PXD007039 | Pride

Parallel Reaction Monitoring of 42 peptide pairs with 15N metabolic labeling

Project description:To examine the properties of 14N and 15N labeled peptides from 15 metabolic labeling samples, we set up targeted quantification using Parallel Reaction Monitoring (PRM) on a high resolution, high accuracy mass spectrometer. We first examined the co-elution of light and heavy peptide forms. Furthermore, we examined the fragment spectra of the light and heavy peptides and compared the pattern of the fragments.

2022-06-09 | PXD030112 | Pride

An Integrated Quantitative Proteomics Workflow for Biomarker Discovery and Validation in Plasma

Project description:Blood plasma is one of the most widely used samples for biomarker discovery research as well as clinical investigations for diagnostic and therapeutic purposes. However, the plasma proteome is extremely complex due to its wide dynamic range of protein concentrations and the presence of high-abundance proteins. Here we are describing an optimized integrated quantitative proteomics pipeline combining the label-free and multiplexed-labeling-based (Isobaric tags for relative and absolute quantitation) proteome profiling methods for biomarker discovery, followed by the targeted approaches for validation of the identified potential marker proteins. In this workflow, the targeted quantitation of proteins is carried out by multiple-reaction monitoring (MRM) and parallel-reaction monitoring (PRM) mass spectrometry. Thus, our approach enables both unbiased screenings of biomarkers and their subsequent selective validation in human plasma. The overall procedure takes only 1–2 days to complete including the time for data acquisition (excluding database searching). This protocol is quick, flexible and eliminates the need for a separate immunoassay-based validation workflow in blood biomarker investigations.

2020-09-07 | PXD017834 | Pride

An integrated multi-omic workflow reveals markers of host-microbe dynamics in clinical cystic fibrosis samples

Project description:Airway microbiota composition correlates with cystic fibrosis (CF) progression, however, microbial drivers of disease remain unclear. MS-based metaproteomics of bronchoalveolar lavage fluid (BALF) offers insights into both host and microbe dynamics and potential interactions. However, detection of microbial proteins, and analysis of their interaction with host proteins is analytically challenging. As a solution, we have developed a novel, integrated workflow coupling deep MS-based BALF analysis, with customized bioinformatic processing of both host and microbial proteins, generating a panel of verified host and microbe peptide candidates suitable for targeted analysis within individual patient samples. We have utilized this workflow in our ongoing work to identify a promising host and microbe peptide panel for application to CF disease progression studies.

2024-06-25 | PXD041457 | Pride

Triggered offset fragmentation allows massively multiplexed target profiling on quadrupole-orbitrap mass spectrometers

Project description:Parallel reaction monitoring (PRM) on quadrupole-orbitrap mass spectrometers has a limited multiplexing capacity which depends heavily on the reproducibility of peptide retention times. To overcome these limitations, we aimed to establish a data acquisition mode that allows retention-time-independent massive multiplexing on quadrupole-orbitrap mass spectrometers. The presented method is based on data-dependent acquisition and is called pseudo-PRM. In principle, high-intensity stable isotope-labeled peptides are used to trigger the repeated fragmentation of the corresponding light peptides. In this way, pseudo-PRM data can be analyzed like normal PRM data with Skyline. We tested pseudo-PRM for the target detection from yeast, human cells, and serum, and assessed the quantification accuracy/precision and sensitivity. Moreover, we analyzed multiplexing of more than 1,000 targets in a single pseudo-PRM run. Finally, we applied pseudo-PRM to quantify vaccinia virus proteins during infection, verifying that pseudo-PRM presents an alternative method for multiplexed target profiling on quadrupole-orbitrap mass spectrometers.

2020-06-19 | PXD017778 | Pride

p300 acetylates Ub and linear Ub dimers

Project description:In this project we present the identification of in vitro acetylation of Ub by the acetyltransferase p300. Free monomeric Ub or M1-linked linear Ub dimers (diUb) were incubated in vitro with p300 in presence or absence of acetyl-Coenzyme A (AcCoA). For identification of the acetylation status of the respective Ub variants parallel reaction monitoring (PRM), a targeted-MS approach, was used.

2022-10-14 | PXD028813 | Pride

Haptenation of Macrophage Migration Inhibitory Factor-Dataset 2

Project description:Firstly, the adductome of he protein macrophage migration inhibitory factor (MIF) with different haptens encompassing different reaction mechanisms and potencies was investigated. After incubation of MIF with the different haptens at 5 times molar excess of hapten, an untargeted Full MS/dd-MS2 experimental approach was used with a QExactive Quadrupole-Orbitrap mass spectrometer. The data were processed with Proteome Discoverer and the suggested sites of adductions were confirmed in parallel reaction monitoring mode (PRM) on the same instruments.

2022-03-18 | PXD031793 | Pride

Study on prognostic biomarkers of HCC based on mass spectrometry

Project description:using DIA, and PRM MS-based workflow to research the prognostic biomarkers of HCC

2022-02-17 | PXD027817 | Pride