Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

A Comprehensive Comparison of Sample Preparation and Normalization Methods for Bottom-Up Proteomics of Extracellular Vesicles

ABSTRACT: In this scientific study, we evaluated sample preparation methods for exosome proteomics along with data normalization techniques. We commenced by extracting exosomal proteins using three different lysis buffers: 8M urea, 1% sodium deoxycholate (SDC), and 0.1% sodium dodecyl sulfate (SDS). Through comparative analysis, we determined that 1% SDC was the most efficient lysis buffer, yielding the highest quality and quantity of protein extracts. Subsequently, we examined various data normalization methods to ensure reliable and reproducible results in proteomic profiling. Our findings highlight the critical role of optimal sample preparation and data normalization in exosome proteomics, which are essential for accurate biomarker discovery and understanding exosomal functions in biological processes. This research provides a foundation for standardized protocols in exosome proteomics and contributes to the enhancement of data quality in this burgeoning field.

ORGANISM(S): Homo Sapiens

SUBMITTER: Haojie Lu

PROVIDER: PXD057883 | iProX | Thu Nov 14 00:00:00 GMT 2024

REPOSITORIES: iProX

ACCESS DATA

Json Xml

Similar Datasets

Navigating Illumina Methylation Data: Biology vs. Technical Artefacts

Project description:Illumina-based BeadChip arrays have revolutionized genome-wide DNA methylation profiling, pushing it into diagnostics. However, comprehensive quality assessment remains a challenge within a wide range of available tissue materials and sample preparation methods. This study tackles two critical issues: differentiating between biological effects and technical artefacts in suboptimal quality samples and the impact of the first sample on the Illumina-like normalization algorithm. We introduce three quality control scores based on global DNA methylation distribution (DB-Score), bin distance from copy number variation analysis (BIN-Score), and consistently methylated CpGs (CM-Score), that rely on biological features rather than internal array controls. These scores were explored and benchmarked across independent study cohorts. Additionally, we reveal deviations in beta values caused by different sample rankings with the Illumina-like normalization algorithm, verified these with whole-genome methylation sequencing data and showed effects on differential DNA methylation analysis. Our findings underscore the necessity of consistently utilizing a pre-defined normalization sample within the ranking process to boost the reproducibility of the Illumina-like normalization algorithm. In conclusion, our study delivers valuable insights, practical recommendations, and R functions designed to enhance the reproducibility and quality assurance of DNA methylation analysis, particularly for challenging sample types.

2024-12-19 | GSE269421 | GEO

dia-PASEF HeLa dilution series 0.2ng - 200ng on Bruker timsTOF Pro 2

Project description:Different sample preparation methods were tested for HeLa proteome analysis. A sample obtained using sodium deoxycholate-based lysis allowed identification of the highest number of proteins. For this sample, a dilution series was acquired in triplicates ranging from 0.2ng to 200ng. All measurements were performed on Bruker timsTOF Pro 2 operated in dia-PASEF mode and analysed library-free using DIA-NN 1.8.

2022-06-20 | PXD029836 | Pride

RevGel-seq: instrument-free single-cell RNA sequencing using a reversible hydrogel for cell-specific barcoding [scRNA-seq_stopping_Scipio]

Project description:Progress in sample preparation for scRNA-seq is reported based on RevGel-seq, a reversible-hydrogel technology. Barcode bead-cell tandems stabilized by a chemical linker are dispersed in the hydrogel in the liquid state. Upon gelation the tandems are immobilized, cell lysis is triggered by detergent diffusion, and RNA molecules are captured on the adjacent barcode beads. After reverse transcription and preparation for cDNA sequencing, bioinformatic analysis reveals performance quality comparable to microfluidic-based technologies.

2023-03-29 | GSE218209 | GEO

RevGel-seq: instrument-free single-cell RNA sequencing using a reversible hydrogel for cell-specific barcoding [PBMC_perf_10X]

2023-03-29 | GSE218208 | GEO

RevGel-seq: instrument-free single-cell RNA sequencing using a reversible hydrogel for cell-specific barcoding [scRNA-seq_ScaleUp_Scipio]

2023-04-30 | GSE218212 | GEO

RevGel-seq: instrument-free single-cell RNA sequencing using a reversible hydrogel for cell-specific barcoding [scRNA-seq_pancreas_Scipio]

2023-03-29 | GSE218211 | GEO

RevGel-seq: instrument-free single-cell RNA sequencing using a reversible hydrogel for cell-specific barcoding [scRNA-seq_Cardio_Scipio]

2023-03-29 | GSE218210 | GEO

RevGel-seq: instrument-free single-cell RNA sequencing using a reversible hydrogel for cell-specific barcoding [PBMC_perf_Scipio]

2023-03-29 | GSE218207 | GEO

RevGel-seq: instrument-free single-cell RNA sequencing using a reversible hydrogel for cell-specific barcoding [NIH-HEK_perf_Scipio]

Project description:Progress in sample preparation for scRNA-seq is reported based on RevGel-seq, a reversible-hydrogel technology. Barcode bead—cell tandems stabilized by a chemical linker are dispersed in the hydrogel in the liquid state. Upon gelation the tandems are immobilized, cell lysis is triggered by detergent diffusion, and RNA molecules are captured on the adjacent barcode beads. After reverse transcription and preparation for cDNA sequencing, bioinformatic analysis reveals performance quality comparable to microfluidic-based technologies.

2023-03-29 | GSE218206 | GEO

ABLE-SWATH - A Case Study and Methodology for OpenSWATH Parameter Optimization Using the ProCan90 Data Set and 45,810 Computational Analysis Runs

Project description:Proteomic methods typically involve lengthy, multi-step sample preparation protocols (14-16 hrs), especially for the lysis and digestion of solid tissue samples or cells. We developed a streamlined proteomic sample preparation protocol termed Accelerated Barocycler Lysis and Extraction (ABLE), that substantially reduces the time and cost of tissue sample processing. ABLE is based on pressure cycling technology (PCT) for rapid tissue solubilisation and reliable, controlled proteolytic digestion. Here, the previously reported PCT protocol was optimised using 1-4 mg biopsy punches from rat kidney. The tissue denaturant urea was substituted with a combination of sodium deoxycholate (SDC) and N-propanol. ABLE produced comparable numbers of protein identifications in half the sample preparation time and with reduced cost, being ready for MS injection in 3 hrs (vs the conventional urea PCT method of 6 hrs). To validate the method, it was applied across a diverse range of rat tissues (kidney, lung, muscle, brain, testis), human HEK 293 cells and ovarian tumours by coupling PCT with SWATH-mass spectrometry (SWATH-MS). There were similar numbers of quantified proteins between ABLE-SWATH and PCT-SWATH methods, with greater than 70% overlap for all sample types, except muscle with 58% overlap. The ABLE tissue processing protocol offers a standardised, high-throughput, efficient and reproducible proteomic preparation method, accelerating sample throughput for any type of MS analysis. Coupled with SWATH-MS, ABLE has the potential to accelerate proteomics analysis towards a clinically relevant turn-around-time.

2019-01-21 | PXD010688 | 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