PXD045737

PXD045737 is an original dataset announced via ProteomeXchange.

Title	Efficient enzyme-free isolation of brain-derived extracellular vesicles
Description	Extracellular vesicles (EVs) have gained significant attention as potential diagnostic and therapeutic tools due to their role in intercellular communication and their ability to encapsulate various biomolecules. However, the isolation of EVs from tissue remains challenging, often requiring enzymatic digestion steps that may introduce unwanted biases and affect EV integrity. In this study, we present an efficient non-enzymatic approach for the isolation of brain-derived extracellular vesicles (BDEVs). Our method employs sequential ultracentrifugation and a density-based gradient to achieve a high yield and purity of BDEVs from brain tissue without any enzymatic digestion. Characterization of the isolated EVs revealed their typical morphology and size distribution, as confirmed by transmission electron microscopy and nanoparticle tracking analysis. We show by western blot that known BDEV markers, such as Flotlin-1, CD81, Alix, and the prion protein (PrP) are present without proteolytic cuts introduced by the enzymatic digestion of the tissue. We also show that the absence ofGM130 ,widely accepted as a negative marker to assess BDEV’s purity, is due to enzymatic digestion rather than lack of this protein in the sample. Moreover, we find no differences in mRNA content between the non-enzymatically and the enzymatically isolated BDEVs, showing that the intraluminal part of the BDEVs is not affected by enzymatic isolation. Similarly, our proteomic analysis shows unaltered protein expression in BDEVs isolated with both methods. However, other proteins, potentially in the BDEVs corona and membrane, are differentially expressed, highlighting the consequences of the enzymatically-based isolation protocol.
HostingRepository	PRIDE
AnnounceDate	2024-11-07
AnnouncementXML	Submission_2024-11-07_07:10:10.726.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Bente Siebels
SpeciesList	scientific name: Mus musculus (Mouse); NCBI TaxID: 10090; scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationList	iodoacetamide derivatized residue
Instrument	Q Exactive; Orbitrap Fusion

Revision	Datetime	Status	ChangeLog Entry
0	2023-09-27 08:47:24	ID requested
⏵ 1	2024-11-07 07:10:11	announced

10.1002/JEV2.70011;

submitter keyword: collagenase-free protocol, prion protein,Brain-derived EVs, BDEVs

Mohsin Shafiq
contact affiliation	Institute of Neuropathology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
contact email	m.shafiq@uke.de
lab head
Bente Siebels
contact affiliation	Section for Mass spectrometry and Proteomics, University Medical Center Hamburg-Eppendorf
contact email	b.siebels@uke.de
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD045737
     1. Label: PRIDE project
     2. Name: Efficient enzyme-free isolation of brain-derived extracellular vesicles