Skip to Main content Skip to Navigation
Theses

Caractérisation, quantification et isolation de vésicules extracellulaires du plasma sanguin à l’aide de nanoparticules d’or ou magnétiques conjuguées à des protéines

Abstract : Extracellular vesicles (EVs) are submicrometric membrane vesicles found in body fluids and produced by cells in response to various stimuli. EVs are involved in numerous physiological processes but also in pathologies as cancers or cardiovascular diseases. Even if EVs are largely studied, our knowledge about them remains limited. This is mainly caused by the difficulties to characterize EVs and by the lack of standardized methods allowing their characterization. The first part of my PhD work focused on the development and optimization of a protein thiolation method. Antibodies modified to expose few thiols were conjugated to gold nanoparticles functionalized with maleimides. The binding of thiolated antibodies to gold nanoparticles was quantitatively studied and optimal conjugation conditions were determined using biochemical methods. The second part of my PhD work concerned the characterization of blood plasma EVs from healthy subjects using transmission electron microscopy (TEM). EVs morphology, size and phenotype were determined by cryo-TEM combined with labelling with protein-conjugated gold nanoparticles. The near-absolute quantification of blood plasma EVs was achieved using an original TEM method based on the direct sedimentation of EVs onto TEM grids. The third part of this study consisted in developing an EV isolation method using AnxA5-conjugated magnetic particles. Conditions allowing total extraction of blood plasma phosphatidylserine-exposing EVs were determined using flow cytometry (FC). This study presents a detailed characterization of blood plasma EVs from healthy subjects and can serve as a reference for future studies on EVs contained in pathological plasmas or other body fluids.
Complete list of metadatas

https://tel.archives-ouvertes.fr/tel-01483569
Contributor : Abes Star :  Contact
Submitted on : Monday, March 6, 2017 - 10:25:10 AM
Last modification on : Thursday, September 3, 2020 - 5:14:23 AM
Long-term archiving on: : Wednesday, June 7, 2017 - 1:18:45 PM

File

LINARES_ROMAIN_2016.pdf
Version validated by the jury (STAR)

Identifiers

  • HAL Id : tel-01483569, version 1

Collections

Citation

Romain Linares. Caractérisation, quantification et isolation de vésicules extracellulaires du plasma sanguin à l’aide de nanoparticules d’or ou magnétiques conjuguées à des protéines. Hématologie. Université de Bordeaux, 2016. Français. ⟨NNT : 2016BORD0344⟩. ⟨tel-01483569⟩

Share

Metrics

Record views

575

Files downloads

443