Mesenchymal Stem Cell-Derived Extracellular Vesicle Isolation and Their Protein Cargo Characterization

Methods Mol Biol. 2021:2259:3-12. doi: 10.1007/978-1-0716-1178-4_1.

Abstract

In the present protocol, extracellular vesicles (EVs) released from a primary culture of human umbilical cord mesenchymal stem cells (MSCs) were isolated by ultracentrifugation processes, characterized by transmission electron microscopy (TEM) and measured by nanoparticle tracking analysis (NTA). Protein was extracted from EVs using RIPA buffer and then was assessed for integrity. The proteomic content of the total EV protein samples was analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS) after labeling by tandem mass tag (TMT). This combined approach allowed the development of an effective strategy to study the protein cargo from MSC-derived EVs.

Keywords: Extracellular vesicles; Mesenchymal stem cells; Tandem mass tag; Transmission electron microscopy.

MeSH terms

  • Cells, Cultured
  • Chromatography, Liquid / methods
  • Culture Media / chemistry
  • Extracellular Vesicles / chemistry*
  • Extracellular Vesicles / ultrastructure*
  • Humans
  • Mesenchymal Stem Cells / chemistry
  • Mesenchymal Stem Cells / cytology*
  • Microscopy, Electron, Transmission / methods
  • Primary Cell Culture / methods
  • Proteins / analysis*
  • Proteins / isolation & purification
  • Proteomics / methods
  • Tandem Mass Spectrometry / methods
  • Umbilical Cord / cytology

Substances

  • Culture Media
  • Proteins