title: Nanoparticle tracking analysis monitors microvesicle and exosome secretion from immune cells.
abstract: Nanoparticle tracking analysis permits the determination of both the size distribution and relative concentration of microvesicles, including exosomes, in the supernatants of cultured cells and biological fluids. We have studied the release of microvesicles from the human lymphoblastoid T-cell lines Jurkat and CEM. Unstimulated, both cell lines release microvesicles in the size range 70-90 nm, which can be depleted from the supernatant by ultracentrifugation at 100 000 g, and by anti-CD45 magnetic beads, and which by immunoblotting also contain the exosome-associated proteins Alix and Tsg101. Incubation with known potentiators of exosome release, the ionophores monensin and A23187, resulted in a significant increase in microvesicle release that was both time and concentration dependent. Mass spectrometric analysis of proteins isolated from ultracentrifuged supernatants of A23187-treated cells revealed the presence of exosome-associated proteins including heat-shock protein 90, tubulin, elongation factor α1, actin and glyceraldehyde 3-phosphate dehydrogenase. Additionally, treatment of peripheral blood monocyte-derived dendritic cells with bacterial lipopolysaccharide displayed an increase in secreted microvesicles. Consequently, nanoparticle tracking analysis can be effectively applied to monitor microvesicle release from cells of the immune system.
authors.affiliation: School of Medicine, University of St Andrews, Fife, UK.
authors.forename: Chin Y
authors.initials: CY
authors.lastname: Soo
authors.forename: Yaqiong
authors.initials: Y
authors.lastname: Song
authors.forename: Ying
authors.initials: Y
authors.lastname: Zheng
authors.forename: Elaine C
authors.initials: EC
authors.lastname: Campbell
authors.forename: Andrew C
authors.initials: AC
authors.lastname: Riches
authors.forename: Frank
authors.initials: F
authors.lastname: Gunn-Moore
authors.forename: Simon J
authors.initials: SJ
authors.lastname: Powis
mesh_terms: D000001:Calcimycin / Q000494:pharmacology; D002135:Calcium-Binding Proteins / Q000737:chemistry; D018797:Cell Cycle Proteins / Q000737:chemistry; D002460:Cell Line; D058948:Cell Tracking / Q000379:methods*; D004268:DNA-Binding Proteins / Q000737:chemistry; D003713:Dendritic Cells / Q000187:drug effects; D056827:Endosomal Sorting Complexes Required for Transport / Q000737:chemistry; D055354:Exosomes / Q000187:drug effects / Q000276:immunology*; D006801:Humans; D018189:Immunomagnetic Separation; D007476:Ionophores / Q000494:pharmacology; D017493:Leukocyte Common Antigens / Q000737:chemistry; D008070:Lipopolysaccharides / Q000494:pharmacology; D008985:Monensin / Q000494:pharmacology; D053758:Nanoparticles; D013601:T-Lymphocytes / Q000187:drug effects / Q000276:immunology*; D014157:Transcription Factors / Q000737:chemistry; D000099177:Tsg101 Protein
chemical_list: D000001:Calcimycin; D002135:Calcium-Binding Proteins; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D056827:Endosomal Sorting Complexes Required for Transport; D007476:Ionophores; D017493:Leukocyte Common Antigens; D008070:Lipopolysaccharides; D008985:Monensin; D014157:Transcription Factors; D000099177:Tsg101 Protein; C117020:PDCD6IP protein, human; C516769:PTPRC protein, human