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You searched for: EV120065 (EV-TRACK ID)

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Experiment number
  • If needed, multiple experiments were identified in a single publication based on differing sample types, separation protocols and/or vesicle types of interest.
Species
  • Species of origin of the EVs.
Separation protocol
  • Gives a short, non-chronological overview of the different steps of the separation protocol.
    • dUC = differential ultracentrifugation
    • DG = density gradient
    • UF = ultrafiltration
    • SEC = size-exclusion chromatography
Experiment number
  • Experiments differ in Vesicle type
Experiment number
  • Experiments differ in Vesicle type
Details EV-TRACK ID Experiment nr. Species Sample type separation protocol First author Year EV-METRIC
EV120065 1/2 Rattus norvegicus/rattus Cell culture supernatant dUC
UF
Hooper C 2012 22%

Study summary

Full title
All authors
Hooper C, Sainz-Fuertes R, Lynham S, Hye A, Killick R, Warley A, Bolondi C, Pocock J, Lovestone S
Journal
BMC Neurosci
Abstract
BACKGROUND: Microglia, the immune effector cells of the CNS and the signaling molecule Wnt, both pla (show more...)BACKGROUND: Microglia, the immune effector cells of the CNS and the signaling molecule Wnt, both play critical roles in neurodevelopment and neurological disease. Here we describe the inducible release of exosomes from primary cultured rat microglia following treatment with recombinant carrier-free Wnt3a. RESULTS: Wnt3a was internalised into microglia, being detectable in early endosomes, and secreted in exosomes through a GSK3-independent mechanism. Electron microscopy demonstrated that exosomes were elliptical, electron-dense (100 nm) vesicles that coalesced with time in vitro. In contrast to microglia, primary cortical neurons released exosomes constitutively and the quantity of exosomes released was not altered by Wnt3a treatment. The proteomic profile of the microglial-derived exosomes was characterised using liquid chromatography-tandem mass spectrometry (LC/MS/MS) and the vesicles were found to be associated with proteins involved in cellular architecture, metabolism, protein synthesis and protein degradation including ?-actin, glyceraldehyde-3-phosphate dehydrogenase, ribosomal subunits and ubiquitin (45 proteins in total). Unlike lipopolysaccharide, Wnt3a did not induce a neurotoxic, pro-inflammatory phenotype in primary microglia. CONCLUSION: These findings reveal a novel mechanism through which Wnt3a signals in microglia resulting in the release of exosomes loaded with proteinaceous cargo. (hide)
EV-METRIC
22% (49th percentile of all experiments on the same sample type)
 Reported
 Not reported
 Not applicable
EV-enriched proteins
Protein analysis: analysis of three or more EV-enriched proteins
non EV-enriched protein
Protein analysis: assessment of a non-EV-enriched protein
qualitative and quantitative analysis
Particle analysis: implementation of both qualitative and quantitative methods
electron microscopy images
Particle analysis: inclusion of a widefield and close-up electron microscopy image
density gradient
Separation method: density gradient, at least as validation of results attributed to EVs
EV density
Separation method: reporting of obtained EV density
ultracentrifugation specifics
Separation method: reporting of g-forces, duration and rotor type of ultracentrifugation steps
antibody specifics
Protein analysis: antibody clone/reference number and dilution
lysate preparation
Protein analysis: lysis buffer composition
Study data
Sample type
Cell culture supernatant
Sample origin
DNF
Focus vesicles
exosomes
Separation protocol
Separation protocol
  • Gives a short, non-chronological overview of the
    different steps of the separation protocol.
    • dUC = differential ultracentrifugation
    • DG = density gradient
    • UF = ultrafiltration
    • SEC = size-exclusion chromatography
dUC + UF
Protein markers
EV: Alix/ Beta-actin
non-EV:
Proteomics
yes
Show all info
Study aim
Biogenesis/Sorting
Sample
Species
Rattus norvegicus/rattus
Sample Type
Cell culture supernatant
Separation Method
Differential ultracentrifugation
dUC: centrifugation steps
Between 10,000 g and 50,000 g
Between 100,000 g and 150,000 g
Pelleting: time(min)
60
Characterization: Protein analysis
Western Blot
Detected EV-associated proteins
Alix/ Beta-actin
ELISA
Detected EV-associated proteins
Beta-actin
Characterization: Particle analysis
EM
EM-type
transmission EM/ immune EM
Proteïns
Beta-actin
Image type
Wide-field
EV120065 2/2 Rattus norvegicus/rattus Cell culture supernatant dUC
UF
Hooper C 2012 22%

Study summary

Full title
All authors
Hooper C, Sainz-Fuertes R, Lynham S, Hye A, Killick R, Warley A, Bolondi C, Pocock J, Lovestone S
Journal
BMC Neurosci
Abstract
BACKGROUND: Microglia, the immune effector cells of the CNS and the signaling molecule Wnt, both pla (show more...)BACKGROUND: Microglia, the immune effector cells of the CNS and the signaling molecule Wnt, both play critical roles in neurodevelopment and neurological disease. Here we describe the inducible release of exosomes from primary cultured rat microglia following treatment with recombinant carrier-free Wnt3a. RESULTS: Wnt3a was internalised into microglia, being detectable in early endosomes, and secreted in exosomes through a GSK3-independent mechanism. Electron microscopy demonstrated that exosomes were elliptical, electron-dense (100 nm) vesicles that coalesced with time in vitro. In contrast to microglia, primary cortical neurons released exosomes constitutively and the quantity of exosomes released was not altered by Wnt3a treatment. The proteomic profile of the microglial-derived exosomes was characterised using liquid chromatography-tandem mass spectrometry (LC/MS/MS) and the vesicles were found to be associated with proteins involved in cellular architecture, metabolism, protein synthesis and protein degradation including ?-actin, glyceraldehyde-3-phosphate dehydrogenase, ribosomal subunits and ubiquitin (45 proteins in total). Unlike lipopolysaccharide, Wnt3a did not induce a neurotoxic, pro-inflammatory phenotype in primary microglia. CONCLUSION: These findings reveal a novel mechanism through which Wnt3a signals in microglia resulting in the release of exosomes loaded with proteinaceous cargo. (hide)
EV-METRIC
22% (49th percentile of all experiments on the same sample type)
 Reported
 Not reported
 Not applicable
EV-enriched proteins
Protein analysis: analysis of three or more EV-enriched proteins
non EV-enriched protein
Protein analysis: assessment of a non-EV-enriched protein
qualitative and quantitative analysis
Particle analysis: implementation of both qualitative and quantitative methods
electron microscopy images
Particle analysis: inclusion of a widefield and close-up electron microscopy image
density gradient
Separation method: density gradient, at least as validation of results attributed to EVs
EV density
Separation method: reporting of obtained EV density
ultracentrifugation specifics
Separation method: reporting of g-forces, duration and rotor type of ultracentrifugation steps
antibody specifics
Protein analysis: antibody clone/reference number and dilution
lysate preparation
Protein analysis: lysis buffer composition
Study data
Sample type
Cell culture supernatant
Sample origin
DNF
Focus vesicles
exosomes
Separation protocol
Separation protocol
  • Gives a short, non-chronological overview of the
    different steps of the separation protocol.
    • dUC = differential ultracentrifugation
    • DG = density gradient
    • UF = ultrafiltration
    • SEC = size-exclusion chromatography
dUC + UF
Protein markers
EV: Alix/ Beta-actin
non-EV:
Proteomics
no
Show all info
Study aim
Biogenesis/Sorting
Sample
Species
Rattus norvegicus/rattus
Sample Type
Cell culture supernatant
Separation Method
Differential ultracentrifugation
dUC: centrifugation steps
Between 10,000 g and 50,000 g
Between 100,000 g and 150,000 g
Equal to or above 150,000 g
Pelleting: time(min)
60
Characterization: Protein analysis
Western Blot
Detected EV-associated proteins
Alix/ Beta-actin
ELISA
Detected EV-associated proteins
Beta-actin
Characterization: Particle analysis
1 - 2 of 2
  • CM = Commercial method
  • dUC = differential ultracentrifugation
  • DG = density gradient
  • UF = ultrafiltration
  • SEC = size-exclusion chromatography
EV-TRACK ID
EV120065
species
Rattus
norvegicus/rattus
sample type
Cell culture
separation protocol
dUC
UF
dUC
UF
Exp. nr.
1
2
EV-METRIC %
22
22