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You searched for: EV210043 (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.
    • (d)(U)C = (differential) (ultra)centrifugation
    • DG = density gradient
    • UF = ultrafiltration
    • SEC = size-exclusion chromatography
    • IAF = immuno-affinity capture
Details EV-TRACK ID Experiment nr. Species Sample type Separation protocol First author Year EV-METRIC
EV210043 1/2 Homo sapiens Cell culture supernatant DG
(d)(U)C
UF
Filtration
Fikatas, Antonios 2021 100%

Study summary

Full title
All authors
Antonios Fikatas, Jonas Dehairs, Sam Noppen, Jordi Doijen, Frank Vanderhoydonc, Eef Meyen, Johannes V Swinnen, Christophe Pannecouque, Dominique Schols
Journal
Viruses
Abstract
To date, no vaccines or antivirals are available against Zika virus (ZIKV). In addition, the mechani (show more...)To date, no vaccines or antivirals are available against Zika virus (ZIKV). In addition, the mechanisms underlying ZIKV-associated pathogenesis of the central nervous system (CNS) are largely unexplored. Getting more insight into the cellular pathways that ZIKV recruits to facilitate infection of susceptible cells will be crucial for establishing an effective treatment strategy. In general, cells secrete a number of vesicles, known as extracellular vesicles (EVs), in response to viral infections. These EVs serve as intercellular communicators. Here, we investigated the role of EVs derived from ZIKV-infected human brain microvascular endothelial cells on the blood–brain barrier (BBB) system. We demonstrated that ZIKV-infected EVs (IEVs) can incorporate viral components, including ZIKV RNA, NS1, and E-protein, and further transfer them to several types of CNS cells. Using label-free impedance-based biosensing, we observed that ZIKV and IEVs can temporally disturb the monolayer integrity of BBB-mimicking cells, possibly by inducing structural rearrangements of the adherent protein VE-cadherin (immunofluorescence staining). Finally, differences in the lipidomic profile between EVs and their parental cells possibly suggest a preferential sorting mechanism of specific lipid species into the vesicles. To conclude, these data suggest that IEVs could be postulated as vehicles (Trojan horse) for ZIKV transmission via the BBB. (hide)
EV-METRIC
100% (99th 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. For the quantitative method, the reporting of measured EV concentration is expected.
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
Control condition
Focus vesicles
extracellular vesicle
Separation protocol
Separation protocol
  • Gives a short, non-chronological overview of the
    different steps of the separation protocol.
    • dUC = (Differential) (ultra)centrifugation
    • DG = density gradient
    • UF = ultrafiltration
    • SEC = size-exclusion chromatography
    • IAF = immuno-affinity capture
DG
(d)(U)C
UF
Filtration
Protein markers
EV: TSG101/ Alix/ CD63
non-EV: Calnexin
Proteomics
no
EV density (g/ml)
1.06-1.16
Show all info
Study aim
Function/Identification of content (omics approaches)
Sample
Species
Homo sapiens
Sample Type
Cell culture supernatant
EV-producing cells
hcMEC/D3
EV-harvesting Medium
EV-depleted medium
Preparation of EDS
Commercial EDS
Cell viability (%)
NA
Separation Method
(Differential) (ultra)centrifugation
dUC: centrifugation steps
Below or equal to 800 g
Between 800 g and 10,000 g
Pelleting performed
Yes
Pelleting: time(min)
1260
Pelleting: rotor type
TH-641
Pelleting: speed (g)
100 000
Wash: volume per pellet (ml)
10
Wash: time (min)
180
Wash: Rotor Type
TH-641
Wash: speed (g)
100 000
Density gradient
Type
Discontinuous
Number of initial discontinuous layers
4
Lowest density fraction
5%
Highest density fraction
40%
Total gradient volume, incl. sample (mL)
10
Sample volume (mL)
1
Orientation
Top-down
Rotor type
TH-641
Speed (g)
100 000
Duration (min)
1080
Fraction volume (mL)
1,5
Fraction processing
None
Filtration steps
0.22µm or 0.2µm
Ultra filtration
Cut-off size (kDa)
10
Membrane type
Regenerated cellulose
Characterization: Protein analysis
Protein Concentration Method
Bradford
Western Blot
Antibody details provided?
No
Detected EV-associated proteins
CD63/ TSG101/ Alix
Not detected contaminants
Calnexin
Characterization: Lipid analysis
Yes
Characterization: Particle analysis
NTA
Report type
Mean
Reported size (nm)
165
EV concentration
Yes
Particle yield
Yes, as number of particles per milliliter of starting sample 1,50E+08
EM
EM-type
Transmission-EM
Image type
Close-up, Wide-field
Report size (nm)
130
EV210043 2/2 Homo sapiens Cell culture supernatant DG
(d)(U)C
UF
Filtration
Fikatas, Antonios 2021 89%

Study summary

Full title
All authors
Antonios Fikatas, Jonas Dehairs, Sam Noppen, Jordi Doijen, Frank Vanderhoydonc, Eef Meyen, Johannes V Swinnen, Christophe Pannecouque, Dominique Schols
Journal
Viruses
Abstract
To date, no vaccines or antivirals are available against Zika virus (ZIKV). In addition, the mechani (show more...)To date, no vaccines or antivirals are available against Zika virus (ZIKV). In addition, the mechanisms underlying ZIKV-associated pathogenesis of the central nervous system (CNS) are largely unexplored. Getting more insight into the cellular pathways that ZIKV recruits to facilitate infection of susceptible cells will be crucial for establishing an effective treatment strategy. In general, cells secrete a number of vesicles, known as extracellular vesicles (EVs), in response to viral infections. These EVs serve as intercellular communicators. Here, we investigated the role of EVs derived from ZIKV-infected human brain microvascular endothelial cells on the blood–brain barrier (BBB) system. We demonstrated that ZIKV-infected EVs (IEVs) can incorporate viral components, including ZIKV RNA, NS1, and E-protein, and further transfer them to several types of CNS cells. Using label-free impedance-based biosensing, we observed that ZIKV and IEVs can temporally disturb the monolayer integrity of BBB-mimicking cells, possibly by inducing structural rearrangements of the adherent protein VE-cadherin (immunofluorescence staining). Finally, differences in the lipidomic profile between EVs and their parental cells possibly suggest a preferential sorting mechanism of specific lipid species into the vesicles. To conclude, these data suggest that IEVs could be postulated as vehicles (Trojan horse) for ZIKV transmission via the BBB. (hide)
EV-METRIC
89% (98th 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. For the quantitative method, the reporting of measured EV concentration is expected.
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
ZIKV-infected
Focus vesicles
extracellular vesicle
Separation protocol
Separation protocol
  • Gives a short, non-chronological overview of the
    different steps of the separation protocol.
    • dUC = (Differential) (ultra)centrifugation
    • DG = density gradient
    • UF = ultrafiltration
    • SEC = size-exclusion chromatography
    • IAF = immuno-affinity capture
DG
(d)(U)C
UF
Filtration
Protein markers
EV: TSG101/ Alix/ CD63
non-EV: Calnexin
Proteomics
no
EV density (g/ml)
1.06-1.16
Show all info
Study aim
Function/Identification of content (omics approaches)
Sample
Species
Homo sapiens
Sample Type
Cell culture supernatant
EV-producing cells
hcMEC/D3
EV-harvesting Medium
EV-depleted medium
Preparation of EDS
Commercial EDS
Cell viability (%)
NA
Separation Method
(Differential) (ultra)centrifugation
dUC: centrifugation steps
Below or equal to 800 g
Between 800 g and 10,000 g
Pelleting performed
Yes
Pelleting: time(min)
1260
Pelleting: rotor type
TH-641
Pelleting: speed (g)
100 000
Wash: volume per pellet (ml)
10
Wash: time (min)
180
Wash: Rotor Type
TH-641
Wash: speed (g)
100 000
Density gradient
Type
Discontinuous
Number of initial discontinuous layers
4
Lowest density fraction
5%
Highest density fraction
40%
Total gradient volume, incl. sample (mL)
10
Sample volume (mL)
1
Orientation
Top-down
Rotor type
TH-641
Speed (g)
100 000
Duration (min)
1080
Fraction volume (mL)
1,5
Fraction processing
None
Filtration steps
0.22µm or 0.2µm
Ultra filtration
Cut-off size (kDa)
10
Membrane type
Regenerated cellulose
Characterization: Protein analysis
Protein Concentration Method
Bradford
Western Blot
Antibody details provided?
No
Detected EV-associated proteins
CD63/ TSG101/ Alix
Not detected contaminants
Calnexin
Characterization: Lipid analysis
Yes
Characterization: Particle analysis
NTA
Report type
Mean
Reported size (nm)
175
EV concentration
Yes
Particle yield
Yes, as number of particles per milliliter of starting sample 1,40E+08
EM
EM-type
Transmission-EM
Image type
Wide-field
Report size (nm)
130
1 - 2 of 2
  • CM = Commercial method
  • dUC = differential ultracentrifugation
  • DG = density gradient
  • UF = ultrafiltration
  • SEC = size-exclusion chromatography
EV-TRACK ID
EV210043
species
Homo sapiens
sample type
Cell culture
cell type
hcMEC/D3
condition
Control condition
ZIKV-infected
separation protocol
DG
(d)(U)C
UF
Filtration
DG
(d)(U)C
UF
Filtration
Exp. nr.
1
2
EV-METRIC %
100
89