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You searched for: EV210180 (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 code Experiment nr. Species Sample type Separation protocol First author Year EV-METRIC
EV210180 1/2 Homo sapiens Cell culture supernatant (d)(U)C Santos, Mark F 2021 78%

Study summary

Full title
All authors
Mark F. Santos, Germana Rappa, Jana Karbanová, Simona Fontana, Maria Antonietta Di Bella, Marshall R. Pope, Barbara Parrino, Stella Maria Cascioferro, Giulio Vistoli, Patrizia Diana, Girolamo Cirrincione, Goffredo O. Arena, Gyunghwi Woo, Kevin Huang, Tony Huynh, Marta Moschetti, Riccardo Alessandro, Denis Corbeil, Aurelio Lorico
Journal
J Extracell Vesicles
Abstract
Extracellular vesicles (EVs) are mediators of intercellular communication under both healthy and pat (show more...)Extracellular vesicles (EVs) are mediators of intercellular communication under both healthy and pathological conditions, including the induction of pro-metastatic traits, but it is not yet known how and where functional cargoes of EVs are delivered to their targets in host cell compartments. We have described that after endocytosis, EVs reach Rab7+ late endosomes and a fraction of these enter the nucleoplasmic reticulum and transport EV biomaterials to the host cell nucleoplasm. Their entry therein and docking to outer nuclear membrane occur through a tripartite complex formed by the proteins VAP-A, ORP3 and Rab7 (VOR complex). Here, we report that the antifungal compound itraconazole (ICZ), but not its main metabolite hydroxy-ICZ or ketoconazole, disrupts the binding of Rab7 to ORP3–VAP-A complexes, leading to inhibition of EV-mediated pro-metastatic morphological changes including cell migration behaviour of colon cancer cells. With novel, smaller chemical drugs, inhibition of the VOR complex was maintained, although the ICZ moieties responsible for antifungal activity and interference with intracellular cholesterol distribution were removed. Knowing that cancer cells hijack their microenvironment and that EVs derived from them determine the pre-metastatic niche, small-sized inhibitors of nuclear transfer of EV cargo into host cells could find cancer therapeutic applications, particularly in combination with direct targeting of cancer cells. (hide)
EV-METRIC
78% (96th 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
(d)(U)C
Protein markers
EV: CD63/ CD81/ Alix/ CD9
non-EV: Histone H1/ Calnexin
Proteomics
no
Show all info
Study aim
Function/Mechanism of uptake/transfer
Sample
Species
Homo sapiens
Sample Type
Cell culture supernatant
EV-producing cells
SW620
EV-harvesting Medium
Serum free medium
Separation Method
(Differential) (ultra)centrifugation
dUC: centrifugation steps
Below or equal to 800 g
Between 800 g and 10,000 g
Between 10,000 g and 50,000 g
Equal to or above 150,000 g
Pelleting performed
Yes
Pelleting: time(min)
60
Pelleting: rotor type
SW 55 Ti
Pelleting: speed (g)
200000
Characterization: Protein analysis
Protein Concentration Method
microBCA
Western Blot
Antibody details provided?
No
Detected EV-associated proteins
CD9/ CD63/ Alix/ CD81
Not detected contaminants
Histone H1/ Calnexin
Detected EV-associated proteins
CD9/ CD63/ CD81
Characterization: Lipid analysis
No
Characterization: Particle analysis
NTA
Report type
Mean
Reported size (nm)
150
EV concentration
Yes
Particle yield
Yes, as number of particles per milliliter of starting sample 4.00E+10
EM
EM-type
Transmission-EM
Image type
Close-up, Wide-field
Report size (nm)
60-130
EV210180 2/2 Homo sapiens Cell culture supernatant (d)(U)C Santos, Mark F 2021 78%

Study summary

Full title
All authors
Mark F. Santos, Germana Rappa, Jana Karbanová, Simona Fontana, Maria Antonietta Di Bella, Marshall R. Pope, Barbara Parrino, Stella Maria Cascioferro, Giulio Vistoli, Patrizia Diana, Girolamo Cirrincione, Goffredo O. Arena, Gyunghwi Woo, Kevin Huang, Tony Huynh, Marta Moschetti, Riccardo Alessandro, Denis Corbeil, Aurelio Lorico
Journal
J Extracell Vesicles
Abstract
Extracellular vesicles (EVs) are mediators of intercellular communication under both healthy and pat (show more...)Extracellular vesicles (EVs) are mediators of intercellular communication under both healthy and pathological conditions, including the induction of pro-metastatic traits, but it is not yet known how and where functional cargoes of EVs are delivered to their targets in host cell compartments. We have described that after endocytosis, EVs reach Rab7+ late endosomes and a fraction of these enter the nucleoplasmic reticulum and transport EV biomaterials to the host cell nucleoplasm. Their entry therein and docking to outer nuclear membrane occur through a tripartite complex formed by the proteins VAP-A, ORP3 and Rab7 (VOR complex). Here, we report that the antifungal compound itraconazole (ICZ), but not its main metabolite hydroxy-ICZ or ketoconazole, disrupts the binding of Rab7 to ORP3–VAP-A complexes, leading to inhibition of EV-mediated pro-metastatic morphological changes including cell migration behaviour of colon cancer cells. With novel, smaller chemical drugs, inhibition of the VOR complex was maintained, although the ICZ moieties responsible for antifungal activity and interference with intracellular cholesterol distribution were removed. Knowing that cancer cells hijack their microenvironment and that EVs derived from them determine the pre-metastatic niche, small-sized inhibitors of nuclear transfer of EV cargo into host cells could find cancer therapeutic applications, particularly in combination with direct targeting of cancer cells. (hide)
EV-METRIC
78% (96th 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
CD9-GFP expression
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
(d)(U)C
Protein markers
EV: CD63/ CD81/ Alix/ CD9
non-EV: Histone H1/ Calnexin
Proteomics
no
Show all info
Study aim
Function/Mechanism of uptake/transfer
Sample
Species
Homo sapiens
Sample Type
Cell culture supernatant
EV-producing cells
FEMX-I
EV-harvesting Medium
Serum free medium
Separation Method
(Differential) (ultra)centrifugation
dUC: centrifugation steps
Below or equal to 800 g
Between 800 g and 10,000 g
Between 10,000 g and 50,000 g
Equal to or above 150,000 g
Pelleting performed
Yes
Pelleting: time(min)
60
Pelleting: rotor type
SW 55 Ti
Pelleting: speed (g)
200000
Characterization: Protein analysis
Protein Concentration Method
microBCA
Western Blot
Antibody details provided?
No
Detected EV-associated proteins
CD9/ CD63/ Alix/ CD81
Not detected contaminants
Histone H1/ Calnexin
Detected EV-associated proteins
CD9/ CD63
Characterization: Lipid analysis
No
Characterization: Particle analysis
NTA
Report type
Mean
Reported size (nm)
120
EV concentration
Yes
Particle yield
Yes, as number of particles per milliliter of starting sample 2.00E+10
EM
EM-type
Transmission-EM
Image type
Close-up, Wide-field
Report size (nm)
40-100
1 - 2 of 2
  • CM = Commercial method
  • dUC = differential ultracentrifugation
  • DG = density gradient
  • UF = ultrafiltration
  • SEC = size-exclusion chromatography
EV-TRACK ID
EV210180
species
Homo sapiens
sample type
Cell culture
cell type
SW620
FEMX-I
condition
Control condition
CD9-GFP expression
separation protocol
(d)(U)C
(d)(U)C
Exp. nr.
1
2
EV-METRIC %
78
78