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You searched for: EV180053 (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
EV180053 1/1 Homo sapiens Cell culture supernatant (d)(U)C
Filtration
Rocha, Sara 2018 75%

Study summary

Full title
All authors
Sara Rocha, Joana Carvalho, Patrícia Oliveira, Maren Voglstaetter, Domitille Schvartz, Andreas R. Thomsen, Nadia Walter, Richa Khanduri, Jean‐Charles Sanchez, Andreas Keller, Carla Oliveira, Irina Nazarenko
Journal
Advanced Science
Abstract
The success of malignant tumors is conditioned by the intercellular communication between tumor cell (show more...)The success of malignant tumors is conditioned by the intercellular communication between tumor cells and their microenvironment, with extracellular vesicles (EVs) acting as main mediators. While the value of 3D conditions to study tumor cells is well established, the impact of cellular architecture on EV content and function is not investigated yet. Here, a recently developed 3D cell culture microwell array is adapted for EV production and a comprehensive comparative analysis of biochemical features, RNA and proteomic profiles of EVs secreted by 2D vs 3D cultures of gastric cancer cells, is performed. 3D cultures are significantly more efficient in producing EVs than 2D cultures. Global upregulation of microRNAs and downregulation of proteins in 3D are observed, indicating their dynamic coregulation in response to cellular architecture, with the ADP‐ribosylation factor 6 signaling pathway significantly downregulated in 3D EVs. The data strengthen the biological relevance of cellular architecture for production and cargo of EVs. (hide)
EV-METRIC
75% (94th 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
Cell Name
MKN74, MKN45
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.
    • (d)(U)C = (differential) (ultra)centrifugation
    • DG = density gradient
    • UF = ultrafiltration
    • SEC = size-exclusion chromatography
    • IAF = immuno-affinity capture
(d)(U)C
Filtration
Protein markers
EV: integrin-alpha6/ catenin-beta1/ integrin-alpha2/ integrin-alpha3/ tubulin/ Flotillin-1/ E-cadherin/ GAPDH/ CD81/ integrin-beta1/ catenin-delta1/ CD9
non-EV: CytochromeC
Proteomics
yes
Show all info
Study aim
New methodological development, Identification of content (omics approaches)
Sample
Species
Homo sapiens
Sample Type
Cell culture supernatant
Sample Condition
Control condition
EV-producing cells
MKN74, MKN45
EV-harvesting Medium
Serum free medium
Cell viability
95
Separation Method
Differential ultracentrifugation
centrifugation steps
Below or equal to 800 g
Between 800 g and 10,000 g
Between 10,000 g and 50,000 g
Between 100,000 g and 150,000 g
Filtration steps
0.22µm or 0.2µm
Characterization: Protein analysis
Protein Concentration Method
Bradford
Protein Concentration
0.2
Western Blot
Antibody details provided?
Yes
Lysis buffer provided?
Yes
Detected EV-associated proteins
integrin-alpha2, integrin-alpha3, integrin-alpha6, integrin-beta1, E-cadherin, catenin-beta1, catenin-delta1, GAPDH, tubulin
Proteomics
Proteomics database
Yes
Characterization: RNA analysis
Proteinase treatment
Moment of Proteinase treatment
After
Proteinase type
Proteinase K
Proteinase concentration
5
RNAse treatment
Moment of RNAse treatment
After
RNAse type
RNase A
RNAse concentration
1
Characterization: Particle analysis
NTA
Report type
Size range/distribution
Reported size (nm)
110
EV concentration
Yes
Particle yield
1.50E+09 particles/million cells
Particle analysis: flow cytometry
Flow cytometer type
Amnis ImageStream
Hardware adjustment
Calibration bead size
200
EM
EM-type
Transmission-EM
Image type
Close-up, Wide-field
Report size (nm)
110
1 - 1 of 1
  • CM = Commercial method
  • dUC = differential ultracentrifugation
  • DG = density gradient
  • UF = ultrafiltration
  • SEC = size-exclusion chromatography
EV-TRACK ID
EV180053
species
Homo sapiens
sample type
Cell culture
cell type
MKN74
MKN45
condition
Control condition
separation protocol
(d)(U)C
Filtration
Exp. nr.
1
EV-METRIC %
75