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Details | EV-TRACK ID | Experiment nr. | Species | Sample type | separation protocol | First author | Year | EV-METRIC |
---|---|---|---|---|---|---|---|---|
EV170030 | 1/1 | Mus musculus | Cell culture supernatant |
DG (d)(U)C |
Driedonks, Tom A. P. | 2018 | 100% | |
Study summaryFull title
All authors
Driedonks TAP, van der Grein SG, Ariyurek Y, Buermans HPJ, Jekel H, Chow FWN, Wauben MHM, Buck AH, 't Hoen PAC, Nolte-'t Hoen ENM
Journal
Cell Mol Life Sci
Abstract
The release and uptake of nano-sized extracellular vesicles (EV) is a highly conserved means of inte (show more...)
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
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 dataSample type
Cell culture supernatant
Cell Name
primary bone marrow dendritic cells
Sample origin
Control condition
Focus vesicles
extracellular vesicle
Separation protocol
Separation protocol
DG
(d)(U)C Adj. k-factor
253.9 (pelleting)
Protein markers
EV: MHC2/ CD63/ CD9/ Galectin-3
non-EV: beta-actin Proteomics
no
Show all info
Study aim
Biomarker, Biogenesis/cargo sorting, Identification of content (omics approaches)
Sample
Species
Mus musculus
Sample Type
Cell culture supernatant
Sample Condition
Control condition
EV-producing cells
primary bone marrow dendritic cells
EV-harvesting Medium
EV-depleted serum
Preparation of EDS
overnight (16h) at >=100,000g
Cell viability
90
Separation Method
Differential ultracentrifugation
centrifugation steps
Below or equal to 800 g
Between 10,000 g and 50,000 g Between 100,000 g and 150,000 g Obtain an EV pellet :
Yes
Pelleting: time(min)
65
Pelleting: rotor type
SW 28
Pelleting: speed (g)
100000
Pelleting: adjusted k-factor
253.9
Density gradient
Density medium
Sucrose
Type
Continuous
Lowest density fraction
0.4M
Highest density fraction
2.5M
Sample volume (mL)
0.15
Orientation
Bottom-up (sample migrates upwards)
Rotor type
SW 40 Ti
Speed (g)
192000
Duration (min)
900-1080
Fraction volume (mL)
1
Fraction processing
Centrifugation
Pelleting: volume per fraction
1
Pelleting: duration (min)
65
Pelleting: rotor type
SW 40 Ti
Pelleting: speed (g)
192000
Pelleting: adjusted k-factor
144.0
Characterization: Protein analysis
Protein Concentration Method
Not determined
Western Blot
Antibody details provided?
Yes
Lysis buffer provided?
Yes
Detected EV-associated proteins
CD9, CD63, MHC2, Galectin-3
Not detected contaminants
beta-actin
Characterization: Particle analysis
NTA
Report type
Size range/distribution
Reported size (nm)
100 - 200
EV concentration
Yes
Particle analysis: flow cytometry
Flow cytometer type
BD Influx
Hardware adjustment
see van der Vlist et al. 2012 Nature Protocols;and Nolte-'t Hoen 2012 Nanomedicine
Calibration bead size
0.1,0.2
EV concentration
Yes
EM
EM-type
Transmission-EM
Image type
Close-up, Wide-field
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