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Experiment number
  • If needed, multiple experiments were identified in a single publication based on differing sample types, isolation protocols and/or vesicle types of interest.
Species
  • Species of origin of the EVs.
Isolation protocol
  • Gives a short, non-chronological overview of the different steps of the isolation protocol.
    • dUC = differential ultracentrifugation
    • DG = density gradient
    • UF = ultrafiltration
    • SEC = size-exclusion chromatography
Details EV-TRACK ID Experiment nr. Species Sample type Isolation protocol First author Year EV-METRIC
EV130042 1/1 Mus musculus Cell culture supernatant 0.2 µm filter
dUC
Sucrose-DG (valid.)
Kotzerke K 2013 56%

Study summary

Full title
All authors
Kotzerke K, Mempel M, Aung T, Wulf GG, Urlaub H, Wenzel D, Schön MP, Braun A
Journal
Exp Dermatol
Abstract
It has long been known that keratinocytes influence cutaneous immunity through secretion of soluble (show more...)It has long been known that keratinocytes influence cutaneous immunity through secretion of soluble factors. Exosomes, small membrane vesicles of endocytotic origin, have been implicated in intercellular communication processes such as the transfer of tumor cell antigens and the activation of recipient dendritic cells (DC). However, little is known about immunomodulatory functions of keratinocyte-derived exosomes. To address this question, we analysed exosome secretion of the murine keratinocyte cell line MPEK under steady state as well as inflammatory conditions (+/- IFN?). These exosomes were readily taken up by bone marrow-derived DC (BMDC) in vitro resulting in a matured phenotype, as evidenced by increased CD40 expression as well as by the production of large amounts of IL-6, IL-10 and IL-12. When the transfer of antigen-specific information through exosomes was investigated, it was found that keratinocytes took up antigen (ovalbumin) and transferred it to their exosomes. However, these antigen-harbouring exosomes failed to induce antigen-specific T cell responses via BMDC. Together, this novel biological function suggests that keratinocytes are able to direct unspecific immune processes but do not elicit specific immune responses. (hide)
EV-METRIC
56% (93rd 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
Isolation method: density gradient, at least as validation of results attributed to EVs
EV density
Isolation method: reporting of obtained EV density
ultracentrifugation specifics
Isolation 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
Focus vesicles
exosomes
Isolation protocol
Isolation protocol
  • Gives a short, non-chronological overview of the
    different steps of the isolation protocol.
    • dUC = differential ultracentrifugation
    • DG = density gradient
    • UF = ultrafiltration
    • SEC = size-exclusion chromatography
0.2 µm filter + dUC + Sucrose-DG (valid.)
Adj. k-factor
256 (pelleting)
Protein markers
EV: Alix/ Flotillin
non-EV: Cell organelle protein
Proteomics
yes
EV density (g/ml)
1.100
Show all info
Study aim
Function
Sample
Species
Mus musculus
Sample Type
Cell culture supernatant
EV-harvesting Medium
serum free
Isolation Method
Differential ultracentrifugation
dUC: centrifugation steps
Between 800 g and 10,000 g
Between 10,000 g and 50,000 g
Between 100,000 g and 150,000 g
Pelleting: time(min)
150
Pelleting: rotor type
SW32
Pelleting: adjusted k-factor
256.0
Density gradient
Only used for validation of main results
1
Density medium
Sucrose
Lowest density fraction
0.25
Highest density fraction
2.25
Orientation
Top-down
Rotor type
SW32
Speed (g)
100000
Filtration steps
0.22µm or 0.2µm
Characterization: Protein analysis
Western Blot
Detected EV-associated proteins
Alix/ Flotillin
Detected contaminants
Cell organelle protein
ELISA
Detected EV-associated proteins
Flotillin
Fluorescent NTA
Characterization: Particle analysis
EM
EM-type
transmission EM
Image type
Close-up
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