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You searched for: EV160007 (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
Experiment number
  • Experiments differ in Sample type
Experiment number
  • Experiments differ in Sample type
Experiment number
  • Experiments differ in Sample type
Details EV-TRACK ID Experiment nr. Species Sample type Separation protocol First author Year EV-METRIC
EV160007 1/3 Homo sapiens Blood plasma (d)(U)C
Filtration
SEC
Hong CS 2016 50%

Study summary

Full title
All authors
Hong CS, Funk S, Muller L, Boyiadzis M, Whiteside TL
Journal
J Extracell Vesicles
Abstract
OBJECTIVE: Isolation from human plasma of exosomes that retain functional and morphological integri (show more...)OBJECTIVE: Isolation from human plasma of exosomes that retain functional and morphological integrity for probing their protein, lipid and nucleic acid content is a priority for the future use of exosomes as biomarkers. A method that meets these criteria and can be scaled up for patient monitoring is thus desirable. METHODS: Plasma specimens (1 mL) of patients with acute myeloid leukaemia (AML) or a head and neck squamous cell carcinoma (HNSCC) were differentially centrifuged, ultrafiltered and fractionated by size exclusion chromatography in small disposable columns (mini-SEC). Exosomes were eluted in phosphate-buffered saline and were evaluated by qNano for particle size and counts, morphology by transmission electron microscopy, protein content, molecular profiles by western blots, and for ability to modify functions of immune cells. RESULTS: Exosomes eluting in fractions #3-5 had a diameter ranging from 50 to 200 nm by qNano, with the fraction #4 containing the bulk of clean, unaggregated exosomes. The exosome elution profiles remained constant for repeated runs of the same plasma. Larger plasma volumes could be fractionated running multiple mini-SEC columns in parallel. Particle concentrations per millilitre of plasma in #4 fractions of AML and HNSCC were comparable and were higher (p<0.003) than those in normal controls. Isolated AML exosomes co-incubated with normal human NK cells inhibited NKG2D expression levels (p<0.004), and HNSCC exosomes suppressed activation (p<0.01) and proliferation of activated T lymphocytes (p<0.03). CONCLUSIONS: Mini-SEC allows for simple and reproducible isolation from human plasma of exosomes retaining structural integrity and functional activity. It enables molecular/functional analysis of the exosome content in serial specimens of human plasma for clinical applications. (hide)
EV-METRIC
50% (83rd 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
Blood plasma
Sample origin
Acute myeloid leukemia
Focus vesicles
exosome
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
Filtration
SEC
Protein markers
EV: TSG101/ PD-1/ CD123/ CD96/ CD44/ CD34/ Pro-TGFbeta1 LAP/ Pro-TGFbeta1LAP/ PD-L1/ CLL-1/ CD9
non-EV: None
Proteomics
no
Show all info
Study aim
Function, Biomarker, New methodological development, Identification of content (omics approaches)
Sample
Species
Homo sapiens
Sample Type
Blood plasma
Separation Method
(Differential) (ultra)centrifugation
dUC: centrifugation steps
Between 800 g and 10,000 g
Between 10,000 g and 50,000 g
Pelleting performed
No
Filtration steps
0.22µm or 0.2µm
Size-exclusion chromatography
Total column volume (mL)
10
Sample volume/column (mL)
1
Resin type
Sepharose CL-2B
Characterization: Protein analysis
Protein Concentration Method
BCA
Protein Yield (µg)
66
Western Blot
Antibody details provided?
Yes
Lysis buffer provided?
Yes
Detected EV-associated proteins
CD9, TSG101, CD44, CD34, CD123, CD96, CLL-1, Pro-TGFbeta1 LAP, PD-1, PD-L1
Characterization: Lipid analysis
No
Characterization: Particle analysis
TRPS
Report type
Mean
Reported size (nm)
77-92
EV concentration
Yes
Particle yield
8.90E+10 particles/ml start sample
EM
EM-type
Transmission-EM
Image type
Wide-field
EV160007 2/3 Homo sapiens Blood plasma (d)(U)C
Filtration
SEC
Hong CS 2016 50%

Study summary

Full title
All authors
Hong CS, Funk S, Muller L, Boyiadzis M, Whiteside TL
Journal
J Extracell Vesicles
Abstract
OBJECTIVE: Isolation from human plasma of exosomes that retain functional and morphological integri (show more...)OBJECTIVE: Isolation from human plasma of exosomes that retain functional and morphological integrity for probing their protein, lipid and nucleic acid content is a priority for the future use of exosomes as biomarkers. A method that meets these criteria and can be scaled up for patient monitoring is thus desirable. METHODS: Plasma specimens (1 mL) of patients with acute myeloid leukaemia (AML) or a head and neck squamous cell carcinoma (HNSCC) were differentially centrifuged, ultrafiltered and fractionated by size exclusion chromatography in small disposable columns (mini-SEC). Exosomes were eluted in phosphate-buffered saline and were evaluated by qNano for particle size and counts, morphology by transmission electron microscopy, protein content, molecular profiles by western blots, and for ability to modify functions of immune cells. RESULTS: Exosomes eluting in fractions #3-5 had a diameter ranging from 50 to 200 nm by qNano, with the fraction #4 containing the bulk of clean, unaggregated exosomes. The exosome elution profiles remained constant for repeated runs of the same plasma. Larger plasma volumes could be fractionated running multiple mini-SEC columns in parallel. Particle concentrations per millilitre of plasma in #4 fractions of AML and HNSCC were comparable and were higher (p<0.003) than those in normal controls. Isolated AML exosomes co-incubated with normal human NK cells inhibited NKG2D expression levels (p<0.004), and HNSCC exosomes suppressed activation (p<0.01) and proliferation of activated T lymphocytes (p<0.03). CONCLUSIONS: Mini-SEC allows for simple and reproducible isolation from human plasma of exosomes retaining structural integrity and functional activity. It enables molecular/functional analysis of the exosome content in serial specimens of human plasma for clinical applications. (hide)
EV-METRIC
50% (83rd 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
Blood plasma
Sample origin
Head and neck squamous cell carcinoma
Focus vesicles
exosome
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
Filtration
SEC
Protein markers
EV: TSG101/ CD39/ PD-1/ CD73/ Cox2/ Fas/ FasL/ PD-L1/ HSP70/ CD9
non-EV: None
Proteomics
no
Show all info
Study aim
Function, Biomarker, New methodological development, Identification of content (omics approaches)
Sample
Species
Homo sapiens
Sample Type
Blood plasma
Separation Method
(Differential) (ultra)centrifugation
dUC: centrifugation steps
Between 800 g and 10,000 g
Between 10,000 g and 50,000 g
Pelleting performed
No
Filtration steps
0.22µm or 0.2µm
Size-exclusion chromatography
Total column volume (mL)
10
Sample volume/column (mL)
1
Resin type
Sepharose CL-2B
Characterization: Protein analysis
Protein Concentration Method
BCA
Protein Yield (µg)
123
Western Blot
Antibody details provided?
Yes
Lysis buffer provided?
Yes
Detected EV-associated proteins
CD9, TSG101, HSP70, Cox2, CD73, CD39, Fas, FasL, PD-1, PD-L1
Characterization: Lipid analysis
No
Characterization: Particle analysis
TRPS
Report type
Mean
Reported size (nm)
73-76
EV concentration
Yes
Particle yield
1.10E+11 particles/ml start sample
EM
EM-type
Transmission-EM
Image type
Wide-field
EV160007 3/3 Homo sapiens Blood plasma (d)(U)C
Filtration
SEC
Hong CS 2016 50%

Study summary

Full title
All authors
Hong CS, Funk S, Muller L, Boyiadzis M, Whiteside TL
Journal
J Extracell Vesicles
Abstract
OBJECTIVE: Isolation from human plasma of exosomes that retain functional and morphological integri (show more...)OBJECTIVE: Isolation from human plasma of exosomes that retain functional and morphological integrity for probing their protein, lipid and nucleic acid content is a priority for the future use of exosomes as biomarkers. A method that meets these criteria and can be scaled up for patient monitoring is thus desirable. METHODS: Plasma specimens (1 mL) of patients with acute myeloid leukaemia (AML) or a head and neck squamous cell carcinoma (HNSCC) were differentially centrifuged, ultrafiltered and fractionated by size exclusion chromatography in small disposable columns (mini-SEC). Exosomes were eluted in phosphate-buffered saline and were evaluated by qNano for particle size and counts, morphology by transmission electron microscopy, protein content, molecular profiles by western blots, and for ability to modify functions of immune cells. RESULTS: Exosomes eluting in fractions #3-5 had a diameter ranging from 50 to 200 nm by qNano, with the fraction #4 containing the bulk of clean, unaggregated exosomes. The exosome elution profiles remained constant for repeated runs of the same plasma. Larger plasma volumes could be fractionated running multiple mini-SEC columns in parallel. Particle concentrations per millilitre of plasma in #4 fractions of AML and HNSCC were comparable and were higher (p<0.003) than those in normal controls. Isolated AML exosomes co-incubated with normal human NK cells inhibited NKG2D expression levels (p<0.004), and HNSCC exosomes suppressed activation (p<0.01) and proliferation of activated T lymphocytes (p<0.03). CONCLUSIONS: Mini-SEC allows for simple and reproducible isolation from human plasma of exosomes retaining structural integrity and functional activity. It enables molecular/functional analysis of the exosome content in serial specimens of human plasma for clinical applications. (hide)
EV-METRIC
50% (83rd 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
Blood plasma
Sample origin
Control condition
Focus vesicles
exosome
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
Filtration
SEC
Protein markers
EV: TSG101/ CD39/ CD123/ PD-1/ CD73/ Cox2/ Fas/ Pro-TGFbeta1 LAP/ Pro-TGFbeta1LAP/ FasL/ PD-L1/ CLL-1/ HSP70/ CD9
non-EV: None
Proteomics
no
Show all info
Study aim
Function, Biomarker, New methodological development, Identification of content (omics approaches)
Sample
Species
Homo sapiens
Sample Type
Blood plasma
Separation Method
(Differential) (ultra)centrifugation
dUC: centrifugation steps
Between 800 g and 10,000 g
Between 10,000 g and 50,000 g
Pelleting performed
No
Filtration steps
0.22µm or 0.2µm
Size-exclusion chromatography
Total column volume (mL)
10
Sample volume/column (mL)
1
Resin type
Sepharose CL-2B
Characterization: Protein analysis
Protein Concentration Method
BCA
Protein Yield (µg)
32
Western Blot
Antibody details provided?
Yes
Lysis buffer provided?
Yes
Detected EV-associated proteins
CD9, TSG101, CD123, CLL-1, Pro-TGFbeta1 LAP, PD-1, HSP70, Cox2, CD73, CD39, Fas, FasL, PD-L1
Characterization: Lipid analysis
No
Characterization: Particle analysis
TRPS
Report type
Mean
Reported size (nm)
69-76
EV concentration
Yes
Particle yield
7.00E+09 particles/ml start sample
EM
EM-type
Transmission-EM
Image type
Wide-field
1 - 3 of 3
  • CM = Commercial method
  • dUC = differential ultracentrifugation
  • DG = density gradient
  • UF = ultrafiltration
  • SEC = size-exclusion chromatography
EV-TRACK ID
EV160007
species
Homo sapiens
sample type
Blood plasma
condition
Acute
myeloid leukemia
Head
and neck squamous cell carcinoma
Control condition
separation protocol
(d)(U)C
Filtration
SEC
(d)(U)C
Filtration
SEC
(d)(U)C
Filtration
SEC
Exp. nr.
1
2
3
EV-METRIC %
50
50
50