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You searched for: EV210020 (EV-TRACK ID)

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Results list Study Tree
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
EV210020 1/9 Homo sapiens MM6 (d)(U)C
Filtration
DG 		Veerman, Rosanne 2021 71%

Study summary

Full title
Molecular evaluation of five different isolation methods for extracellular vesicles reveals different clinical applicability and subcellular origin
All authors
Rosanne E. Veerman, Loes Teeuwen, Paulo Czarnewski, Gözde Güclüler Akpinar, AnnSofi Sandberg, Xiaofang Cao, Maria Pernemalm, Lukas M. Orre, Susanne Gabrielsson, Maria Eldh
Journal
J Extracell Vesicles
Abstract
Extracellular vesicles (EVs) are increasingly tested as therapeutic vehicles and biomarkers, but sti (show more...)Extracellular vesicles (EVs) are increasingly tested as therapeutic vehicles and biomarkers, but still EV subtypes are not fully characterised. To isolate EVs with few co-isolated entities, a combination of methods is needed. However, this is time-consuming and requires large sample volumes, often not feasible in most clinical studies or in studies where small sample volumes are available. Therefore, we compared EVs rendered by five commonly used methods based on different principles from conditioned cell medium and 250 μl or 3 ml plasma, that is, precipitation (ExoQuick ULTRA), membrane affinity (exoEasy Maxi Kit), size-exclusion chromatography (qEVoriginal), iodixanol gradient (OptiPrep), and phosphatidylserine affinity (MagCapture). EVs were characterised by electron microscopy, Nanoparticle Tracking Analysis, Bioanalyzer, flow cytometry, and LC-MS/MS. The different methods yielded samples of different morphology, particle size, and proteomic profile. For the conditioned medium, Izon 35 isolated the highest number of EV proteins followed by exoEasy, which also isolated fewer non-EV proteins. For the plasma samples, exoEasy isolated a high number of EV proteins and few non-EV proteins, while Izon 70 isolated the most EV proteins. We conclude that no method is perfect for all studies, rather, different methods are suited depending on sample type and interest in EV subtype, in addition to sample volume and budget. (hide)
EV-METRIC
71% (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
Blood plasma
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
(Differential) (ultra)centrifugation
Filtration
Density gradient
Protein markers
EV: CD81/ CD63/ CD9
non-EV: None
Proteomics
yes
EV density (g/ml)
1.12-1.19
Show all info
Study aim
Technical analysis comparing/optimizing EV-related methods
Sample
Species
Homo sapiens
Sample Type
Cell culture supernatant
EV-producing cells
MM6
EV-harvesting Medium
EV-depleted medium
Preparation of EDS
overnight (16h) at >=100,000g
Cell viability (%)
90
Separation Method
(Differential) (ultra)centrifugation
dUC: centrifugation steps
Below or equal to 800 g
Between 800 g and 10,000 g
Pelleting performed
No
Density gradient
Type
Discontinuous
Number of initial discontinuous layers
3
Lowest density fraction
10%
Highest density fraction
50%
Total gradient volume, incl. sample (mL)
10.5
Sample volume (mL)
0.5
Orientation
Top-down
Rotor type
SW 32.1 Ti
Speed (g)
130000
Duration (min)
960
Fraction volume (mL)
0.5
Fraction processing
Centrifugation
Pelleting: volume per fraction
65
Pelleting: duration (min)
120
Pelleting: rotor type
Type 45 Ti
Pelleting: speed (g)
100000
Filtration steps
> 0.45 µm,
Characterization: Protein analysis
Protein Concentration Method
Bradford
Flow cytometry specific beads
Antibody details provided?
No
Antibody dilution provided?
No
Detected EV-associated proteins
CD9/ CD63/ CD81
Proteomics database
No
Characterization: RNA analysis
RNA analysis
Type
Capillary electrophoresis (e.g. Bioanalyzer)
Database
No
Proteinase treatment
No
RNAse treatment
No
Characterization: Lipid analysis
No
Characterization: Particle analysis
NTA
Report type
Modus
Reported size (nm)
126
Particle analysis: flow cytometry
Flow cytometer type
BD FACS Canto II
Hardware adjustment
Calibration bead size
4
EM
EM-type
Transmission-EM
Image type
Close-up, Wide-field
EV210020 5/9 Homo sapiens MM6 (d)(U)C
Filtration
DG 		Veerman, Rosanne 2021 71%

Study summary

Full title
Molecular evaluation of five different isolation methods for extracellular vesicles reveals different clinical applicability and subcellular origin
All authors
Rosanne E. Veerman, Loes Teeuwen, Paulo Czarnewski, Gözde Güclüler Akpinar, AnnSofi Sandberg, Xiaofang Cao, Maria Pernemalm, Lukas M. Orre, Susanne Gabrielsson, Maria Eldh
Journal
J Extracell Vesicles
Abstract
Extracellular vesicles (EVs) are increasingly tested as therapeutic vehicles and biomarkers, but sti (show more...)Extracellular vesicles (EVs) are increasingly tested as therapeutic vehicles and biomarkers, but still EV subtypes are not fully characterised. To isolate EVs with few co-isolated entities, a combination of methods is needed. However, this is time-consuming and requires large sample volumes, often not feasible in most clinical studies or in studies where small sample volumes are available. Therefore, we compared EVs rendered by five commonly used methods based on different principles from conditioned cell medium and 250 μl or 3 ml plasma, that is, precipitation (ExoQuick ULTRA), membrane affinity (exoEasy Maxi Kit), size-exclusion chromatography (qEVoriginal), iodixanol gradient (OptiPrep), and phosphatidylserine affinity (MagCapture). EVs were characterised by electron microscopy, Nanoparticle Tracking Analysis, Bioanalyzer, flow cytometry, and LC-MS/MS. The different methods yielded samples of different morphology, particle size, and proteomic profile. For the conditioned medium, Izon 35 isolated the highest number of EV proteins followed by exoEasy, which also isolated fewer non-EV proteins. For the plasma samples, exoEasy isolated a high number of EV proteins and few non-EV proteins, while Izon 70 isolated the most EV proteins. We conclude that no method is perfect for all studies, rather, different methods are suited depending on sample type and interest in EV subtype, in addition to sample volume and budget. (hide)
EV-METRIC
71% (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
Cell culture supernatant
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
(Differential) (ultra)centrifugation
Filtration
Density gradient
Protein markers
EV: CD81/ CD63/ CD9
non-EV: None
Proteomics
yes
EV density (g/ml)
1.12-1.19
Show all info
Study aim
Technical analysis comparing/optimizing EV-related methods
Sample
Species
Homo sapiens
Sample Type
Cell culture supernatant
EV-producing cells
MM6
EV-harvesting Medium
EV-depleted medium
Preparation of EDS
overnight (16h) at >=100,000g
Cell viability (%)
90
Separation Method
(Differential) (ultra)centrifugation
dUC: centrifugation steps
Below or equal to 800 g
Between 800 g and 10,000 g
Pelleting performed
No
Density gradient
Type
Discontinuous
Number of initial discontinuous layers
3
Lowest density fraction
10%
Highest density fraction
50%
Total gradient volume, incl. sample (mL)
10.5
Sample volume (mL)
0.5
Orientation
Top-down
Rotor type
SW 32.1 Ti
Speed (g)
120000
Duration (min)
1440
Fraction volume (mL)
0.5
Fraction processing
Centrifugation
Pelleting: volume per fraction
65
Pelleting: duration (min)
120
Pelleting: rotor type
Type 45 Ti
Pelleting: speed (g)
100000
Filtration steps
> 0.45 µm,
Characterization: Protein analysis
Protein Concentration Method
Bradford
Flow cytometry specific beads
Antibody details provided?
No
Antibody dilution provided?
No
Detected EV-associated proteins
CD9/ CD63/ CD81
Proteomics database
No
Characterization: RNA analysis
RNA analysis
Type
Capillary electrophoresis (e.g. Bioanalyzer)
Database
No
Proteinase treatment
No
RNAse treatment
No
Characterization: Lipid analysis
No
Characterization: Particle analysis
NTA
Report type
Modus
Reported size (nm)
92
Particle analysis: flow cytometry
Flow cytometer type
BD FACS Canto II
Hardware adjustment
Calibration bead size
4
EM
EM-type
Transmission-EM
Image type
Close-up, Wide-field
EV210020 2/9 Homo sapiens MM6 (d)(U)C
Filtration
ExoQuick 		Veerman, Rosanne 2021 43%

Study summary

Full title
Molecular evaluation of five different isolation methods for extracellular vesicles reveals different clinical applicability and subcellular origin
All authors
Rosanne E. Veerman, Loes Teeuwen, Paulo Czarnewski, Gözde Güclüler Akpinar, AnnSofi Sandberg, Xiaofang Cao, Maria Pernemalm, Lukas M. Orre, Susanne Gabrielsson, Maria Eldh
Journal
J Extracell Vesicles
Abstract
Extracellular vesicles (EVs) are increasingly tested as therapeutic vehicles and biomarkers, but sti (show more...)Extracellular vesicles (EVs) are increasingly tested as therapeutic vehicles and biomarkers, but still EV subtypes are not fully characterised. To isolate EVs with few co-isolated entities, a combination of methods is needed. However, this is time-consuming and requires large sample volumes, often not feasible in most clinical studies or in studies where small sample volumes are available. Therefore, we compared EVs rendered by five commonly used methods based on different principles from conditioned cell medium and 250 μl or 3 ml plasma, that is, precipitation (ExoQuick ULTRA), membrane affinity (exoEasy Maxi Kit), size-exclusion chromatography (qEVoriginal), iodixanol gradient (OptiPrep), and phosphatidylserine affinity (MagCapture). EVs were characterised by electron microscopy, Nanoparticle Tracking Analysis, Bioanalyzer, flow cytometry, and LC-MS/MS. The different methods yielded samples of different morphology, particle size, and proteomic profile. For the conditioned medium, Izon 35 isolated the highest number of EV proteins followed by exoEasy, which also isolated fewer non-EV proteins. For the plasma samples, exoEasy isolated a high number of EV proteins and few non-EV proteins, while Izon 70 isolated the most EV proteins. We conclude that no method is perfect for all studies, rather, different methods are suited depending on sample type and interest in EV subtype, in addition to sample volume and budget. (hide)
EV-METRIC
43% (81st 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
Blood plasma
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
(Differential) (ultra)centrifugation
Filtration
Commercial method
Protein markers
EV: CD81/ CD63/ CD9
non-EV: None
Proteomics
yes
Show all info
Study aim
Technical analysis comparing/optimizing EV-related methods
Sample
Species
Homo sapiens
Sample Type
Cell culture supernatant
EV-producing cells
MM6
EV-harvesting Medium
EV-depleted medium
Preparation of EDS
overnight (16h) at >=100,000g
Cell viability (%)
90
Separation Method
(Differential) (ultra)centrifugation
dUC: centrifugation steps
Below or equal to 800 g
Between 800 g and 10,000 g
Pelleting performed
No
Filtration steps
> 0.45 µm,
Commercial kit
ExoQuick
Characterization: Protein analysis
Protein Concentration Method
Bradford
Flow cytometry specific beads
Antibody details provided?
No
Antibody dilution provided?
No
Detected EV-associated proteins
CD9/ CD63/ CD81
Proteomics database
No
Characterization: RNA analysis
RNA analysis
Type
Capillary electrophoresis (e.g. Bioanalyzer)
Database
No
Proteinase treatment
No
RNAse treatment
No
Characterization: Lipid analysis
No
Characterization: Particle analysis
NTA
Report type
Modus
Reported size (nm)
126
Particle analysis: flow cytometry
Flow cytometer type
BD FACS Canto II
Hardware adjustment
Calibration bead size
4
EM
EM-type
Transmission-EM
Image type
Close-up, Wide-field
EV210020 3/9 Homo sapiens MM6 (d)(U)C
Filtration
Other;exoEasy 		Veerman, Rosanne 2021 43%

Study summary

Full title
Molecular evaluation of five different isolation methods for extracellular vesicles reveals different clinical applicability and subcellular origin
All authors
Rosanne E. Veerman, Loes Teeuwen, Paulo Czarnewski, Gözde Güclüler Akpinar, AnnSofi Sandberg, Xiaofang Cao, Maria Pernemalm, Lukas M. Orre, Susanne Gabrielsson, Maria Eldh
Journal
J Extracell Vesicles
Abstract
Extracellular vesicles (EVs) are increasingly tested as therapeutic vehicles and biomarkers, but sti (show more...)Extracellular vesicles (EVs) are increasingly tested as therapeutic vehicles and biomarkers, but still EV subtypes are not fully characterised. To isolate EVs with few co-isolated entities, a combination of methods is needed. However, this is time-consuming and requires large sample volumes, often not feasible in most clinical studies or in studies where small sample volumes are available. Therefore, we compared EVs rendered by five commonly used methods based on different principles from conditioned cell medium and 250 μl or 3 ml plasma, that is, precipitation (ExoQuick ULTRA), membrane affinity (exoEasy Maxi Kit), size-exclusion chromatography (qEVoriginal), iodixanol gradient (OptiPrep), and phosphatidylserine affinity (MagCapture). EVs were characterised by electron microscopy, Nanoparticle Tracking Analysis, Bioanalyzer, flow cytometry, and LC-MS/MS. The different methods yielded samples of different morphology, particle size, and proteomic profile. For the conditioned medium, Izon 35 isolated the highest number of EV proteins followed by exoEasy, which also isolated fewer non-EV proteins. For the plasma samples, exoEasy isolated a high number of EV proteins and few non-EV proteins, while Izon 70 isolated the most EV proteins. We conclude that no method is perfect for all studies, rather, different methods are suited depending on sample type and interest in EV subtype, in addition to sample volume and budget. (hide)
EV-METRIC
43% (81st 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
Blood plasma
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
(Differential) (ultra)centrifugation
Filtration
Commercial method
Protein markers
EV: CD81/ CD63/ CD9
non-EV: None
Proteomics
yes
Show all info
Study aim
Technical analysis comparing/optimizing EV-related methods
Sample
Species
Homo sapiens
Sample Type
Cell culture supernatant
EV-producing cells
MM6
EV-harvesting Medium
EV-depleted medium
Preparation of EDS
overnight (16h) at >=100,000g
Cell viability (%)
90
Separation Method
(Differential) (ultra)centrifugation
dUC: centrifugation steps
Below or equal to 800 g
Between 800 g and 10,000 g
Pelleting performed
No
Filtration steps
> 0.45 µm,
Commercial kit
Other;exoEasy
Characterization: Protein analysis
Protein Concentration Method
Bradford
Flow cytometry specific beads
Antibody details provided?
No
Antibody dilution provided?
No
Detected EV-associated proteins
CD9/ CD63/ CD81
Proteomics database
No
Characterization: RNA analysis
RNA analysis
Type
Capillary electrophoresis (e.g. Bioanalyzer)
Database
No
Proteinase treatment
No
RNAse treatment
No
Characterization: Lipid analysis
No
Characterization: Particle analysis
NTA
Report type
Modus
Reported size (nm)
126
Particle analysis: flow cytometry
Flow cytometer type
BD FACS Canto II
Hardware adjustment
Calibration bead size
4
EM
EM-type
Transmission-EM
Image type
Close-up, Wide-field
EV210020 4/9 Homo sapiens MM6 (d)(U)C
Filtration
qEV 		Veerman, Rosanne 2021 43%

Study summary

Full title
Molecular evaluation of five different isolation methods for extracellular vesicles reveals different clinical applicability and subcellular origin
All authors
Rosanne E. Veerman, Loes Teeuwen, Paulo Czarnewski, Gözde Güclüler Akpinar, AnnSofi Sandberg, Xiaofang Cao, Maria Pernemalm, Lukas M. Orre, Susanne Gabrielsson, Maria Eldh
Journal
J Extracell Vesicles
Abstract
Extracellular vesicles (EVs) are increasingly tested as therapeutic vehicles and biomarkers, but sti (show more...)Extracellular vesicles (EVs) are increasingly tested as therapeutic vehicles and biomarkers, but still EV subtypes are not fully characterised. To isolate EVs with few co-isolated entities, a combination of methods is needed. However, this is time-consuming and requires large sample volumes, often not feasible in most clinical studies or in studies where small sample volumes are available. Therefore, we compared EVs rendered by five commonly used methods based on different principles from conditioned cell medium and 250 μl or 3 ml plasma, that is, precipitation (ExoQuick ULTRA), membrane affinity (exoEasy Maxi Kit), size-exclusion chromatography (qEVoriginal), iodixanol gradient (OptiPrep), and phosphatidylserine affinity (MagCapture). EVs were characterised by electron microscopy, Nanoparticle Tracking Analysis, Bioanalyzer, flow cytometry, and LC-MS/MS. The different methods yielded samples of different morphology, particle size, and proteomic profile. For the conditioned medium, Izon 35 isolated the highest number of EV proteins followed by exoEasy, which also isolated fewer non-EV proteins. For the plasma samples, exoEasy isolated a high number of EV proteins and few non-EV proteins, while Izon 70 isolated the most EV proteins. We conclude that no method is perfect for all studies, rather, different methods are suited depending on sample type and interest in EV subtype, in addition to sample volume and budget. (hide)
EV-METRIC
43% (81st 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
Blood plasma
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
(Differential) (ultra)centrifugation
Filtration
Commercial method
Protein markers
EV: CD81/ CD63/ CD9
non-EV: None
Proteomics
yes
Show all info
Study aim
Technical analysis comparing/optimizing EV-related methods
Sample
Species
Homo sapiens
Sample Type
Cell culture supernatant
EV-producing cells
MM6
EV-harvesting Medium
EV-depleted medium
Preparation of EDS
overnight (16h) at >=100,000g
Cell viability (%)
90
Separation Method
(Differential) (ultra)centrifugation
dUC: centrifugation steps
Below or equal to 800 g
Between 800 g and 10,000 g
Pelleting performed
No
Filtration steps
> 0.45 µm,
Commercial kit
qEV
Characterization: Protein analysis
Protein Concentration Method
Bradford
Flow cytometry specific beads
Antibody details provided?
No
Antibody dilution provided?
No
Detected EV-associated proteins
CD9/ CD63/ CD81
Proteomics database
No
Characterization: RNA analysis
RNA analysis
Type
Capillary electrophoresis (e.g. Bioanalyzer)
Database
No
Proteinase treatment
No
RNAse treatment
No
Characterization: Lipid analysis
No
Characterization: Particle analysis
NTA
Report type
Modus
Reported size (nm)
126
Particle analysis: flow cytometry
Flow cytometer type
BD FACS Canto II
Hardware adjustment
Calibration bead size
4
EM
EM-type
Transmission-EM
Image type
Close-up, Wide-field
EV210020 6/9 Homo sapiens MM6 (d)(U)C
Filtration
ExoQuick 		Veerman, Rosanne 2021 43%

Study summary

Full title
Molecular evaluation of five different isolation methods for extracellular vesicles reveals different clinical applicability and subcellular origin
All authors
Rosanne E. Veerman, Loes Teeuwen, Paulo Czarnewski, Gözde Güclüler Akpinar, AnnSofi Sandberg, Xiaofang Cao, Maria Pernemalm, Lukas M. Orre, Susanne Gabrielsson, Maria Eldh
Journal
J Extracell Vesicles
Abstract
Extracellular vesicles (EVs) are increasingly tested as therapeutic vehicles and biomarkers, but sti (show more...)Extracellular vesicles (EVs) are increasingly tested as therapeutic vehicles and biomarkers, but still EV subtypes are not fully characterised. To isolate EVs with few co-isolated entities, a combination of methods is needed. However, this is time-consuming and requires large sample volumes, often not feasible in most clinical studies or in studies where small sample volumes are available. Therefore, we compared EVs rendered by five commonly used methods based on different principles from conditioned cell medium and 250 μl or 3 ml plasma, that is, precipitation (ExoQuick ULTRA), membrane affinity (exoEasy Maxi Kit), size-exclusion chromatography (qEVoriginal), iodixanol gradient (OptiPrep), and phosphatidylserine affinity (MagCapture). EVs were characterised by electron microscopy, Nanoparticle Tracking Analysis, Bioanalyzer, flow cytometry, and LC-MS/MS. The different methods yielded samples of different morphology, particle size, and proteomic profile. For the conditioned medium, Izon 35 isolated the highest number of EV proteins followed by exoEasy, which also isolated fewer non-EV proteins. For the plasma samples, exoEasy isolated a high number of EV proteins and few non-EV proteins, while Izon 70 isolated the most EV proteins. We conclude that no method is perfect for all studies, rather, different methods are suited depending on sample type and interest in EV subtype, in addition to sample volume and budget. (hide)
EV-METRIC
43% (81st 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
Cell culture supernatant
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
(Differential) (ultra)centrifugation
Filtration
Commercial method
Protein markers
EV: CD81/ CD63/ CD9
non-EV: None
Proteomics
yes
Show all info
Study aim
Technical analysis comparing/optimizing EV-related methods
Sample
Species
Homo sapiens
Sample Type
Cell culture supernatant
EV-producing cells
MM6
EV-harvesting Medium
EV-depleted medium
Preparation of EDS
overnight (16h) at >=100,000g
Cell viability (%)
90
Separation Method
(Differential) (ultra)centrifugation
dUC: centrifugation steps
Below or equal to 800 g
Between 800 g and 10,000 g
Pelleting performed
No
Filtration steps
> 0.45 µm,
Commercial kit
ExoQuick
Characterization: Protein analysis
Protein Concentration Method
Bradford
Flow cytometry specific beads
Antibody details provided?
No
Antibody dilution provided?
No
Detected EV-associated proteins
CD9/ CD63/ CD81
Proteomics database
No
Characterization: RNA analysis
RNA analysis
Type
Capillary electrophoresis (e.g. Bioanalyzer)
Database
No
Proteinase treatment
No
RNAse treatment
No
Characterization: Lipid analysis
No
Characterization: Particle analysis
NTA
Report type
Modus
Reported size (nm)
92
Particle analysis: flow cytometry
Flow cytometer type
BD FACS Canto II
Hardware adjustment
Calibration bead size
4
EM
EM-type
Transmission-EM
Image type
Close-up, Wide-field
EV210020 7/9 Homo sapiens MM6 (d)(U)C
Filtration
exoEasy;Other 		Veerman, Rosanne 2021 43%

Study summary

Full title
Molecular evaluation of five different isolation methods for extracellular vesicles reveals different clinical applicability and subcellular origin
All authors
Rosanne E. Veerman, Loes Teeuwen, Paulo Czarnewski, Gözde Güclüler Akpinar, AnnSofi Sandberg, Xiaofang Cao, Maria Pernemalm, Lukas M. Orre, Susanne Gabrielsson, Maria Eldh
Journal
J Extracell Vesicles
Abstract
Extracellular vesicles (EVs) are increasingly tested as therapeutic vehicles and biomarkers, but sti (show more...)Extracellular vesicles (EVs) are increasingly tested as therapeutic vehicles and biomarkers, but still EV subtypes are not fully characterised. To isolate EVs with few co-isolated entities, a combination of methods is needed. However, this is time-consuming and requires large sample volumes, often not feasible in most clinical studies or in studies where small sample volumes are available. Therefore, we compared EVs rendered by five commonly used methods based on different principles from conditioned cell medium and 250 μl or 3 ml plasma, that is, precipitation (ExoQuick ULTRA), membrane affinity (exoEasy Maxi Kit), size-exclusion chromatography (qEVoriginal), iodixanol gradient (OptiPrep), and phosphatidylserine affinity (MagCapture). EVs were characterised by electron microscopy, Nanoparticle Tracking Analysis, Bioanalyzer, flow cytometry, and LC-MS/MS. The different methods yielded samples of different morphology, particle size, and proteomic profile. For the conditioned medium, Izon 35 isolated the highest number of EV proteins followed by exoEasy, which also isolated fewer non-EV proteins. For the plasma samples, exoEasy isolated a high number of EV proteins and few non-EV proteins, while Izon 70 isolated the most EV proteins. We conclude that no method is perfect for all studies, rather, different methods are suited depending on sample type and interest in EV subtype, in addition to sample volume and budget. (hide)
EV-METRIC
43% (81st 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
Cell culture supernatant
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
(Differential) (ultra)centrifugation
Filtration
Commercial method
Protein markers
EV: CD81/ CD63/ CD9
non-EV: None
Proteomics
yes
Show all info
Study aim
Technical analysis comparing/optimizing EV-related methods
Sample
Species
Homo sapiens
Sample Type
Cell culture supernatant
EV-producing cells
MM6
EV-harvesting Medium
EV-depleted medium
Preparation of EDS
overnight (16h) at >=100,000g
Cell viability (%)
90
Separation Method
(Differential) (ultra)centrifugation
dUC: centrifugation steps
Below or equal to 800 g
Between 800 g and 10,000 g
Pelleting performed
No
Filtration steps
> 0.45 µm,
Commercial kit
exoEasy;Other
Characterization: Protein analysis
Protein Concentration Method
Bradford
Flow cytometry specific beads
Antibody details provided?
No
Antibody dilution provided?
No
Detected EV-associated proteins
CD9/ CD63/ CD81
Proteomics database
No
Characterization: RNA analysis
RNA analysis
Type
Capillary electrophoresis (e.g. Bioanalyzer)
Database
No
Proteinase treatment
No
RNAse treatment
No
Characterization: Lipid analysis
No
Characterization: Particle analysis
NTA
Report type
Modus
Reported size (nm)
92
Particle analysis: flow cytometry
Flow cytometer type
BD FACS Canto II
Hardware adjustment
Calibration bead size
4
EM
EM-type
Transmission-EM
Image type
Close-up, Wide-field
EV210020 8/9 Homo sapiens MM6 (d)(U)C
Filtration
qEV 		Veerman, Rosanne 2021 43%

Study summary

Full title
Molecular evaluation of five different isolation methods for extracellular vesicles reveals different clinical applicability and subcellular origin
All authors
Rosanne E. Veerman, Loes Teeuwen, Paulo Czarnewski, Gözde Güclüler Akpinar, AnnSofi Sandberg, Xiaofang Cao, Maria Pernemalm, Lukas M. Orre, Susanne Gabrielsson, Maria Eldh
Journal
J Extracell Vesicles
Abstract
Extracellular vesicles (EVs) are increasingly tested as therapeutic vehicles and biomarkers, but sti (show more...)Extracellular vesicles (EVs) are increasingly tested as therapeutic vehicles and biomarkers, but still EV subtypes are not fully characterised. To isolate EVs with few co-isolated entities, a combination of methods is needed. However, this is time-consuming and requires large sample volumes, often not feasible in most clinical studies or in studies where small sample volumes are available. Therefore, we compared EVs rendered by five commonly used methods based on different principles from conditioned cell medium and 250 μl or 3 ml plasma, that is, precipitation (ExoQuick ULTRA), membrane affinity (exoEasy Maxi Kit), size-exclusion chromatography (qEVoriginal), iodixanol gradient (OptiPrep), and phosphatidylserine affinity (MagCapture). EVs were characterised by electron microscopy, Nanoparticle Tracking Analysis, Bioanalyzer, flow cytometry, and LC-MS/MS. The different methods yielded samples of different morphology, particle size, and proteomic profile. For the conditioned medium, Izon 35 isolated the highest number of EV proteins followed by exoEasy, which also isolated fewer non-EV proteins. For the plasma samples, exoEasy isolated a high number of EV proteins and few non-EV proteins, while Izon 70 isolated the most EV proteins. We conclude that no method is perfect for all studies, rather, different methods are suited depending on sample type and interest in EV subtype, in addition to sample volume and budget. (hide)
EV-METRIC
43% (81st 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
Cell culture supernatant
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
(Differential) (ultra)centrifugation
Filtration
Commercial method
Protein markers
EV: CD81/ CD63/ CD9
non-EV: None
Proteomics
yes
Show all info
Study aim
Technical analysis comparing/optimizing EV-related methods
Sample
Species
Homo sapiens
Sample Type
Cell culture supernatant
EV-producing cells
MM6
EV-harvesting Medium
EV-depleted medium
Preparation of EDS
overnight (16h) at >=100,000g
Cell viability (%)
90
Separation Method
(Differential) (ultra)centrifugation
dUC: centrifugation steps
Below or equal to 800 g
Between 800 g and 10,000 g
Pelleting performed
No
Filtration steps
> 0.45 µm,
Commercial kit
qEV
Characterization: Protein analysis
Protein Concentration Method
Bradford
Flow cytometry specific beads
Antibody details provided?
No
Antibody dilution provided?
No
Detected EV-associated proteins
CD9/ CD63/ CD81
Proteomics database
No
Characterization: RNA analysis
RNA analysis
Type
Capillary electrophoresis (e.g. Bioanalyzer)
Database
No
Proteinase treatment
No
RNAse treatment
No
Characterization: Lipid analysis
No
Characterization: Particle analysis
NTA
Report type
Modus
Reported size (nm)
92
Particle analysis: flow cytometry
Flow cytometer type
BD FACS Canto II
Hardware adjustment
Calibration bead size
4
EM
EM-type
Transmission-EM
Image type
Close-up, Wide-field
EV210020 9/9 Homo sapiens MM6 (d)(U)C
Filtration
MagCapture;Other 		Veerman, Rosanne 2021 43%

Study summary

Full title
Molecular evaluation of five different isolation methods for extracellular vesicles reveals different clinical applicability and subcellular origin
All authors
Rosanne E. Veerman, Loes Teeuwen, Paulo Czarnewski, Gözde Güclüler Akpinar, AnnSofi Sandberg, Xiaofang Cao, Maria Pernemalm, Lukas M. Orre, Susanne Gabrielsson, Maria Eldh
Journal
J Extracell Vesicles
Abstract
Extracellular vesicles (EVs) are increasingly tested as therapeutic vehicles and biomarkers, but sti (show more...)Extracellular vesicles (EVs) are increasingly tested as therapeutic vehicles and biomarkers, but still EV subtypes are not fully characterised. To isolate EVs with few co-isolated entities, a combination of methods is needed. However, this is time-consuming and requires large sample volumes, often not feasible in most clinical studies or in studies where small sample volumes are available. Therefore, we compared EVs rendered by five commonly used methods based on different principles from conditioned cell medium and 250 μl or 3 ml plasma, that is, precipitation (ExoQuick ULTRA), membrane affinity (exoEasy Maxi Kit), size-exclusion chromatography (qEVoriginal), iodixanol gradient (OptiPrep), and phosphatidylserine affinity (MagCapture). EVs were characterised by electron microscopy, Nanoparticle Tracking Analysis, Bioanalyzer, flow cytometry, and LC-MS/MS. The different methods yielded samples of different morphology, particle size, and proteomic profile. For the conditioned medium, Izon 35 isolated the highest number of EV proteins followed by exoEasy, which also isolated fewer non-EV proteins. For the plasma samples, exoEasy isolated a high number of EV proteins and few non-EV proteins, while Izon 70 isolated the most EV proteins. We conclude that no method is perfect for all studies, rather, different methods are suited depending on sample type and interest in EV subtype, in addition to sample volume and budget. (hide)
EV-METRIC
43% (81st 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
Cell culture supernatant
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
(Differential) (ultra)centrifugation
Filtration
Commercial method
Protein markers
EV: CD81/ CD63/ CD9
non-EV: None
Proteomics
yes
Show all info
Study aim
Technical analysis comparing/optimizing EV-related methods
Sample
Species
Homo sapiens
Sample Type
Cell culture supernatant
EV-producing cells
MM6
EV-harvesting Medium
EV-depleted medium
Preparation of EDS
overnight (16h) at >=100,000g
Cell viability (%)
90
Separation Method
(Differential) (ultra)centrifugation
dUC: centrifugation steps
Below or equal to 800 g
Between 800 g and 10,000 g
Pelleting performed
No
Filtration steps
> 0.45 µm,
Commercial kit
MagCapture;Other
Characterization: Protein analysis
Protein Concentration Method
Bradford
Flow cytometry specific beads
Antibody details provided?
No
Antibody dilution provided?
No
Detected EV-associated proteins
CD9/ CD63/ CD81
Proteomics database
No
Characterization: RNA analysis
RNA analysis
Type
Capillary electrophoresis (e.g. Bioanalyzer)
Database
No
Proteinase treatment
No
RNAse treatment
No
Characterization: Lipid analysis
No
Characterization: Particle analysis
NTA
Report type
Modus
Reported size (nm)
92
Particle analysis: flow cytometry
Flow cytometer type
BD FACS Canto II
Hardware adjustment
Calibration bead size
4
EM
EM-type
Transmission-EM
Image type
Close-up, Wide-field
1 - 9 of 9
  • CM = Commercial method
  • dUC = differential ultracentrifugation
  • DG = density gradient
  • UF = ultrafiltration
  • SEC = size-exclusion chromatography
EV-TRACK ID
EV210020
species
Homo
sapiens
sample type
Cell
culture
cell type
MM6
condition
Blood
plasma
Cell
culture
Blood
plasma
Blood
plasma
Blood
plasma
Cell
culture
Cell
culture
Cell
culture
Cell
culture
separation protocol
dUC
Filtration
Density
gradient
dUC
Filtration
Density
gradient
dUC
Filtration
ExoQuick
dUC
Filtration
Other
exoEasy
dUC
Filtration
qEV
dUC
Filtration
ExoQuick
dUC
Filtration
exoEasy
Other
dUC
Filtration
qEV
dUC
Filtration
MagCapture
Other
Exp. nr.
1
5
2
3
4
6
7
8
9
EV-METRIC %
71
71
43
43
43
43
43
43
43