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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 code Experiment nr. Species Sample type Separation protocol First author Year EV-METRIC
EV210254 1/3 Homo sapiens Cell culture supernatant (d)(U)C/ Filtration Imanbekova, Meruyert 2021 50%

Study summary

Full title
All authors
Meruyert Imanbekova, Sorina Suarasan, Tatu Rojalin, Rachel R. Mizenko, Silvia Hilt, Meghna Mathur, Paula Lepine, Michael Nicouleau, Nguyen-Vi Mohamed, Thomas M. Durcan, Randy P. Carney, John C. Voss and Sebastian Wachsmann-Hogiu
Journal
Nanoscale advances
Abstract
One of the hallmarks of Alzheimer's disease (AD) pathogenesis is believed to be the production and d (show more...)One of the hallmarks of Alzheimer's disease (AD) pathogenesis is believed to be the production and deposition of amyloid-beta (Aβ) peptide into extracellular plaques. Existing research indicates that extracellular vesicles (EVs) can carry Aβ associated with AD. However, characterization of the EVs-associated Aβ and its conformational variants has yet to be realized. Raman spectroscopy is a label-free and non-destructive method that is able to assess the biochemical composition of EVs. This study reports for the first time the Raman spectroscopic fingerprint of the Aβ present in the molecular cargo of small extracellular vesicles (sEVs). Raman spectra were measured from sEVs isolated from Alzheimer's disease cell culture model, where secretion of Aβ is regulated by tetracycline promoter, and from midbrain organoids. The averaged spectra of each sEV group showed considerable variation as a reflection of the biochemical content of sEVs. Spectral analysis identified more intense Raman peaks at 1650 cm−1 and 2930 cm−1 attributable to the Aβ peptide incorporated in sEVs produced by the Alzheimer's cell culture model. Subsequent analysis of the spectra by principal component analysis differentiated the sEVs of the Alzheimer's disease cell culture model from the control groups of sEVs. Moreover, the results indicate that Aβ associated with secreted sEVs has a α-helical secondary structure and the size of a monomer or small oligomer. Furthermore, by analyzing the lipid content of sEVs we identified altered fatty acid chain lengths in sEVs that carry Aβ that may affect the fluidity of the EV membrane. Overall, our findings provide evidence supporting the use of Raman spectroscopy for the identification and characterization of sEVs associated with potential biomarkers of neurological disorders such as toxic proteins. (hide)
EV-METRIC
50% (79th 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
Control condition
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
Protein markers
EV: CD9/ CD63/ CD81/ AB
non-EV: None
Proteomics
no
Show all info
Study aim
Biomarker
Sample
Species
Homo sapiens
Sample Type
Cell culture supernatant
EV-producing cells
MC65
EV-harvesting Medium
Serum free medium
Separation Method
(Differential) (ultra)centrifugation
dUC: centrifugation steps
Below or equal to 800 g
Between 800 g and 10,000 g
Between 10,000 g and 50,000 g
Equal to or above 150,000 g
Pelleting: time(min)
90
Pelleting: rotor type
SW 28
Pelleting: speed (g)
120000
Wash: volume per pellet (ml)
Not specified
Wash: time (min)
90
Wash: Rotor Type
SW 28
Wash: speed (g)
120000
Filtration steps
Between 0.1 and 0.2 ?m
Characterization: Protein analysis
Protein Concentration Method
Not determined
Detected EV-associated proteins
AB
Detected EV-associated proteins
CD9/ CD63/ CD81
Characterization: Lipid analysis
Yes
Characterization: Particle analysis
NTA
Report type
Mean
Reported size (nm)
157.3 nm
EV concentration
Yes
EM
EM-type
Transmission-EM
Image type
Close-up
Report size (nm)
150
EV210254 2/3 Homo sapiens Cell culture supernatant (d)(U)C/ Filtration Imanbekova, Meruyert 2021 50%

Study summary

Full title
All authors
Meruyert Imanbekova, Sorina Suarasan, Tatu Rojalin, Rachel R. Mizenko, Silvia Hilt, Meghna Mathur, Paula Lepine, Michael Nicouleau, Nguyen-Vi Mohamed, Thomas M. Durcan, Randy P. Carney, John C. Voss and Sebastian Wachsmann-Hogiu
Journal
Nanoscale advances
Abstract
One of the hallmarks of Alzheimer's disease (AD) pathogenesis is believed to be the production and d (show more...)One of the hallmarks of Alzheimer's disease (AD) pathogenesis is believed to be the production and deposition of amyloid-beta (Aβ) peptide into extracellular plaques. Existing research indicates that extracellular vesicles (EVs) can carry Aβ associated with AD. However, characterization of the EVs-associated Aβ and its conformational variants has yet to be realized. Raman spectroscopy is a label-free and non-destructive method that is able to assess the biochemical composition of EVs. This study reports for the first time the Raman spectroscopic fingerprint of the Aβ present in the molecular cargo of small extracellular vesicles (sEVs). Raman spectra were measured from sEVs isolated from Alzheimer's disease cell culture model, where secretion of Aβ is regulated by tetracycline promoter, and from midbrain organoids. The averaged spectra of each sEV group showed considerable variation as a reflection of the biochemical content of sEVs. Spectral analysis identified more intense Raman peaks at 1650 cm−1 and 2930 cm−1 attributable to the Aβ peptide incorporated in sEVs produced by the Alzheimer's cell culture model. Subsequent analysis of the spectra by principal component analysis differentiated the sEVs of the Alzheimer's disease cell culture model from the control groups of sEVs. Moreover, the results indicate that Aβ associated with secreted sEVs has a α-helical secondary structure and the size of a monomer or small oligomer. Furthermore, by analyzing the lipid content of sEVs we identified altered fatty acid chain lengths in sEVs that carry Aβ that may affect the fluidity of the EV membrane. Overall, our findings provide evidence supporting the use of Raman spectroscopy for the identification and characterization of sEVs associated with potential biomarkers of neurological disorders such as toxic proteins. (hide)
EV-METRIC
50% (79th 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
Tetracycline-treated
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
Protein markers
EV: CD9/ CD63/ CD81/ AB
non-EV: None
Proteomics
no
Show all info
Study aim
Biomarker
Sample
Species
Homo sapiens
Sample Type
Cell culture supernatant
EV-producing cells
MC65
EV-harvesting Medium
Serum free medium
Separation Method
(Differential) (ultra)centrifugation
dUC: centrifugation steps
Below or equal to 800 g
Between 800 g and 10,000 g
Between 10,000 g and 50,000 g
Equal to or above 150,000 g
Pelleting: time(min)
90
Pelleting: rotor type
SW 28
Pelleting: speed (g)
120000
Wash: volume per pellet (ml)
Not specified
Wash: time (min)
90
Wash: Rotor Type
SW 28
Wash: speed (g)
120000
Filtration steps
Between 0.1 and 0.2 ?m
Characterization: Protein analysis
Protein Concentration Method
Not determined
Detected EV-associated proteins
AB
Detected EV-associated proteins
CD9/ CD63/ CD81
Characterization: Lipid analysis
Yes
Characterization: Particle analysis
NTA
Report type
Mean
Reported size (nm)
164.1 nm
EV concentration
Yes
EM
EM-type
Transmission-EM
Image type
Close-up
Report size (nm)
150
EV210254 3/3 Homo sapiens Cell culture supernatant (d)(U)C/ Filtration Imanbekova, Meruyert 2021 50%

Study summary

Full title
All authors
Meruyert Imanbekova, Sorina Suarasan, Tatu Rojalin, Rachel R. Mizenko, Silvia Hilt, Meghna Mathur, Paula Lepine, Michael Nicouleau, Nguyen-Vi Mohamed, Thomas M. Durcan, Randy P. Carney, John C. Voss and Sebastian Wachsmann-Hogiu
Journal
Nanoscale advances
Abstract
One of the hallmarks of Alzheimer's disease (AD) pathogenesis is believed to be the production and d (show more...)One of the hallmarks of Alzheimer's disease (AD) pathogenesis is believed to be the production and deposition of amyloid-beta (Aβ) peptide into extracellular plaques. Existing research indicates that extracellular vesicles (EVs) can carry Aβ associated with AD. However, characterization of the EVs-associated Aβ and its conformational variants has yet to be realized. Raman spectroscopy is a label-free and non-destructive method that is able to assess the biochemical composition of EVs. This study reports for the first time the Raman spectroscopic fingerprint of the Aβ present in the molecular cargo of small extracellular vesicles (sEVs). Raman spectra were measured from sEVs isolated from Alzheimer's disease cell culture model, where secretion of Aβ is regulated by tetracycline promoter, and from midbrain organoids. The averaged spectra of each sEV group showed considerable variation as a reflection of the biochemical content of sEVs. Spectral analysis identified more intense Raman peaks at 1650 cm−1 and 2930 cm−1 attributable to the Aβ peptide incorporated in sEVs produced by the Alzheimer's cell culture model. Subsequent analysis of the spectra by principal component analysis differentiated the sEVs of the Alzheimer's disease cell culture model from the control groups of sEVs. Moreover, the results indicate that Aβ associated with secreted sEVs has a α-helical secondary structure and the size of a monomer or small oligomer. Furthermore, by analyzing the lipid content of sEVs we identified altered fatty acid chain lengths in sEVs that carry Aβ that may affect the fluidity of the EV membrane. Overall, our findings provide evidence supporting the use of Raman spectroscopy for the identification and characterization of sEVs associated with potential biomarkers of neurological disorders such as toxic proteins. (hide)
EV-METRIC
50% (79th 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
Control condition
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
Protein markers
EV: CD9/ CD63/ CD81/ AB
non-EV: None
Proteomics
no
Show all info
Study aim
Biomarker
Sample
Species
Homo sapiens
Sample Type
Cell culture supernatant
EV-producing cells
midbrain organoids
EV-harvesting Medium
Serum free medium
Separation Method
(Differential) (ultra)centrifugation
dUC: centrifugation steps
Below or equal to 800 g
Between 800 g and 10,000 g
Between 10,000 g and 50,000 g
Equal to or above 150,000 g
Pelleting: time(min)
90
Pelleting: rotor type
SW 28
Pelleting: speed (g)
120000
Wash: volume per pellet (ml)
Not specified
Wash: time (min)
90
Wash: Rotor Type
SW 28
Wash: speed (g)
120000
Filtration steps
Between 0.1 and 0.2 ?m
Characterization: Protein analysis
Protein Concentration Method
Not determined
Detected EV-associated proteins
AB
Detected EV-associated proteins
CD9/ CD63/ CD81
Characterization: Lipid analysis
Yes
Characterization: Particle analysis
NTA
Report type
Mean
Reported size (nm)
293.5 nm
EV concentration
Yes
EM
EM-type
Transmission-EM
Image type
Close-up
Report size (nm)
150
1 - 3 of 3
  • CM = Commercial method
  • dUC = differential ultracentrifugation
  • DG = density gradient
  • UF = ultrafiltration
  • SEC = size-exclusion chromatography
EV-TRACK ID
EV210254
species
Homo sapiens
sample type
Cell culture
cell type
MC65
MC65
midbrain organoids
condition
Control condition
Tetracycline-treated
Control condition
separation protocol
dUC/ Filtration
dUC/ Filtration
dUC/ Filtration
Exp. nr.
1
2
3
EV-METRIC %
50
50
50