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You searched for: EV200174 (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
Details EV-TRACK code Experiment nr. Species Sample type Separation protocol First author Year EV-METRIC
EV200174 1/2 Bos taurus 1% skim milk (d)(U)C
Sukreet, Sonal 2022 67%

Study summary

Full title
All authors
Sonal Sukreet, Camila Pereira Braga, Thuy T An, Jiri Adamec, Juan Cui, Janos Zempleni
Journal
J Nutr
Abstract
Background: Bovine milk exosomes (BMEs) harbor regulatory proteins, lipids and microRNAs. Consumptio (show more...)Background: Bovine milk exosomes (BMEs) harbor regulatory proteins, lipids and microRNAs. Consumption of an exosome and RNA-depleted (ERD) diet elicited phenotypes compared to controls fed an exosome and RNA-sufficient (ERS) diet in mice. All other ingredients were identical in the diets. ERD and ERS diets were prepared by substituting ultrasonicated and non-ultrasonicated milk, respectively, for casein in the AIN-93G formulation. Objective: The objective of this study was to assess the effect of ultrasonication of milk on exosome content and bioavailability, and cargo content. Methods: Bovine milk was ultrasonicated and exosomes were isolated by ultracentrifugation (USE); controls were not ultrasonicated (NSE). Exosome count, size and morphology were assessed using a nanoparticle tracker and electron microscopy. RNAs, lipids and proteins were analyzed by RNA-sequencing and mass spectrometry. Intestinal transport, bioavailability and distribution were measured by using fluorophore-labeled USEs and NSEs in Caco-2 cells, FHs 74 Int cells and C57BL/6J mice (n = 3; age 6 - 8 weeks). Results: The exosome count was 76 ± 22% lower in USE compared to NSE (P < 0.05). Ultrasonication caused a degradation of up to 100% of microRNAs. USE and NSE contained 145 and 332 unique lipid signatures, respectively (P < 0.05). We detected total of 525 and 484 proteins in USE and NSE. The uptake of USEs decreased by 46 ± 30% and 40 ± 27% compared to NSEs in Caco-2 and FHs 74 Int cells, respectively (P < 0.05). The hepatic accumulation of USEs was 48% ± 28% lower than the accumulation of NSEs in mice (P < 0.05). Conclusions: Ultrasonication of milk depletes bioavailable BMEs in studies of Caco-2 cells, FHs 74 Int cells and C57BL/6J mice and causes a near-complete degradation of microRNA cargos. (hide)
EV-METRIC
67% (50th 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
1% skim milk
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
(Differential) (ultra)centrifugation
No extra separation steps
Protein markers
EV: TSG101/ Alix/ CD63/ CD9/ CD81
non-EV: Beta-integrin/ lactalbumin/ Histone-H3
Proteomics
yes
Show all info
Study aim
Function/New methodological development/Identification of content (omics approaches)/Mechanism of uptake/transfer
Sample
Species
Bos taurus
Sample Type
1% skim milk
Separation Method
(Differential) (ultra)centrifugation
dUC: centrifugation steps
Between 10,000 g and 50,000 g
Between 100,000 g and 150,000 g
Between 50,000 g and 100,000 g
Pelleting: time(min)
90
Pelleting: rotor type
Fiberlite-F37L-8x100 rotor
Pelleting: speed (g)
120,000
Wash: volume per pellet (ml)
30 mL
Wash: time (min)
90
Wash: Rotor Type
Fiberlite-F37L-8x100 rotor
Wash: speed (g)
120,000
Characterization: Protein analysis
Protein Concentration Method
BCA;Qubit
Western Blot
Antibody details provided?
No
Detected EV-associated proteins
CD9/ CD63/ TSG101/ Alix/ CD81
Detected contaminants
lactalbumin
Not detected contaminants
Beta-integrin/ Histone-H3
Proteomics database
No
Characterization: RNA analysis
RNA analysis
Type
(RT)(q)PCR;RNA sequencing
Database
Yes
Proteinase treatment
No
RNAse treatment
No
Characterization: Lipid analysis
Yes
Characterization: Particle analysis
NTA
Report type
Mean
Reported size (nm)
117+/-44
EV concentration
Yes
Particle yield
Yes, per milliliter of starting sample 5.90E+11
EM
EM-type
Transmission-EM/ Scanning-EM
Image type
Close-up, Wide-field
EV200174 2/2 Bos taurus 1% skim milk (d)(U)C Sukreet, Sonal 2022 67%

Study summary

Full title
All authors
Sonal Sukreet, Camila Pereira Braga, Thuy T An, Jiri Adamec, Juan Cui, Janos Zempleni
Journal
J Nutr
Abstract
Background: Bovine milk exosomes (BMEs) harbor regulatory proteins, lipids and microRNAs. Consumptio (show more...)Background: Bovine milk exosomes (BMEs) harbor regulatory proteins, lipids and microRNAs. Consumption of an exosome and RNA-depleted (ERD) diet elicited phenotypes compared to controls fed an exosome and RNA-sufficient (ERS) diet in mice. All other ingredients were identical in the diets. ERD and ERS diets were prepared by substituting ultrasonicated and non-ultrasonicated milk, respectively, for casein in the AIN-93G formulation. Objective: The objective of this study was to assess the effect of ultrasonication of milk on exosome content and bioavailability, and cargo content. Methods: Bovine milk was ultrasonicated and exosomes were isolated by ultracentrifugation (USE); controls were not ultrasonicated (NSE). Exosome count, size and morphology were assessed using a nanoparticle tracker and electron microscopy. RNAs, lipids and proteins were analyzed by RNA-sequencing and mass spectrometry. Intestinal transport, bioavailability and distribution were measured by using fluorophore-labeled USEs and NSEs in Caco-2 cells, FHs 74 Int cells and C57BL/6J mice (n = 3; age 6 - 8 weeks). Results: The exosome count was 76 ± 22% lower in USE compared to NSE (P < 0.05). Ultrasonication caused a degradation of up to 100% of microRNAs. USE and NSE contained 145 and 332 unique lipid signatures, respectively (P < 0.05). We detected total of 525 and 484 proteins in USE and NSE. The uptake of USEs decreased by 46 ± 30% and 40 ± 27% compared to NSEs in Caco-2 and FHs 74 Int cells, respectively (P < 0.05). The hepatic accumulation of USEs was 48% ± 28% lower than the accumulation of NSEs in mice (P < 0.05). Conclusions: Ultrasonication of milk depletes bioavailable BMEs in studies of Caco-2 cells, FHs 74 Int cells and C57BL/6J mice and causes a near-complete degradation of microRNA cargos. (hide)
EV-METRIC
67% (50th 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
1% skim milk
Sample origin
Sonicated milk
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
(Differential) (ultra)centrifugation
Protein markers
EV: CD81/ Alix/ TSG101/ CD63/ CD9
non-EV: Beta-integrin/ lactalbumin/ Histone H3
Proteomics
yes
Show all info
Study aim
Function/New methodological development/Identification of content (omics approaches)/Mechanism of uptake/transfer
Sample
Species
Bos taurus
Sample Type
1% skim milk
Separation Method
(Differential) (ultra)centrifugation
dUC: centrifugation steps
Between 10,000 g and 50,000 g
Between 100,000 g and 150,000 g
Between 50,000 g and 100,000 g
Pelleting: time(min)
90
Pelleting: rotor type
Fiberlite-F37L-8x100 rotor
Pelleting: speed (g)
120,000
Wash: volume per pellet (ml)
30 mL
Wash: time (min)
90
Wash: Rotor Type
Fiberlite-F37L-8x100 rotor
Wash: speed (g)
120,000
Characterization: Protein analysis
Protein Concentration Method
BCA;Qubit
Western Blot
Antibody details provided?
No
Detected EV-associated proteins
CD9/ CD63/ TSG101
Not detected EV-associated proteins
CD81/ Alix
Detected contaminants
lactalbumin
Not detected contaminants
Beta-integrin/ Histone H3
Proteomics database
No
Characterization: RNA analysis
RNA analysis
Type
(RT)(q)PCR;RNAsequencing
Database
Yes
Proteinase treatment
No
RNAse treatment
No
Characterization: Lipid analysis
Yes
Characterization: Particle analysis
NTA
Report type
Mean
Reported size (nm)
218+/-99
EV concentration
Yes
Particle yield
Yes, per milliliter of starting sample 1.40E+11
EM
EM-type
Transmission-EM/ Scanning-EM
Image type
Close-up, Wide-field
1 - 2 of 2
  • CM = Commercial method
  • dUC = differential ultracentrifugation
  • DG = density gradient
  • UF = ultrafiltration
  • SEC = size-exclusion chromatography
EV-TRACK ID
EV200174
species
Bos taurus
sample type
1% skim milk
condition
Control condition
Sonicated milk
separation protocol
dUC
No extra separation steps
dUC
Exp. nr.
1
2
EV-METRIC %
67
67