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You searched for: EV160012 (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.
    • dUC = differential ultracentrifugation
    • DG = density gradient
    • UF = ultrafiltration
    • SEC = size-exclusion chromatography
Details EV-TRACK ID Experiment nr. Species Sample type separation protocol First author Year EV-METRIC
EV160012 1/8 Homo sapiens Cell culture supernatant (Differential) (ultra)centrifugation
Filtration
Hannafon BN 2016 33%

Study summary

Full title
All authors
Hannafon BN, Trigoso YD, Calloway CL, Zhao YD, Lum DH, Welm AL, Zhao ZJ, Blick KE, Dooley WC, Ding WQ.
Journal
Breast Cancer Res Treat
Abstract
BACKGROUND: microRNAs are promising candidate breast cancer biomarkers due to their cancer-specific (show more...)BACKGROUND: microRNAs are promising candidate breast cancer biomarkers due to their cancer-specific expression profiles. However, efforts to develop circulating breast cancer biomarkers are challenged by the heterogeneity of microRNAs in the blood. To overcome this challenge, we aimed to develop a molecular profile of microRNAs specifically secreted from breast cancer cells. Our first step towards this direction relates to capturing and analyzing the contents of exosomes, which are small secretory vesicles that selectively encapsulate microRNAs indicative of their cell of origin. To our knowledge, circulating exosome microRNAs have not been well-evaluated as biomarkers for breast cancer diagnosis or monitoring. METHODS: Exosomes were collected from the conditioned media of human breast cancer cell lines, mouse plasma of patient-derived orthotopic xenograft models (PDX), and human plasma samples. Exosomes were verified by electron microscopy, nanoparticle tracking analysis, and western blot. Cellular and exosome microRNAs from breast cancer cell lines were profiled by next-generation small RNA sequencing. Plasma exosome microRNA expression was analyzed by qRT-PCR analysis. RESULTS: Small RNA sequencing and qRT-PCR analysis showed that several microRNAs are selectively encapsulated or highly enriched in breast cancer exosomes. Importantly, the selectively enriched exosome microRNA, human miR-1246, was detected at significantly higher levels in exosomes isolated from PDX mouse plasma, indicating that tumor exosome microRNAs are released into the circulation and can serve as plasma biomarkers for breast cancer. This observation was extended to human plasma samples where miR-1246 and miR-21 were detected at significantly higher levels in the plasma exosomes of 16 patients with breast cancer as compared to the plasma exosomes of healthy control subjects. Receiver operating characteristic curve analysis indicated that the combination of plasma exosome miR-1246 and miR-21 is a better indicator of breast cancer than their individual levels. CONCLUSIONS: Our results demonstrate that certain microRNA species, such as miR-21 and miR-1246, are selectively enriched in human breast cancer exosomes and significantly elevated in the plasma of patients with breast cancer. These findings indicate a potential new strategy to selectively analyze plasma breast cancer microRNAs indicative of the presence of breast cancer. (hide)
EV-METRIC
33% (67th 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
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
exosome
Separation protocol
Separation protocol
  • Gives a short, non-chronological overview of the
    different steps of the separation protocol.
    • dUC = differential ultracentrifugation
    • DG = density gradient
    • UF = ultrafiltration
    • SEC = size-exclusion chromatography
(Differential) (ultra)centrifugation + Filtration
Protein markers
EV: CD63
non-EV:
Proteomics
no
Show all info
Study aim
Biomarker/Identification of content (omics approaches)
Sample
Species
Homo sapiens
Sample Type
Cell culture supernatant
Sample Condition
Control condition
EV-producing cells
MCF7
EV-harvesting Medium
EV-depleted medium
Preparation of EDS
2h at 100,000g
Separation Method
Differential ultracentrifugation
dUC: centrifugation steps
Between 10,000 g and 50,000 g
Between 100,000 g and 150,000 g
Pelleting: time(min)
60
Pelleting: rotor type
Not specified
Pelleting: speed (g)
100000
Wash: volume per pellet (ml)
10
Wash: time (min)
60
Wash: Rotor Type
Not specified
Wash: speed (g)
100000
Filtration steps
0.22µm or 0.2µm
Characterization: Protein analysis
Protein Concentration Method
Bradford
Western Blot
Detected EV-associated proteins
CD63
Characterization: Particle analysis
NTA
Report type
Mean
Reported size (nm)
103.3
EV concentration
Yes
EM
EM-type
Immuno-EM
Proteïns
CD63
Image type
Close-up, Wide-field
EV160012 2/8 Homo sapiens Cell culture supernatant (Differential) (ultra)centrifugation
Filtration
Hannafon BN 2016 33%

Study summary

Full title
All authors
Hannafon BN, Trigoso YD, Calloway CL, Zhao YD, Lum DH, Welm AL, Zhao ZJ, Blick KE, Dooley WC, Ding WQ.
Journal
Breast Cancer Res Treat
Abstract
BACKGROUND: microRNAs are promising candidate breast cancer biomarkers due to their cancer-specific (show more...)BACKGROUND: microRNAs are promising candidate breast cancer biomarkers due to their cancer-specific expression profiles. However, efforts to develop circulating breast cancer biomarkers are challenged by the heterogeneity of microRNAs in the blood. To overcome this challenge, we aimed to develop a molecular profile of microRNAs specifically secreted from breast cancer cells. Our first step towards this direction relates to capturing and analyzing the contents of exosomes, which are small secretory vesicles that selectively encapsulate microRNAs indicative of their cell of origin. To our knowledge, circulating exosome microRNAs have not been well-evaluated as biomarkers for breast cancer diagnosis or monitoring. METHODS: Exosomes were collected from the conditioned media of human breast cancer cell lines, mouse plasma of patient-derived orthotopic xenograft models (PDX), and human plasma samples. Exosomes were verified by electron microscopy, nanoparticle tracking analysis, and western blot. Cellular and exosome microRNAs from breast cancer cell lines were profiled by next-generation small RNA sequencing. Plasma exosome microRNA expression was analyzed by qRT-PCR analysis. RESULTS: Small RNA sequencing and qRT-PCR analysis showed that several microRNAs are selectively encapsulated or highly enriched in breast cancer exosomes. Importantly, the selectively enriched exosome microRNA, human miR-1246, was detected at significantly higher levels in exosomes isolated from PDX mouse plasma, indicating that tumor exosome microRNAs are released into the circulation and can serve as plasma biomarkers for breast cancer. This observation was extended to human plasma samples where miR-1246 and miR-21 were detected at significantly higher levels in the plasma exosomes of 16 patients with breast cancer as compared to the plasma exosomes of healthy control subjects. Receiver operating characteristic curve analysis indicated that the combination of plasma exosome miR-1246 and miR-21 is a better indicator of breast cancer than their individual levels. CONCLUSIONS: Our results demonstrate that certain microRNA species, such as miR-21 and miR-1246, are selectively enriched in human breast cancer exosomes and significantly elevated in the plasma of patients with breast cancer. These findings indicate a potential new strategy to selectively analyze plasma breast cancer microRNAs indicative of the presence of breast cancer. (hide)
EV-METRIC
33% (67th 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
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
exosome
Separation protocol
Separation protocol
  • Gives a short, non-chronological overview of the
    different steps of the separation protocol.
    • dUC = differential ultracentrifugation
    • DG = density gradient
    • UF = ultrafiltration
    • SEC = size-exclusion chromatography
(Differential) (ultra)centrifugation + Filtration
Protein markers
EV: CD63
non-EV:
Proteomics
no
Show all info
Study aim
Biomarker/Identification of content (omics approaches)
Sample
Species
Homo sapiens
Sample Type
Cell culture supernatant
Sample Condition
Control condition
EV-producing cells
MDAMB231
EV-harvesting Medium
EV-depleted medium
Preparation of EDS
2h at 100,000g
Separation Method
Differential ultracentrifugation
dUC: centrifugation steps
Between 10,000 g and 50,000 g
Between 100,000 g and 150,000 g
Pelleting: time(min)
60
Pelleting: rotor type
Not specified
Pelleting: speed (g)
100000
Wash: volume per pellet (ml)
10
Wash: time (min)
60
Wash: Rotor Type
Not specified
Wash: speed (g)
100000
Filtration steps
0.22µm or 0.2µm
Characterization: Protein analysis
Protein Concentration Method
Bradford
Western Blot
Detected EV-associated proteins
CD63
Characterization: Particle analysis
NTA
Report type
Mean
Reported size (nm)
99.6
EV concentration
Yes
EM
EM-type
Immuno-EM
Proteïns
CD63
Image type
Close-up, Wide-field
EV160012 3/8 Homo sapiens Cell culture supernatant (Differential) (ultra)centrifugation
Filtration
Hannafon BN 2016 11%

Study summary

Full title
All authors
Hannafon BN, Trigoso YD, Calloway CL, Zhao YD, Lum DH, Welm AL, Zhao ZJ, Blick KE, Dooley WC, Ding WQ.
Journal
Breast Cancer Res Treat
Abstract
BACKGROUND: microRNAs are promising candidate breast cancer biomarkers due to their cancer-specific (show more...)BACKGROUND: microRNAs are promising candidate breast cancer biomarkers due to their cancer-specific expression profiles. However, efforts to develop circulating breast cancer biomarkers are challenged by the heterogeneity of microRNAs in the blood. To overcome this challenge, we aimed to develop a molecular profile of microRNAs specifically secreted from breast cancer cells. Our first step towards this direction relates to capturing and analyzing the contents of exosomes, which are small secretory vesicles that selectively encapsulate microRNAs indicative of their cell of origin. To our knowledge, circulating exosome microRNAs have not been well-evaluated as biomarkers for breast cancer diagnosis or monitoring. METHODS: Exosomes were collected from the conditioned media of human breast cancer cell lines, mouse plasma of patient-derived orthotopic xenograft models (PDX), and human plasma samples. Exosomes were verified by electron microscopy, nanoparticle tracking analysis, and western blot. Cellular and exosome microRNAs from breast cancer cell lines were profiled by next-generation small RNA sequencing. Plasma exosome microRNA expression was analyzed by qRT-PCR analysis. RESULTS: Small RNA sequencing and qRT-PCR analysis showed that several microRNAs are selectively encapsulated or highly enriched in breast cancer exosomes. Importantly, the selectively enriched exosome microRNA, human miR-1246, was detected at significantly higher levels in exosomes isolated from PDX mouse plasma, indicating that tumor exosome microRNAs are released into the circulation and can serve as plasma biomarkers for breast cancer. This observation was extended to human plasma samples where miR-1246 and miR-21 were detected at significantly higher levels in the plasma exosomes of 16 patients with breast cancer as compared to the plasma exosomes of healthy control subjects. Receiver operating characteristic curve analysis indicated that the combination of plasma exosome miR-1246 and miR-21 is a better indicator of breast cancer than their individual levels. CONCLUSIONS: Our results demonstrate that certain microRNA species, such as miR-21 and miR-1246, are selectively enriched in human breast cancer exosomes and significantly elevated in the plasma of patients with breast cancer. These findings indicate a potential new strategy to selectively analyze plasma breast cancer microRNAs indicative of the presence of breast cancer. (hide)
EV-METRIC
11% (26th 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
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
exosome
Separation protocol
Separation protocol
  • Gives a short, non-chronological overview of the
    different steps of the separation protocol.
    • dUC = differential ultracentrifugation
    • DG = density gradient
    • UF = ultrafiltration
    • SEC = size-exclusion chromatography
(Differential) (ultra)centrifugation + Filtration
Protein markers
EV: CD63
non-EV:
Proteomics
no
Show all info
Study aim
Biomarker/Identification of content (omics approaches)
Sample
Species
Homo sapiens
Sample Type
Cell culture supernatant
Sample Condition
Control condition
EV-producing cells
MCF10A
EV-harvesting Medium
EV-depleted medium
Preparation of EDS
2h at 100,000g
Separation Method
Differential ultracentrifugation
dUC: centrifugation steps
Between 10,000 g and 50,000 g
Between 100,000 g and 150,000 g
Pelleting: time(min)
60
Pelleting: rotor type
Not specified
Pelleting: speed (g)
100000
Wash: volume per pellet (ml)
10
Wash: time (min)
60
Wash: Rotor Type
Not specified
Wash: speed (g)
100000
Filtration steps
0.22µm or 0.2µm
Characterization: Protein analysis
Protein Concentration Method
Bradford
Western Blot
Detected EV-associated proteins
CD63
Characterization: Particle analysis
NTA
Report type
Mean
Reported size (nm)
126.4
EV concentration
Yes
EV160012 4/8 Homo sapiens Cell culture supernatant (Differential) (ultra)centrifugation
Filtration
Hannafon BN 2016 0%

Study summary

Full title
All authors
Hannafon BN, Trigoso YD, Calloway CL, Zhao YD, Lum DH, Welm AL, Zhao ZJ, Blick KE, Dooley WC, Ding WQ.
Journal
Breast Cancer Res Treat
Abstract
BACKGROUND: microRNAs are promising candidate breast cancer biomarkers due to their cancer-specific (show more...)BACKGROUND: microRNAs are promising candidate breast cancer biomarkers due to their cancer-specific expression profiles. However, efforts to develop circulating breast cancer biomarkers are challenged by the heterogeneity of microRNAs in the blood. To overcome this challenge, we aimed to develop a molecular profile of microRNAs specifically secreted from breast cancer cells. Our first step towards this direction relates to capturing and analyzing the contents of exosomes, which are small secretory vesicles that selectively encapsulate microRNAs indicative of their cell of origin. To our knowledge, circulating exosome microRNAs have not been well-evaluated as biomarkers for breast cancer diagnosis or monitoring. METHODS: Exosomes were collected from the conditioned media of human breast cancer cell lines, mouse plasma of patient-derived orthotopic xenograft models (PDX), and human plasma samples. Exosomes were verified by electron microscopy, nanoparticle tracking analysis, and western blot. Cellular and exosome microRNAs from breast cancer cell lines were profiled by next-generation small RNA sequencing. Plasma exosome microRNA expression was analyzed by qRT-PCR analysis. RESULTS: Small RNA sequencing and qRT-PCR analysis showed that several microRNAs are selectively encapsulated or highly enriched in breast cancer exosomes. Importantly, the selectively enriched exosome microRNA, human miR-1246, was detected at significantly higher levels in exosomes isolated from PDX mouse plasma, indicating that tumor exosome microRNAs are released into the circulation and can serve as plasma biomarkers for breast cancer. This observation was extended to human plasma samples where miR-1246 and miR-21 were detected at significantly higher levels in the plasma exosomes of 16 patients with breast cancer as compared to the plasma exosomes of healthy control subjects. Receiver operating characteristic curve analysis indicated that the combination of plasma exosome miR-1246 and miR-21 is a better indicator of breast cancer than their individual levels. CONCLUSIONS: Our results demonstrate that certain microRNA species, such as miR-21 and miR-1246, are selectively enriched in human breast cancer exosomes and significantly elevated in the plasma of patients with breast cancer. These findings indicate a potential new strategy to selectively analyze plasma breast cancer microRNAs indicative of the presence of breast cancer. (hide)
EV-METRIC
0% (median: 22% 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
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
exosome
Separation protocol
Separation protocol
  • Gives a short, non-chronological overview of the
    different steps of the separation protocol.
    • dUC = differential ultracentrifugation
    • DG = density gradient
    • UF = ultrafiltration
    • SEC = size-exclusion chromatography
(Differential) (ultra)centrifugation + Filtration
Protein markers
EV:
non-EV:
Proteomics
no
Show all info
Study aim
Biomarker/Identification of content (omics approaches)
Sample
Species
Homo sapiens
Sample Type
Cell culture supernatant
Sample Condition
Control condition
EV-producing cells
ZR-75-1 / T47D / BT20 / BT-474 / SK-BR-3
EV-harvesting Medium
EV-depleted medium
Preparation of EDS
2h at 100,000g
Separation Method
Differential ultracentrifugation
dUC: centrifugation steps
Between 10,000 g and 50,000 g
Between 100,000 g and 150,000 g
Pelleting: time(min)
60
Pelleting: rotor type
Not specified
Pelleting: speed (g)
100000
Wash: volume per pellet (ml)
10
Wash: time (min)
60
Wash: Rotor Type
Not specified
Wash: speed (g)
100000
Filtration steps
0.22µm or 0.2µm
Protein Concentration Method
Bradford
Characterization: Particle analysis
EV160012 5/8 Homo sapiens Blood plasma ExoQuick Hannafon BN 2016 0%

Study summary

Full title
All authors
Hannafon BN, Trigoso YD, Calloway CL, Zhao YD, Lum DH, Welm AL, Zhao ZJ, Blick KE, Dooley WC, Ding WQ.
Journal
Breast Cancer Res Treat
Abstract
BACKGROUND: microRNAs are promising candidate breast cancer biomarkers due to their cancer-specific (show more...)BACKGROUND: microRNAs are promising candidate breast cancer biomarkers due to their cancer-specific expression profiles. However, efforts to develop circulating breast cancer biomarkers are challenged by the heterogeneity of microRNAs in the blood. To overcome this challenge, we aimed to develop a molecular profile of microRNAs specifically secreted from breast cancer cells. Our first step towards this direction relates to capturing and analyzing the contents of exosomes, which are small secretory vesicles that selectively encapsulate microRNAs indicative of their cell of origin. To our knowledge, circulating exosome microRNAs have not been well-evaluated as biomarkers for breast cancer diagnosis or monitoring. METHODS: Exosomes were collected from the conditioned media of human breast cancer cell lines, mouse plasma of patient-derived orthotopic xenograft models (PDX), and human plasma samples. Exosomes were verified by electron microscopy, nanoparticle tracking analysis, and western blot. Cellular and exosome microRNAs from breast cancer cell lines were profiled by next-generation small RNA sequencing. Plasma exosome microRNA expression was analyzed by qRT-PCR analysis. RESULTS: Small RNA sequencing and qRT-PCR analysis showed that several microRNAs are selectively encapsulated or highly enriched in breast cancer exosomes. Importantly, the selectively enriched exosome microRNA, human miR-1246, was detected at significantly higher levels in exosomes isolated from PDX mouse plasma, indicating that tumor exosome microRNAs are released into the circulation and can serve as plasma biomarkers for breast cancer. This observation was extended to human plasma samples where miR-1246 and miR-21 were detected at significantly higher levels in the plasma exosomes of 16 patients with breast cancer as compared to the plasma exosomes of healthy control subjects. Receiver operating characteristic curve analysis indicated that the combination of plasma exosome miR-1246 and miR-21 is a better indicator of breast cancer than their individual levels. CONCLUSIONS: Our results demonstrate that certain microRNA species, such as miR-21 and miR-1246, are selectively enriched in human breast cancer exosomes and significantly elevated in the plasma of patients with breast cancer. These findings indicate a potential new strategy to selectively analyze plasma breast cancer microRNAs indicative of the presence of breast cancer. (hide)
EV-METRIC
0% (median: 14% 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
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 ultracentrifugation
    • DG = density gradient
    • UF = ultrafiltration
    • SEC = size-exclusion chromatography
ExoQuick
Protein markers
EV:
non-EV:
Proteomics
no
Show all info
Study aim
Biomarker/Identification of content (omics approaches)
Sample
Species
Homo sapiens
Sample Type
Blood plasma
Sample Condition
Control condition
Separation Method
Commercial kit
ExoQuick
Protein Concentration Method
Bradford
EM
EM-type
Transmission-EM
Image type
Close-up
EV160012 6/8 Homo sapiens Blood plasma ExoQuick Hannafon BN 2016 0%

Study summary

Full title
All authors
Hannafon BN, Trigoso YD, Calloway CL, Zhao YD, Lum DH, Welm AL, Zhao ZJ, Blick KE, Dooley WC, Ding WQ.
Journal
Breast Cancer Res Treat
Abstract
BACKGROUND: microRNAs are promising candidate breast cancer biomarkers due to their cancer-specific (show more...)BACKGROUND: microRNAs are promising candidate breast cancer biomarkers due to their cancer-specific expression profiles. However, efforts to develop circulating breast cancer biomarkers are challenged by the heterogeneity of microRNAs in the blood. To overcome this challenge, we aimed to develop a molecular profile of microRNAs specifically secreted from breast cancer cells. Our first step towards this direction relates to capturing and analyzing the contents of exosomes, which are small secretory vesicles that selectively encapsulate microRNAs indicative of their cell of origin. To our knowledge, circulating exosome microRNAs have not been well-evaluated as biomarkers for breast cancer diagnosis or monitoring. METHODS: Exosomes were collected from the conditioned media of human breast cancer cell lines, mouse plasma of patient-derived orthotopic xenograft models (PDX), and human plasma samples. Exosomes were verified by electron microscopy, nanoparticle tracking analysis, and western blot. Cellular and exosome microRNAs from breast cancer cell lines were profiled by next-generation small RNA sequencing. Plasma exosome microRNA expression was analyzed by qRT-PCR analysis. RESULTS: Small RNA sequencing and qRT-PCR analysis showed that several microRNAs are selectively encapsulated or highly enriched in breast cancer exosomes. Importantly, the selectively enriched exosome microRNA, human miR-1246, was detected at significantly higher levels in exosomes isolated from PDX mouse plasma, indicating that tumor exosome microRNAs are released into the circulation and can serve as plasma biomarkers for breast cancer. This observation was extended to human plasma samples where miR-1246 and miR-21 were detected at significantly higher levels in the plasma exosomes of 16 patients with breast cancer as compared to the plasma exosomes of healthy control subjects. Receiver operating characteristic curve analysis indicated that the combination of plasma exosome miR-1246 and miR-21 is a better indicator of breast cancer than their individual levels. CONCLUSIONS: Our results demonstrate that certain microRNA species, such as miR-21 and miR-1246, are selectively enriched in human breast cancer exosomes and significantly elevated in the plasma of patients with breast cancer. These findings indicate a potential new strategy to selectively analyze plasma breast cancer microRNAs indicative of the presence of breast cancer. (hide)
EV-METRIC
0% (median: 14% 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
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
Breast cancer
Focus vesicles
exosome
Separation protocol
Separation protocol
  • Gives a short, non-chronological overview of the
    different steps of the separation protocol.
    • dUC = differential ultracentrifugation
    • DG = density gradient
    • UF = ultrafiltration
    • SEC = size-exclusion chromatography
ExoQuick
Protein markers
EV:
non-EV:
Proteomics
no
Show all info
Study aim
Biomarker/Identification of content (omics approaches)
Sample
Species
Homo sapiens
Sample Type
Blood plasma
Sample Condition
Breast cancer
Separation Method
Commercial kit
ExoQuick
Protein Concentration Method
Bradford
EV160012 7/8 Mus musculus Blood plasma Immunoaffinity capture (non-commercial)
ExoQuick
Hannafon BN 2016 0%

Study summary

Full title
All authors
Hannafon BN, Trigoso YD, Calloway CL, Zhao YD, Lum DH, Welm AL, Zhao ZJ, Blick KE, Dooley WC, Ding WQ.
Journal
Breast Cancer Res Treat
Abstract
BACKGROUND: microRNAs are promising candidate breast cancer biomarkers due to their cancer-specific (show more...)BACKGROUND: microRNAs are promising candidate breast cancer biomarkers due to their cancer-specific expression profiles. However, efforts to develop circulating breast cancer biomarkers are challenged by the heterogeneity of microRNAs in the blood. To overcome this challenge, we aimed to develop a molecular profile of microRNAs specifically secreted from breast cancer cells. Our first step towards this direction relates to capturing and analyzing the contents of exosomes, which are small secretory vesicles that selectively encapsulate microRNAs indicative of their cell of origin. To our knowledge, circulating exosome microRNAs have not been well-evaluated as biomarkers for breast cancer diagnosis or monitoring. METHODS: Exosomes were collected from the conditioned media of human breast cancer cell lines, mouse plasma of patient-derived orthotopic xenograft models (PDX), and human plasma samples. Exosomes were verified by electron microscopy, nanoparticle tracking analysis, and western blot. Cellular and exosome microRNAs from breast cancer cell lines were profiled by next-generation small RNA sequencing. Plasma exosome microRNA expression was analyzed by qRT-PCR analysis. RESULTS: Small RNA sequencing and qRT-PCR analysis showed that several microRNAs are selectively encapsulated or highly enriched in breast cancer exosomes. Importantly, the selectively enriched exosome microRNA, human miR-1246, was detected at significantly higher levels in exosomes isolated from PDX mouse plasma, indicating that tumor exosome microRNAs are released into the circulation and can serve as plasma biomarkers for breast cancer. This observation was extended to human plasma samples where miR-1246 and miR-21 were detected at significantly higher levels in the plasma exosomes of 16 patients with breast cancer as compared to the plasma exosomes of healthy control subjects. Receiver operating characteristic curve analysis indicated that the combination of plasma exosome miR-1246 and miR-21 is a better indicator of breast cancer than their individual levels. CONCLUSIONS: Our results demonstrate that certain microRNA species, such as miR-21 and miR-1246, are selectively enriched in human breast cancer exosomes and significantly elevated in the plasma of patients with breast cancer. These findings indicate a potential new strategy to selectively analyze plasma breast cancer microRNAs indicative of the presence of breast cancer. (hide)
EV-METRIC
0% (median: 14% 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
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
PDX breast cancer model
Focus vesicles
exosome
Separation protocol
Separation protocol
  • Gives a short, non-chronological overview of the
    different steps of the separation protocol.
    • dUC = differential ultracentrifugation
    • DG = density gradient
    • UF = ultrafiltration
    • SEC = size-exclusion chromatography
Immunoaffinity capture (non-commercial) + ExoQuick
Protein markers
EV:
non-EV:
Proteomics
no
Show all info
Study aim
Biomarker/Identification of content (omics approaches)
Sample
Species
Mus musculus
Sample Type
Blood plasma
Sample Condition
PDX breast cancer model
Separation Method
Commercial kit
ExoQuick
Protein Concentration Method
Bradford
Characterization: Particle analysis
NTA
Report type
Mean
Reported size (nm)
84.6
EV concentration
Yes
EV160012 8/8 Mus musculus Blood plasma Immunoaffinity capture (non-commercial)
ExoQuick
Hannafon BN 2016 0%

Study summary

Full title
All authors
Hannafon BN, Trigoso YD, Calloway CL, Zhao YD, Lum DH, Welm AL, Zhao ZJ, Blick KE, Dooley WC, Ding WQ.
Journal
Breast Cancer Res Treat
Abstract
BACKGROUND: microRNAs are promising candidate breast cancer biomarkers due to their cancer-specific (show more...)BACKGROUND: microRNAs are promising candidate breast cancer biomarkers due to their cancer-specific expression profiles. However, efforts to develop circulating breast cancer biomarkers are challenged by the heterogeneity of microRNAs in the blood. To overcome this challenge, we aimed to develop a molecular profile of microRNAs specifically secreted from breast cancer cells. Our first step towards this direction relates to capturing and analyzing the contents of exosomes, which are small secretory vesicles that selectively encapsulate microRNAs indicative of their cell of origin. To our knowledge, circulating exosome microRNAs have not been well-evaluated as biomarkers for breast cancer diagnosis or monitoring. METHODS: Exosomes were collected from the conditioned media of human breast cancer cell lines, mouse plasma of patient-derived orthotopic xenograft models (PDX), and human plasma samples. Exosomes were verified by electron microscopy, nanoparticle tracking analysis, and western blot. Cellular and exosome microRNAs from breast cancer cell lines were profiled by next-generation small RNA sequencing. Plasma exosome microRNA expression was analyzed by qRT-PCR analysis. RESULTS: Small RNA sequencing and qRT-PCR analysis showed that several microRNAs are selectively encapsulated or highly enriched in breast cancer exosomes. Importantly, the selectively enriched exosome microRNA, human miR-1246, was detected at significantly higher levels in exosomes isolated from PDX mouse plasma, indicating that tumor exosome microRNAs are released into the circulation and can serve as plasma biomarkers for breast cancer. This observation was extended to human plasma samples where miR-1246 and miR-21 were detected at significantly higher levels in the plasma exosomes of 16 patients with breast cancer as compared to the plasma exosomes of healthy control subjects. Receiver operating characteristic curve analysis indicated that the combination of plasma exosome miR-1246 and miR-21 is a better indicator of breast cancer than their individual levels. CONCLUSIONS: Our results demonstrate that certain microRNA species, such as miR-21 and miR-1246, are selectively enriched in human breast cancer exosomes and significantly elevated in the plasma of patients with breast cancer. These findings indicate a potential new strategy to selectively analyze plasma breast cancer microRNAs indicative of the presence of breast cancer. (hide)
EV-METRIC
0% (median: 14% 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
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 ultracentrifugation
    • DG = density gradient
    • UF = ultrafiltration
    • SEC = size-exclusion chromatography
Immunoaffinity capture (non-commercial) + ExoQuick
Protein markers
EV:
non-EV:
Proteomics
no
Show all info
Study aim
Biomarker/Identification of content (omics approaches)
Sample
Species
Mus musculus
Sample Type
Blood plasma
Sample Condition
Control condition
Separation Method
Commercial kit
ExoQuick
Protein Concentration Method
Bradford
Characterization: Particle analysis
1 - 8 of 8
  • CM = Commercial method
  • dUC = differential ultracentrifugation
  • DG = density gradient
  • UF = ultrafiltration
  • SEC = size-exclusion chromatography
EV-TRACK ID
EV160012
species
Homo
sapiens
Homo
sapiens
Homo
sapiens
Homo
sapiens
Homo
sapiens
Homo
sapiens
Mus
musculus
Mus
musculus
sample type
Cell
culture
Cell
culture
Cell
culture
Cell
culture
Blood
plasma
Blood
plasma
Blood
plasma
Blood
plasma
cell type
MCF7
MDAMB231
MCF10A
ZR-75-1
T47D
BT20
BT-474
SK-BR-3
NA
NA
NA
NA
medium
EV-depleted
medium
EV-depleted
medium
EV-depleted
medium
EV-depleted
medium
NA
NA
NA
NA
condition
Control
condition
Control
condition
Control
condition
Control
condition
Control
condition
Breast
cancer
PDX
breast
cancer
model
Control
condition
separation protocol
dUC
Filtration
dUC
Filtration
dUC
Filtration
dUC
Filtration
ExoQuick
ExoQuick
IAF
capture
(non-commercial)
ExoQuick
IAF
capture
(non-commercial)
ExoQuick
Exp. nr.
1
2
3
4
5
6
7
8
EV-METRIC %
33
33
11
0
0
0
0
0