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Details	EV-TRACK ID	Experiment nr.	Species	Sample type	Separation protocol	First author	Year	EV-METRIC
	EV200193	1/4	Homo sapiens	Urine	(d)(U)C	Bryzgunova, Olga E	2016	25%
Study summary Full title Comparative Study of Extracellular Vesicles from the Urine of Healthy Individuals and Prostate Cancer Patients All authors Olga E Bryzgunova, Marat M Zaripov, Tatyana E Skvortsova, Evgeny A Lekchnov, Alina E Grigor'eva, Ivan A Zaporozhchenko, Evgeny S Morozkin, Elena I Ryabchikova, Yuri B Yurchenko, Vladimir E Voitsitskiy, Pavel P Laktionov Journal PLoS One Abstract Recent studies suggest that extracellular vesicles may be the key to timely diagnosis and monitoring (show more...)Recent studies suggest that extracellular vesicles may be the key to timely diagnosis and monitoring of genito-urological malignancies. In this study we investigated the composition and content of extracellular vesicles found in the urine of healthy donors and prostate cancer patients. Urine of 14 PCa patients and 20 healthy volunteers was clarified by low-speed centrifugation and total extracellular vesicles fraction was obtain by high-speed centrifugation. The exosome-enriched fraction was obtained by filtration of total extracellular vesicles through a 0.1 μm pore filter. Transmission electron microscopy showed that cell-free urine in both groups contained vesicles from 20 to 230 nm. Immunogold staining after ultrafiltration demonstrated that 95% and 90% of extracellular vesicles in healthy individuals and cancer patients, respectively, were exosomes. Protein, DNA and RNA concentrations as well as size distribution of extracellular vesicles in both fractions were analyzed. Only 75% of the total protein content of extracellular vesicles was associated with exosomes which amounted to 90-95% of all vesicles. Median DNA concentrations in total extracellular vesicles and exosome-enriched fractions were 18 pg/ml and 2.6 pg/ml urine, correspondingly. Urine extracellular vesicles carried a population of RNA molecules 25 nt to 200 nt in concentration of no more than 290 pg/ml of urine. Additionally, concentrations of miR-19b, miR-25, miR-125b, and miR-205 were quantified by qRT-PCR. MiRNAs were shown to be differently distributed between different fractions of extracellular vesicles. Detection of miR-19b versus miR-16 in total vesicles and exosome-enriched fractions achieved 100%/93% and 95%/79% specificity/sensitivity in distinguishing cancer patients from healthy individuals, respectively, demonstrating the diagnostic value of urine extracellular vesicles. (hide) EV-METRIC 25% (56th 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 Urine Sample origin Prostate cancer 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 Protein markers EV: CD9/ CD63/ CD24 non-EV: None Proteomics no Show all info Study aim Biomarker Sample Species Homo sapiens Sample Type Urine Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Below or equal to 800 g Between 10,000 g and 50,000 g Between 100,000 g and 150,000 g Pelleting performed Yes Pelleting: time(min) 90 Pelleting: rotor type Not specified Pelleting: speed (g) 100000 Wash: volume per pellet (ml) 10 Wash: time (min) 90 Wash: Rotor Type Not specified Wash: speed (g) 100000 Filtration steps 0.2µm > x > 0.1µm Characterization: Protein analysis None Protein Concentration Method Fluorometric assay (e.g. Qubit, NanoOrange,...) Characterization: RNA analysis RNA analysis Type (RT)(q)PCR Database No Proteinase treatment No RNAse treatment No Characterization: Lipid analysis No EM EM-type Immuno-EM/ Transmission-EM EM protein Other;CD9;CD63;CD24 Image type Close-up, Wide-field Report size (nm) 20-230

	EV200193	2/4	Homo sapiens	Urine	(d)(U)C Filtration	Bryzgunova, Olga E	2016	25%
Study summary Full title Comparative Study of Extracellular Vesicles from the Urine of Healthy Individuals and Prostate Cancer Patients All authors Olga E Bryzgunova, Marat M Zaripov, Tatyana E Skvortsova, Evgeny A Lekchnov, Alina E Grigor'eva, Ivan A Zaporozhchenko, Evgeny S Morozkin, Elena I Ryabchikova, Yuri B Yurchenko, Vladimir E Voitsitskiy, Pavel P Laktionov Journal PLoS One Abstract Recent studies suggest that extracellular vesicles may be the key to timely diagnosis and monitoring (show more...)Recent studies suggest that extracellular vesicles may be the key to timely diagnosis and monitoring of genito-urological malignancies. In this study we investigated the composition and content of extracellular vesicles found in the urine of healthy donors and prostate cancer patients. Urine of 14 PCa patients and 20 healthy volunteers was clarified by low-speed centrifugation and total extracellular vesicles fraction was obtain by high-speed centrifugation. The exosome-enriched fraction was obtained by filtration of total extracellular vesicles through a 0.1 μm pore filter. Transmission electron microscopy showed that cell-free urine in both groups contained vesicles from 20 to 230 nm. Immunogold staining after ultrafiltration demonstrated that 95% and 90% of extracellular vesicles in healthy individuals and cancer patients, respectively, were exosomes. Protein, DNA and RNA concentrations as well as size distribution of extracellular vesicles in both fractions were analyzed. Only 75% of the total protein content of extracellular vesicles was associated with exosomes which amounted to 90-95% of all vesicles. Median DNA concentrations in total extracellular vesicles and exosome-enriched fractions were 18 pg/ml and 2.6 pg/ml urine, correspondingly. Urine extracellular vesicles carried a population of RNA molecules 25 nt to 200 nt in concentration of no more than 290 pg/ml of urine. Additionally, concentrations of miR-19b, miR-25, miR-125b, and miR-205 were quantified by qRT-PCR. MiRNAs were shown to be differently distributed between different fractions of extracellular vesicles. Detection of miR-19b versus miR-16 in total vesicles and exosome-enriched fractions achieved 100%/93% and 95%/79% specificity/sensitivity in distinguishing cancer patients from healthy individuals, respectively, demonstrating the diagnostic value of urine extracellular vesicles. (hide) EV-METRIC 25% (56th 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 Urine Sample origin Prostate cancer 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/ CD24 non-EV: None Proteomics no Show all info Study aim Biomarker Sample Species Homo sapiens Sample Type Urine Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Below or equal to 800 g Between 10,000 g and 50,000 g Between 100,000 g and 150,000 g Pelleting performed Yes Pelleting: time(min) 90 Pelleting: rotor type Not specified Pelleting: speed (g) 100000 Wash: volume per pellet (ml) 10 Wash: time (min) 90 Wash: Rotor Type Not specified Wash: speed (g) 100000 Filtration steps 0.2µm > x > 0.1µm EV-subtype Distinction between multiple subtypes Size Used subtypes 30–100 nm enriched Characterization: Protein analysis None Protein Concentration Method Fluorometric assay (e.g. Qubit, NanoOrange,...) Characterization: RNA analysis RNA analysis Type (RT)(q)PCR;Capillary electrophoresis (e.g. Bioanalyzer) Database No Proteinase treatment No RNAse treatment No Characterization: Lipid analysis No EM EM-type Immuno-EM/ Transmission-EM EM protein Other;CD9;CD63;CD24 Image type Close-up, Wide-field Report size (nm) 30-100

	EV200193	3/4	Homo sapiens	Urine	(d)(U)C	Bryzgunova, Olga E	2016	25%
Study summary Full title Comparative Study of Extracellular Vesicles from the Urine of Healthy Individuals and Prostate Cancer Patients All authors Olga E Bryzgunova, Marat M Zaripov, Tatyana E Skvortsova, Evgeny A Lekchnov, Alina E Grigor'eva, Ivan A Zaporozhchenko, Evgeny S Morozkin, Elena I Ryabchikova, Yuri B Yurchenko, Vladimir E Voitsitskiy, Pavel P Laktionov Journal PLoS One Abstract Recent studies suggest that extracellular vesicles may be the key to timely diagnosis and monitoring (show more...)Recent studies suggest that extracellular vesicles may be the key to timely diagnosis and monitoring of genito-urological malignancies. In this study we investigated the composition and content of extracellular vesicles found in the urine of healthy donors and prostate cancer patients. Urine of 14 PCa patients and 20 healthy volunteers was clarified by low-speed centrifugation and total extracellular vesicles fraction was obtain by high-speed centrifugation. The exosome-enriched fraction was obtained by filtration of total extracellular vesicles through a 0.1 μm pore filter. Transmission electron microscopy showed that cell-free urine in both groups contained vesicles from 20 to 230 nm. Immunogold staining after ultrafiltration demonstrated that 95% and 90% of extracellular vesicles in healthy individuals and cancer patients, respectively, were exosomes. Protein, DNA and RNA concentrations as well as size distribution of extracellular vesicles in both fractions were analyzed. Only 75% of the total protein content of extracellular vesicles was associated with exosomes which amounted to 90-95% of all vesicles. Median DNA concentrations in total extracellular vesicles and exosome-enriched fractions were 18 pg/ml and 2.6 pg/ml urine, correspondingly. Urine extracellular vesicles carried a population of RNA molecules 25 nt to 200 nt in concentration of no more than 290 pg/ml of urine. Additionally, concentrations of miR-19b, miR-25, miR-125b, and miR-205 were quantified by qRT-PCR. MiRNAs were shown to be differently distributed between different fractions of extracellular vesicles. Detection of miR-19b versus miR-16 in total vesicles and exosome-enriched fractions achieved 100%/93% and 95%/79% specificity/sensitivity in distinguishing cancer patients from healthy individuals, respectively, demonstrating the diagnostic value of urine extracellular vesicles. (hide) EV-METRIC 25% (56th 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 Urine 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 Protein markers EV: CD9/ CD63/ CD24 non-EV: None Proteomics no Show all info Study aim Biomarker Sample Species Homo sapiens Sample Type Urine Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Below or equal to 800 g Between 10,000 g and 50,000 g Between 100,000 g and 150,000 g Pelleting performed Yes Pelleting: time(min) 90 Pelleting: rotor type Not specified Pelleting: speed (g) 100000 Wash: volume per pellet (ml) 10 Wash: time (min) 90 Wash: Rotor Type Not specified Wash: speed (g) 100000 Filtration steps 0.2µm > x > 0.1µm Characterization: Protein analysis None Protein Concentration Method Fluorometric assay (e.g. Qubit, NanoOrange,...) Characterization: RNA analysis RNA analysis Type (RT)(q)PCR Database No Proteinase treatment No RNAse treatment No Characterization: Lipid analysis No EM EM-type Immuno-EM/ Transmission-EM EM protein Other;CD9;CD63;CD24 Image type Close-up, Wide-field Report size (nm) 20-230

	EV200193	4/4	Homo sapiens	Urine	(d)(U)C Filtration	Bryzgunova, Olga E	2016	25%
Study summary Full title Comparative Study of Extracellular Vesicles from the Urine of Healthy Individuals and Prostate Cancer Patients All authors Olga E Bryzgunova, Marat M Zaripov, Tatyana E Skvortsova, Evgeny A Lekchnov, Alina E Grigor'eva, Ivan A Zaporozhchenko, Evgeny S Morozkin, Elena I Ryabchikova, Yuri B Yurchenko, Vladimir E Voitsitskiy, Pavel P Laktionov Journal PLoS One Abstract Recent studies suggest that extracellular vesicles may be the key to timely diagnosis and monitoring (show more...)Recent studies suggest that extracellular vesicles may be the key to timely diagnosis and monitoring of genito-urological malignancies. In this study we investigated the composition and content of extracellular vesicles found in the urine of healthy donors and prostate cancer patients. Urine of 14 PCa patients and 20 healthy volunteers was clarified by low-speed centrifugation and total extracellular vesicles fraction was obtain by high-speed centrifugation. The exosome-enriched fraction was obtained by filtration of total extracellular vesicles through a 0.1 μm pore filter. Transmission electron microscopy showed that cell-free urine in both groups contained vesicles from 20 to 230 nm. Immunogold staining after ultrafiltration demonstrated that 95% and 90% of extracellular vesicles in healthy individuals and cancer patients, respectively, were exosomes. Protein, DNA and RNA concentrations as well as size distribution of extracellular vesicles in both fractions were analyzed. Only 75% of the total protein content of extracellular vesicles was associated with exosomes which amounted to 90-95% of all vesicles. Median DNA concentrations in total extracellular vesicles and exosome-enriched fractions were 18 pg/ml and 2.6 pg/ml urine, correspondingly. Urine extracellular vesicles carried a population of RNA molecules 25 nt to 200 nt in concentration of no more than 290 pg/ml of urine. Additionally, concentrations of miR-19b, miR-25, miR-125b, and miR-205 were quantified by qRT-PCR. MiRNAs were shown to be differently distributed between different fractions of extracellular vesicles. Detection of miR-19b versus miR-16 in total vesicles and exosome-enriched fractions achieved 100%/93% and 95%/79% specificity/sensitivity in distinguishing cancer patients from healthy individuals, respectively, demonstrating the diagnostic value of urine extracellular vesicles. (hide) EV-METRIC 25% (56th 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 Urine 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/ CD24 non-EV: None Proteomics no Show all info Study aim Biomarker Sample Species Homo sapiens Sample Type Urine Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Below or equal to 800 g Between 10,000 g and 50,000 g Between 100,000 g and 150,000 g Pelleting performed Yes Pelleting: time(min) 90 Pelleting: rotor type Not specified Pelleting: speed (g) 100000 Wash: volume per pellet (ml) 10 Wash: time (min) 90 Wash: Rotor Type Not specified Wash: speed (g) 100000 Filtration steps 0.2µm > x > 0.1µm EV-subtype Distinction between multiple subtypes Size Used subtypes 30–100 nm enriched Characterization: Protein analysis None Protein Concentration Method Fluorometric assay (e.g. Qubit, NanoOrange,...) Characterization: RNA analysis RNA analysis Type (RT)(q)PCR;Capillary electrophoresis (e.g. Bioanalyzer) Database No Proteinase treatment No RNAse treatment No Characterization: Lipid analysis No EM EM-type Immuno-EM/ Transmission-EM EM protein Other;CD9;CD63;CD24 Image type Close-up, Wide-field Report size (nm) 30-100

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EV-TRACK ID	EV200193
species	Homo sapiens
sample type	Urine
condition	Prostate cancer	Prostate cancer	Control condition	Control condition
separation protocol	dUC	dUC Filtration	dUC	dUC Filtration

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