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Details	EV-TRACK ID	Experiment nr.	Species	Sample type	Separation protocol	First author	Year	EV-METRIC
	EV210360	1/2	Homo sapiens	Urine	DG (d)(U)C	Abe H	2018	33%
Study summary Full title Urinary Exosomal mRNA of WT1 as Diagnostic and Prognostic Biomarker for Diabetic Nephropathy. All authors Abe H, Sakurai A, Ono H, Hayashi S, Yoshimoto S, Ochi A, Ueda S, Nishimura K, Shibata E, Tamaki M, Kishi F, Kishi S, Murakami T, Nagai K, Doi T Journal J Med Invest Abstract Diabetic nephropathy (DN) is the major cause of end-stage renal failure and is associated with incre (show more...)Diabetic nephropathy (DN) is the major cause of end-stage renal failure and is associated with increased morbidity and mortality as compared to other causes of renal disease. Albuminuria is often the first clinical indicator of the presence of DN. However, albuminuria or proteinuria is a common symptom in patients with various renal disorders. Therefore, specific biomarkers for the diagnosis of DN are required. A primary hallmark of DN is the progressive damage and death of glomerular podocytes, resulting in the leaking of proteins into the urine. Urinary exosomes released by podocytes are microvesicles containing information of the originated cells. Podocyte-derived signal transduction factors (PDSTFs) are good candidates to assess podocyte injuries. The profile of PDSTFs in urinary exosomes from patients with DN is different from that from patients with minimal change nehrotic syndrome. In addition, PDSTFs molecules in exosomes were derived from primary murine podocytes under high glucose conditions. Among PDSTFs in urinary exosomes, Wilms tumor 1 (WT1) levels reflected damage of diabetic glomeruli in the patients. Urinary exosomal WT1 can predict the decline in eGFR for the following several years. In conclusion, urinary exosomal WT1 is a useful biomarker to improve risk stratification in patients with DN. J. Med. Invest. 65:208-215, August, 2018. (hide) EV-METRIC 33% (65th 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 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 Density gradient (Differential) (ultra)centrifugation Protein markers EV: None 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 Between 800 g and 10,000 g Between 100,000 g and 150,000 g Equal to or above 150,000 g Between 50,000 g and 100,000 g Pelleting performed Yes Pelleting: rotor type RP-65 Pelleting: speed (g) 70000 Density gradient Type Continuous Lowest density fraction 0.25 M Highest density fraction 2 M Total gradient volume, incl. sample (mL) 30 Sample volume (mL) 5 Orientation Top-down Rotor type RPS-28SA Speed (g) 100000 Duration (min) 1200 Fraction volume (mL) 2 Fraction processing Centrifugation Pelleting: volume per fraction 5 Pelleting: duration (min) 60 Pelleting: rotor type RPS-50-2 Pelleting: speed (g) 200000 Characterization: Protein analysis Protein Concentration Method Not determined Western Blot Antibody details provided? No Antibody dilution provided? Yes Detected EV-associated proteins CD63/ CD81 Characterization: RNA analysis RNA analysis Type (RT)(q)PCR Database No Proteinase treatment No RNAse treatment No Characterization: Lipid analysis No

	EV210360	2/2	Homo sapiens	Urine	DG (d)(U)C	Abe H	2018	33%
Study summary Full title Urinary Exosomal mRNA of WT1 as Diagnostic and Prognostic Biomarker for Diabetic Nephropathy. All authors Abe H, Sakurai A, Ono H, Hayashi S, Yoshimoto S, Ochi A, Ueda S, Nishimura K, Shibata E, Tamaki M, Kishi F, Kishi S, Murakami T, Nagai K, Doi T Journal J Med Invest Abstract Diabetic nephropathy (DN) is the major cause of end-stage renal failure and is associated with incre (show more...)Diabetic nephropathy (DN) is the major cause of end-stage renal failure and is associated with increased morbidity and mortality as compared to other causes of renal disease. Albuminuria is often the first clinical indicator of the presence of DN. However, albuminuria or proteinuria is a common symptom in patients with various renal disorders. Therefore, specific biomarkers for the diagnosis of DN are required. A primary hallmark of DN is the progressive damage and death of glomerular podocytes, resulting in the leaking of proteins into the urine. Urinary exosomes released by podocytes are microvesicles containing information of the originated cells. Podocyte-derived signal transduction factors (PDSTFs) are good candidates to assess podocyte injuries. The profile of PDSTFs in urinary exosomes from patients with DN is different from that from patients with minimal change nehrotic syndrome. In addition, PDSTFs molecules in exosomes were derived from primary murine podocytes under high glucose conditions. Among PDSTFs in urinary exosomes, Wilms tumor 1 (WT1) levels reflected damage of diabetic glomeruli in the patients. Urinary exosomal WT1 can predict the decline in eGFR for the following several years. In conclusion, urinary exosomal WT1 is a useful biomarker to improve risk stratification in patients with DN. J. Med. Invest. 65:208-215, August, 2018. (hide) EV-METRIC 33% (65th 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 Proteinuria patients 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 Density gradient (Differential) (ultra)centrifugation Protein markers EV: None 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 Between 800 g and 10,000 g Between 100,000 g and 150,000 g Equal to or above 150,000 g Between 50,000 g and 100,000 g Pelleting performed Yes Pelleting: rotor type RP-65 Pelleting: speed (g) 70000 Density gradient Type Continuous Lowest density fraction 0.25 M Highest density fraction 2 M Total gradient volume, incl. sample (mL) 30 Sample volume (mL) 5 Orientation Top-down Rotor type RPS-28SA Speed (g) 100000 Duration (min) 1200 Fraction volume (mL) 2 Fraction processing Centrifugation Pelleting: volume per fraction 5 Pelleting: duration (min) 60 Pelleting: rotor type RPS-50-2 Pelleting: speed (g) 200000 Characterization: Protein analysis Protein Concentration Method Not determined Western Blot Antibody details provided? No Antibody dilution provided? Yes Detected EV-associated proteins CD63/ CD81 Characterization: RNA analysis RNA analysis Type (RT)(q)PCR Database No Proteinase treatment No RNAse treatment No Characterization: Lipid analysis No

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EV-TRACK ID	EV210360
species	Homo sapiens
sample type	Urine
condition	Control condition	Proteinuria patients
separation protocol	Density gradient/ dUC	Density gradient/ dUC
Exp. nr.	1	2
EV-METRIC %	33	33