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Details	EV-TRACK ID	Experiment nr.	Species	Sample type	Separation protocol	First author	Year	EV-METRIC
	EV100025	1/2	Rattus norvegicus	BSp73AS	(d)(U)C DG	Nazarenko I	2010	44%
Study summary Full title Cell surface tetraspanin Tspan8 contributes to molecular pathways of exosome-induced endothelial cell activation. All authors Nazarenko I, Rana S, Baumann A, McAlear J, Hellwig A, Trendelenburg M, Lochnit G, Preissner KT, Zöller M Journal J Cell Sci Abstract Tumor-derived exosomes containing the tetraspanin Tspan8 can efficiently induce angiogenesis in tumo (show more...)Tumor-derived exosomes containing the tetraspanin Tspan8 can efficiently induce angiogenesis in tumors and tumor-free tissues. However, little information exists on exosome-endothelial cell (EC) interactions or the proangiogenic role of tetraspanins, which are a constitutive component of exosomes. In this study, we used a rat adenocarcinoma model (AS-Tspan8) to explore the effects of exosomal Tspan8 on angiogenesis. Tspan8 contributed to a selective recruitment of proteins and mRNA into exosomes, including CD106 and CD49d, which were implicated in exosome-EC binding and EC internalization. We found that EC internalized Tspan8-CD49d complex-containing exosomes. Exosome uptake induced vascular endothelial growth factor (VEGF)-independent regulation of several angiogenesis-related genes, including von Willebrand factor, Tspan8, chemokines CXCL5 and MIF, chemokine receptor CCR1, and, together with VEGF, VEGF receptor 2. EC uptake of Tspan8-CD49d complex-containing exosomes was accompanied by enhanced EC proliferation, migration, sprouting, and maturation of EC progenitors. Unraveling these new pathways of exosome-initiated EC regulation could provide new options for therapeutic interference with tumor-induced angiogenesis. (hide) EV-METRIC 44% (84th 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 Tspan8 overexpression 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 (d)(U)C DG Protein markers EV: TSG101/ CD49d/ Bag6/ CD71/ CD49c/ CD49b/ HSP90/ CD151/ Mac2BP/ LAMP1/ HSP70/ Tspan8/ CD61/ CD9/ CD106 non-EV: None Proteomics yes Show all info Study aim Function, Identification of content (omics approaches) Sample Species Rattus norvegicus Sample Type Cell culture supernatant EV-producing cells BSp73AS EV-harvesting Medium Serum free medium Cell viability (%) NA 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 Between 100,000 g and 150,000 g Pelleting performed Yes Pelleting: time(min) 90 Pelleting: speed (g) 100000 Wash: time (min) 90 Wash: speed (g) 100000 Density gradient Only used for validation of main results Yes Type Continuous Lowest density fraction 0.25 M Highest density fraction 2.0 M Sample volume (mL) 0.2 Orientation Bottom-up (sample migrates upwards) Rotor type SW 41 Ti Speed (g) 150000 Duration (min) 900 Fraction volume (mL) 1 Fraction processing Centrifugation Pelleting: volume per fraction 11 Pelleting: duration (min) 2 Pelleting: rotor type SW 41 Ti Pelleting: speed (g) 120000 Pelleting: adjusted k-factor 213.2 Characterization: Protein analysis Protein Concentration Method BCA Western Blot Detected EV-associated proteins CD9, CD71, HSP70, HSP90, TSG101, CD151, LAMP1, Tspan8, Bag6, CD49b, CD49c, CD49d, CD61, Mac2BP, CD106 Characterization: RNA analysis Proteinase treatment No RNAse treatment Yes Moment of RNAse treatment After RNAse type RNase A RNAse concentration 5 Characterization: Lipid analysis No Characterization: Particle analysis EM EM-type Transmission-EM Image type Close-up Extra information this is publication from 2010. To that time point no NTA, DLS or further technologies were established

	EV100025	2/2	Rattus norvegicus	BSp73AS	(d)(U)C DG	Nazarenko I	2010	44%
Study summary Full title Cell surface tetraspanin Tspan8 contributes to molecular pathways of exosome-induced endothelial cell activation. All authors Nazarenko I, Rana S, Baumann A, McAlear J, Hellwig A, Trendelenburg M, Lochnit G, Preissner KT, Zöller M Journal J Cell Sci Abstract Tumor-derived exosomes containing the tetraspanin Tspan8 can efficiently induce angiogenesis in tumo (show more...)Tumor-derived exosomes containing the tetraspanin Tspan8 can efficiently induce angiogenesis in tumors and tumor-free tissues. However, little information exists on exosome-endothelial cell (EC) interactions or the proangiogenic role of tetraspanins, which are a constitutive component of exosomes. In this study, we used a rat adenocarcinoma model (AS-Tspan8) to explore the effects of exosomal Tspan8 on angiogenesis. Tspan8 contributed to a selective recruitment of proteins and mRNA into exosomes, including CD106 and CD49d, which were implicated in exosome-EC binding and EC internalization. We found that EC internalized Tspan8-CD49d complex-containing exosomes. Exosome uptake induced vascular endothelial growth factor (VEGF)-independent regulation of several angiogenesis-related genes, including von Willebrand factor, Tspan8, chemokines CXCL5 and MIF, chemokine receptor CCR1, and, together with VEGF, VEGF receptor 2. EC uptake of Tspan8-CD49d complex-containing exosomes was accompanied by enhanced EC proliferation, migration, sprouting, and maturation of EC progenitors. Unraveling these new pathways of exosome-initiated EC regulation could provide new options for therapeutic interference with tumor-induced angiogenesis. (hide) EV-METRIC 44% (84th 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 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 (d)(U)C DG Protein markers EV: CD49c/ CD61/ CD49b/ CD151/ Mac2BP/ HSP70/ CD9/ CD106 non-EV: None Proteomics yes Show all info Study aim Function, Identification of content (omics approaches) Sample Species Rattus norvegicus Sample Type Cell culture supernatant EV-producing cells BSp73AS EV-harvesting Medium Serum free medium Cell viability (%) NA 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 Between 100,000 g and 150,000 g Pelleting performed Yes Pelleting: time(min) 90 Pelleting: speed (g) 100000 Wash: time (min) 90 Wash: speed (g) 100000 Density gradient Only used for validation of main results Yes Type Continuous Lowest density fraction 0.25 M Highest density fraction 2.0 M Sample volume (mL) 0.2 Orientation Bottom-up (sample migrates upwards) Rotor type SW 41 Ti Speed (g) 150000 Duration (min) 900 Fraction volume (mL) 1 Fraction processing Centrifugation Pelleting: volume per fraction 11 Pelleting: duration (min) 2 Pelleting: rotor type SW 41 Ti Pelleting: speed (g) 120000 Pelleting: adjusted k-factor 213.2 Characterization: Protein analysis Protein Concentration Method BCA Western Blot Detected EV-associated proteins CD151, CD9, CD49b, CD49c, CD61, Mac2BP, CD106, HSP70 Characterization: RNA analysis Proteinase treatment No RNAse treatment Yes Moment of RNAse treatment After RNAse type RNase A RNAse concentration 5 Characterization: Lipid analysis No Characterization: Particle analysis None Extra information this is publication from 2010. To that time point no NTA, DLS or further technologies were established

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EV-TRACK ID	EV100025
species	Rattus norvegicus
sample type	Cell culture
cell type	BSp73AS
condition	Tspan8 overexpression	Control condition
separation protocol	(d)(U)C DG	(d)(U)C DG
Exp. nr.	1	2
EV-METRIC %	44	44