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Details	EV-TRACK ID	Experiment nr.	Species	Sample type	Separation protocol	First author	Year	EV-METRIC
	EV210366	1/6	Homo sapiens	Primary Lymphatic Endothelial Cells	(d)(U)C ExoQuick Filtration	Brown M	2018	50%
Study summary Full title Lymphatic exosomes promote dendritic cell migration along guidance cues. All authors Brown M, Johnson LA, Leone DA, Majek P, Vaahtomeri K, Senfter D, Bukosza N, Schachner H, Asfour G, Langer B, Hauschild R, Parapatics K, Hong YK, Bennett KL, Kain R, Detmar M, Sixt M, Jackson DG, Kerjaschki D Journal J Cell Biol Abstract Lymphatic endothelial cells (LECs) release extracellular chemokines to guide the migration of dendri (show more...)Lymphatic endothelial cells (LECs) release extracellular chemokines to guide the migration of dendritic cells. In this study, we report that LECs also release basolateral exosome-rich endothelial vesicles (EEVs) that are secreted in greater numbers in the presence of inflammatory cytokines and accumulate in the perivascular stroma of small lymphatic vessels in human chronic inflammatory diseases. Proteomic analyses of EEV fractions identified >1,700 cargo proteins and revealed a dominant motility-promoting protein signature. In vitro and ex vivo EEV fractions augmented cellular protrusion formation in a CX3CL1/fractalkine-dependent fashion and enhanced the directional migratory response of human dendritic cells along guidance cues. We conclude that perilymphatic LEC exosomes enhance exploratory behavior and thus promote directional migration of CX3CR1-expressing cells in complex tissue environments. (hide) EV-METRIC 50% (87th 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-rich endothelial vesicles 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 Commercial method Filtration Protein markers EV: CX3CL1/ CD9 non-EV: None Proteomics yes Show all info Study aim Function/Identification of content (omics approaches) Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells Primary Lymphatic Endothelial Cells EV-harvesting Medium EV-depleted medium Preparation of EDS Commercial EDS Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Pelleting performed No Filtration steps 0.22µm or 0.2µm Commercial kit ExoQuick Characterization: Protein analysis Protein Concentration Method BCA Western Blot Antibody details provided? No Lysis buffer provided? Yes Detected EV-associated proteins CD9/ CX3CL1 Proteomics database No Characterization: Lipid analysis No Characterization: Particle analysis NTA Report type Size range/distribution Reported size (nm) 25 / 75 / 125 / 175 / 225 / 275 EV concentration Yes EM EM-type Transmission-EM Image type Close-up Report size (nm) 133

	EV210366	3/6	Homo sapiens	Primary Lymphatic Endothelial Cells	(d)(U)C ExoQuick Filtration	Brown M	2018	50%
Study summary Full title Lymphatic exosomes promote dendritic cell migration along guidance cues. All authors Brown M, Johnson LA, Leone DA, Majek P, Vaahtomeri K, Senfter D, Bukosza N, Schachner H, Asfour G, Langer B, Hauschild R, Parapatics K, Hong YK, Bennett KL, Kain R, Detmar M, Sixt M, Jackson DG, Kerjaschki D Journal J Cell Biol Abstract Lymphatic endothelial cells (LECs) release extracellular chemokines to guide the migration of dendri (show more...)Lymphatic endothelial cells (LECs) release extracellular chemokines to guide the migration of dendritic cells. In this study, we report that LECs also release basolateral exosome-rich endothelial vesicles (EEVs) that are secreted in greater numbers in the presence of inflammatory cytokines and accumulate in the perivascular stroma of small lymphatic vessels in human chronic inflammatory diseases. Proteomic analyses of EEV fractions identified >1,700 cargo proteins and revealed a dominant motility-promoting protein signature. In vitro and ex vivo EEV fractions augmented cellular protrusion formation in a CX3CL1/fractalkine-dependent fashion and enhanced the directional migratory response of human dendritic cells along guidance cues. We conclude that perilymphatic LEC exosomes enhance exploratory behavior and thus promote directional migration of CX3CR1-expressing cells in complex tissue environments. (hide) EV-METRIC 50% (87th 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 TNF- containing EV harvesting media (7ng/mL) Focus vesicles Exosome-rich endothelial vesicles 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 Commercial method Filtration Protein markers EV: CX3CL1/ CD9 non-EV: None Proteomics yes Show all info Study aim Function/Identification of content (omics approaches) Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells Primary Lymphatic Endothelial Cells EV-harvesting Medium EV-depleted medium Preparation of EDS Commercial EDS Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Pelleting performed No Filtration steps 0.22µm or 0.2µm Commercial kit ExoQuick Characterization: Protein analysis Protein Concentration Method BCA Western Blot Antibody details provided? No Lysis buffer provided? Yes Detected EV-associated proteins CD9/ CX3CL1 Proteomics database No Characterization: Lipid analysis No Characterization: Particle analysis NTA Report type Size range/distribution Reported size (nm) 25 / 75 / 125 / 175 / 225 / 275 EV concentration Yes EM EM-type Transmission-EM Image type Close-up Report size (nm) 83

	EV210366	2/6	Homo sapiens	Primary Lymphatic Endothelial Cells	(d)(U)C ExoQuick Filtration DG	Brown M	2018	37%
Study summary Full title Lymphatic exosomes promote dendritic cell migration along guidance cues. All authors Brown M, Johnson LA, Leone DA, Majek P, Vaahtomeri K, Senfter D, Bukosza N, Schachner H, Asfour G, Langer B, Hauschild R, Parapatics K, Hong YK, Bennett KL, Kain R, Detmar M, Sixt M, Jackson DG, Kerjaschki D Journal J Cell Biol Abstract Lymphatic endothelial cells (LECs) release extracellular chemokines to guide the migration of dendri (show more...)Lymphatic endothelial cells (LECs) release extracellular chemokines to guide the migration of dendritic cells. In this study, we report that LECs also release basolateral exosome-rich endothelial vesicles (EEVs) that are secreted in greater numbers in the presence of inflammatory cytokines and accumulate in the perivascular stroma of small lymphatic vessels in human chronic inflammatory diseases. Proteomic analyses of EEV fractions identified >1,700 cargo proteins and revealed a dominant motility-promoting protein signature. In vitro and ex vivo EEV fractions augmented cellular protrusion formation in a CX3CL1/fractalkine-dependent fashion and enhanced the directional migratory response of human dendritic cells along guidance cues. We conclude that perilymphatic LEC exosomes enhance exploratory behavior and thus promote directional migration of CX3CR1-expressing cells in complex tissue environments. (hide) EV-METRIC 37% (79th 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-rich endothelial vesicles 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 Commercial method Filtration Density gradient Protein markers EV: CD63 non-EV: None Proteomics no Show all info Study aim Function/Identification of content (omics approaches) Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells Primary Lymphatic Endothelial Cells EV-harvesting Medium EV-depleted medium Preparation of EDS Commercial EDS Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Pelleting performed No Density gradient Only used for validation of main results Yes Type Discontinuous Number of initial discontinuous layers 4 Lowest density fraction 5% Highest density fraction 40% Total gradient volume, incl. sample (mL) 12 Sample volume (mL) 1 Orientation Top-down Rotor type P40ST (Hitachi) Speed (g) 100000 Duration (min) 960 Fraction volume (mL) 1 Filtration steps 0.22µm or 0.2µm Commercial kit ExoQuick Characterization: Protein analysis Protein Concentration Method Not determined Western Blot Antibody details provided? No Lysis buffer provided? Yes Detected EV-associated proteins CD63 Characterization: Lipid analysis No Characterization: Particle analysis None

	EV210366	4/6	Homo sapiens	Primary Lymphatic Endothelial Cells	(d)(U)C ExoQuick Filtration DG	Brown M	2018	37%
Study summary Full title Lymphatic exosomes promote dendritic cell migration along guidance cues. All authors Brown M, Johnson LA, Leone DA, Majek P, Vaahtomeri K, Senfter D, Bukosza N, Schachner H, Asfour G, Langer B, Hauschild R, Parapatics K, Hong YK, Bennett KL, Kain R, Detmar M, Sixt M, Jackson DG, Kerjaschki D Journal J Cell Biol Abstract Lymphatic endothelial cells (LECs) release extracellular chemokines to guide the migration of dendri (show more...)Lymphatic endothelial cells (LECs) release extracellular chemokines to guide the migration of dendritic cells. In this study, we report that LECs also release basolateral exosome-rich endothelial vesicles (EEVs) that are secreted in greater numbers in the presence of inflammatory cytokines and accumulate in the perivascular stroma of small lymphatic vessels in human chronic inflammatory diseases. Proteomic analyses of EEV fractions identified >1,700 cargo proteins and revealed a dominant motility-promoting protein signature. In vitro and ex vivo EEV fractions augmented cellular protrusion formation in a CX3CL1/fractalkine-dependent fashion and enhanced the directional migratory response of human dendritic cells along guidance cues. We conclude that perilymphatic LEC exosomes enhance exploratory behavior and thus promote directional migration of CX3CR1-expressing cells in complex tissue environments. (hide) EV-METRIC 37% (79th 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 TNF- containing EV harvesting media (7ng/mL) Focus vesicles Exosome-rich endothelial vesicles 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 Commercial method Filtration Density gradient Protein markers EV: CD63 non-EV: None Proteomics no Show all info Study aim Function/Identification of content (omics approaches) Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells Primary Lymphatic Endothelial Cells EV-harvesting Medium EV-depleted medium Preparation of EDS Commercial EDS Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Pelleting performed No Density gradient Only used for validation of main results Yes Type Discontinuous Number of initial discontinuous layers 4 Lowest density fraction 5% Highest density fraction 40% Total gradient volume, incl. sample (mL) 12 Sample volume (mL) 1 Orientation Top-down Rotor type P40ST (Hitachi) Speed (g) 100000 Duration (min) 960 Fraction volume (mL) 1 Filtration steps 0.22µm or 0.2µm Commercial kit ExoQuick Characterization: Protein analysis Protein Concentration Method Not determined Western Blot Antibody details provided? No Lysis buffer provided? Yes Detected EV-associated proteins CD63 Characterization: Lipid analysis No Characterization: Particle analysis None

	EV210366	5/6	Homo sapiens	Primary Lymphatic Endothelial Cells	(d)(U)C ExoQuick Filtration No Extra Isolation steps	Brown M	2018	0%
Study summary Full title Lymphatic exosomes promote dendritic cell migration along guidance cues. All authors Brown M, Johnson LA, Leone DA, Majek P, Vaahtomeri K, Senfter D, Bukosza N, Schachner H, Asfour G, Langer B, Hauschild R, Parapatics K, Hong YK, Bennett KL, Kain R, Detmar M, Sixt M, Jackson DG, Kerjaschki D Journal J Cell Biol Abstract Lymphatic endothelial cells (LECs) release extracellular chemokines to guide the migration of dendri (show more...)Lymphatic endothelial cells (LECs) release extracellular chemokines to guide the migration of dendritic cells. In this study, we report that LECs also release basolateral exosome-rich endothelial vesicles (EEVs) that are secreted in greater numbers in the presence of inflammatory cytokines and accumulate in the perivascular stroma of small lymphatic vessels in human chronic inflammatory diseases. Proteomic analyses of EEV fractions identified >1,700 cargo proteins and revealed a dominant motility-promoting protein signature. In vitro and ex vivo EEV fractions augmented cellular protrusion formation in a CX3CL1/fractalkine-dependent fashion and enhanced the directional migratory response of human dendritic cells along guidance cues. We conclude that perilymphatic LEC exosomes enhance exploratory behavior and thus promote directional migration of CX3CR1-expressing cells in complex tissue environments. (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. 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 GW4869-containing EV harvesting media (10M) Focus vesicles Exosome-rich endothelial vesicles 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 Commercial method Filtration No Extra Isolation steps Protein markers EV: CD63 non-EV: None Proteomics no Show all info Study aim Function/Identification of content (omics approaches) Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells Primary Lymphatic Endothelial Cells EV-harvesting Medium EV-depleted medium Preparation of EDS Commercial EDS Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Pelleting performed No Filtration steps 0.22µm or 0.2µm Commercial kit ExoQuick Other Name other separation method No Extra Isolation steps Characterization: Protein analysis None Protein Concentration Method BCA Characterization: Lipid analysis No Characterization: Particle analysis None

	EV210366	6/6	Homo sapiens	Primary Lymphatic Endothelial Cells	(d)(U)C ExoQuick Filtration	Brown M	2018	0%
Study summary Full title Lymphatic exosomes promote dendritic cell migration along guidance cues. All authors Brown M, Johnson LA, Leone DA, Majek P, Vaahtomeri K, Senfter D, Bukosza N, Schachner H, Asfour G, Langer B, Hauschild R, Parapatics K, Hong YK, Bennett KL, Kain R, Detmar M, Sixt M, Jackson DG, Kerjaschki D Journal J Cell Biol Abstract Lymphatic endothelial cells (LECs) release extracellular chemokines to guide the migration of dendri (show more...)Lymphatic endothelial cells (LECs) release extracellular chemokines to guide the migration of dendritic cells. In this study, we report that LECs also release basolateral exosome-rich endothelial vesicles (EEVs) that are secreted in greater numbers in the presence of inflammatory cytokines and accumulate in the perivascular stroma of small lymphatic vessels in human chronic inflammatory diseases. Proteomic analyses of EEV fractions identified >1,700 cargo proteins and revealed a dominant motility-promoting protein signature. In vitro and ex vivo EEV fractions augmented cellular protrusion formation in a CX3CL1/fractalkine-dependent fashion and enhanced the directional migratory response of human dendritic cells along guidance cues. We conclude that perilymphatic LEC exosomes enhance exploratory behavior and thus promote directional migration of CX3CR1-expressing cells in complex tissue environments. (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. 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 GW4869 (10M) and TNF- (7ng/mL) containing EV harvesting media Focus vesicles Exosome-rich endothelial vesicles 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 Commercial method Filtration Protein markers EV: CD63 non-EV: None Proteomics no Show all info Study aim Function/Identification of content (omics approaches) Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells Primary Lymphatic Endothelial Cells EV-harvesting Medium EV-depleted medium Preparation of EDS Commercial EDS Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Pelleting performed No Filtration steps 0.22µm or 0.2µm Commercial kit ExoQuick Characterization: Protein analysis None Protein Concentration Method BCA Characterization: Lipid analysis No Characterization: Particle analysis None

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EV-TRACK ID	EV210366
species	Homo sapiens
sample type	Cell culture
cell type	Primary Lymphatic Endothelial Cells
condition	Control condition	TNF- containing EV harvesting media (7ng/mL)	Control condition	TNF- containing EV harvesting media (7ng/mL)	GW4869-containing EV harvesting media (10M)	GW4869 (10M) and TNF- (7ng/mL) containing EV harvesting media
separation protocol	dUC/ ExoQuick/ Filtration	dUC/ ExoQuick/ Filtration	dUC/ ExoQuick/ Filtration/ Density gradient	dUC/ ExoQuick/ Filtration/ Density gradient	dUC/ ExoQuick/ Filtration/ No Extra Isolation steps	dUC/ ExoQuick/ Filtration
Exp. nr.	1	3	2	4	5	6
EV-METRIC %	50	50	37	37	0	0