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Details	EV-TRACK ID	Experiment nr.	Species	Sample type	Separation protocol	First author	Year	EV-METRIC
	EV210281	3/16	Homo sapiens	MDAMB231	(d)(U)C UF qEV Immunoaffinity capture	Grisard, Eleonora	2021	75%
Study summary Full title Homosalate boosts the release of tumor-derived Extracellular Vesicles with anti-anoikis properties All authors Eleonora Grisard, Aurianne Lescure, Nathalie Nevo, Maxime Corbé, Mabel Jouve, Gregory Lavieu, Alain Joliot, Elaine Del Nery, Lorena Martin-Jaular, Clotilde Théry Journal bioRxiv Abstract Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular (show more...)Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular vesicles (EVs). EVs could have different subcellular origin, composition and functional properties, but tools to distinguish between EV subtypes are scarce. Here, we tagged CD63-or CD9-positive EVs secreted by triple negative breast cancer cells with Nanoluciferase enzyme, to set-up a miniaturized method to quantify secretion of these two EV subtypes directly in the supernatant of cells. We performed a cell-based high-content screening to identify clinically-approved drugs able to affect EV secretion. One of the identified hits is Homosalate, an anti-inflammatory drug found in sunscreens which robustly increased EVs’release. Comparing EVs induced by Homosalate with those induced by Bafilomycin A1, we discovered that: 1) the two drugs act on EVs generated indistinct subcellular compartmentsand 2) EVs released upon treatment with Homosalate, but not with Bafilomycin A1, conferred anti-anoikis properties to another recipient tumor cell line. In conclusion, we identified a new drug modifying EV release and demonstrated that under influence of different drugs, triple negative breast cancer cells release EV subpopulations from different subcellular origins harboring distinct functional properties. (hide) EV-METRIC 75% (96th 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 DMSO treated 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 Ultrafiltration qEV Immunoaffinity capture Protein markers EV: CD9/ CD63/ CD81/ syntenin/ CD98 non-EV: Gapdh Proteomics no Show all info Study aim Function/Biogenesis/cargo sorting Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells MDAMB231 EV-harvesting Medium Serum free medium Cell viability (%) 80 Cell count 27000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Pelleting performed No Ultra filtration Cut-off size (kDa) 10 kDa Membrane type PES Commercial kit qEV Immunoaffinity capture Selected surface protein(s) CD9/ CD63 Other Name other separation method qEV Other Name other separation method Immunoaffinity capture Characterization: Protein analysis Protein Concentration Method Not determined Western Blot Antibody details provided? Yes Antibody dilution provided? Yes Lysis buffer provided? Yes Detected EV-associated proteins CD9/ CD63/ CD81/ syntenin/ CD98 Detected contaminants Gapdh Characterization: Lipid analysis No Characterization: Particle analysis NTA Report type Size range/distribution Reported size (nm) 100 EV concentration Yes Particle yield number of particles per million cells: 5.00e+9 EM EM-type Transmission-EM Image type Wide-field Report size (nm) 125

	EV210281	4/16	Homo sapiens	MDAMB231	(d)(U)C UF qEV Immunoaffinity capture	Grisard, Eleonora	2021	75%
Study summary Full title Homosalate boosts the release of tumor-derived Extracellular Vesicles with anti-anoikis properties All authors Eleonora Grisard, Aurianne Lescure, Nathalie Nevo, Maxime Corbé, Mabel Jouve, Gregory Lavieu, Alain Joliot, Elaine Del Nery, Lorena Martin-Jaular, Clotilde Théry Journal bioRxiv Abstract Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular (show more...)Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular vesicles (EVs). EVs could have different subcellular origin, composition and functional properties, but tools to distinguish between EV subtypes are scarce. Here, we tagged CD63-or CD9-positive EVs secreted by triple negative breast cancer cells with Nanoluciferase enzyme, to set-up a miniaturized method to quantify secretion of these two EV subtypes directly in the supernatant of cells. We performed a cell-based high-content screening to identify clinically-approved drugs able to affect EV secretion. One of the identified hits is Homosalate, an anti-inflammatory drug found in sunscreens which robustly increased EVs’release. Comparing EVs induced by Homosalate with those induced by Bafilomycin A1, we discovered that: 1) the two drugs act on EVs generated indistinct subcellular compartmentsand 2) EVs released upon treatment with Homosalate, but not with Bafilomycin A1, conferred anti-anoikis properties to another recipient tumor cell line. In conclusion, we identified a new drug modifying EV release and demonstrated that under influence of different drugs, triple negative breast cancer cells release EV subpopulations from different subcellular origins harboring distinct functional properties. (hide) EV-METRIC 75% (96th 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 Homosalate treated 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 Ultrafiltration qEV Immunoaffinity capture Protein markers EV: CD9/ CD63/ CD81/ syntenin/ CD98 non-EV: Gapdh Proteomics no Show all info Study aim Function/Biogenesis/cargo sorting Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells MDAMB231 EV-harvesting Medium Serum free medium Cell viability (%) 80 Cell count 27000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Pelleting performed No Ultra filtration Cut-off size (kDa) 10 kDa Membrane type PES Commercial kit qEV Immunoaffinity capture Selected surface protein(s) CD9/ CD63 Other Name other separation method qEV Other Name other separation method Immunoaffinity capture Characterization: Protein analysis Protein Concentration Method Not determined Western Blot Antibody details provided? Yes Antibody dilution provided? Yes Lysis buffer provided? Yes Detected EV-associated proteins CD9/ CD63/ CD81/ syntenin/ CD98 Detected contaminants Gapdh Characterization: Lipid analysis No Characterization: Particle analysis NTA Report type Size range/distribution Reported size (nm) 100 EV concentration Yes Particle yield number of particles per million cells: 5.00e+9 EM EM-type Transmission-EM Image type Wide-field Report size (nm) 125

	EV210281	1/16	Homo sapiens	MDAMB231 Nanoluc-CD63	(d)(U)C UF qEV	Grisard, Eleonora	2021	50%
Study summary Full title Homosalate boosts the release of tumor-derived Extracellular Vesicles with anti-anoikis properties All authors Eleonora Grisard, Aurianne Lescure, Nathalie Nevo, Maxime Corbé, Mabel Jouve, Gregory Lavieu, Alain Joliot, Elaine Del Nery, Lorena Martin-Jaular, Clotilde Théry Journal bioRxiv Abstract Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular (show more...)Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular vesicles (EVs). EVs could have different subcellular origin, composition and functional properties, but tools to distinguish between EV subtypes are scarce. Here, we tagged CD63-or CD9-positive EVs secreted by triple negative breast cancer cells with Nanoluciferase enzyme, to set-up a miniaturized method to quantify secretion of these two EV subtypes directly in the supernatant of cells. We performed a cell-based high-content screening to identify clinically-approved drugs able to affect EV secretion. One of the identified hits is Homosalate, an anti-inflammatory drug found in sunscreens which robustly increased EVs’release. Comparing EVs induced by Homosalate with those induced by Bafilomycin A1, we discovered that: 1) the two drugs act on EVs generated indistinct subcellular compartmentsand 2) EVs released upon treatment with Homosalate, but not with Bafilomycin A1, conferred anti-anoikis properties to another recipient tumor cell line. In conclusion, we identified a new drug modifying EV release and demonstrated that under influence of different drugs, triple negative breast cancer cells release EV subpopulations from different subcellular origins harboring distinct functional properties. (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 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 Ultrafiltration qEV Protein markers EV: CD9/ CD63/ syntenin/ CD98/ 14-3-3 non-EV: None Proteomics no Show all info Study aim Function/Biogenesis/cargo sorting Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells MDAMB231 Nanoluc-CD63 EV-harvesting Medium Serum free medium Cell count 70000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Pelleting performed No Ultra filtration Cut-off size (kDa) 10 kDa Membrane type PES Commercial kit qEV Other Name other separation method qEV Characterization: Protein analysis Protein Concentration Method BCA Protein Yield (µg) measured in all qEV fractions F7-24 peaked at around 300ug in F21 Western Blot Antibody details provided? Yes Antibody dilution provided? Yes Lysis buffer provided? Yes Detected EV-associated proteins CD9/ CD63/ syntenin/ CD98 Not detected EV-associated proteins 14-3-3 Characterization: Lipid analysis No Characterization: Particle analysis NTA EV concentration Yes Particle yield number of particles per million cells: 2.00e+9 EM EM-type Transmission-EM Image type Wide-field Report size (nm) 125

	EV210281	2/16	Homo sapiens	MDAMB231 Nluc-CD9	(d)(U)C UF qEV	Grisard, Eleonora	2021	50%
Study summary Full title Homosalate boosts the release of tumor-derived Extracellular Vesicles with anti-anoikis properties All authors Eleonora Grisard, Aurianne Lescure, Nathalie Nevo, Maxime Corbé, Mabel Jouve, Gregory Lavieu, Alain Joliot, Elaine Del Nery, Lorena Martin-Jaular, Clotilde Théry Journal bioRxiv Abstract Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular (show more...)Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular vesicles (EVs). EVs could have different subcellular origin, composition and functional properties, but tools to distinguish between EV subtypes are scarce. Here, we tagged CD63-or CD9-positive EVs secreted by triple negative breast cancer cells with Nanoluciferase enzyme, to set-up a miniaturized method to quantify secretion of these two EV subtypes directly in the supernatant of cells. We performed a cell-based high-content screening to identify clinically-approved drugs able to affect EV secretion. One of the identified hits is Homosalate, an anti-inflammatory drug found in sunscreens which robustly increased EVs’release. Comparing EVs induced by Homosalate with those induced by Bafilomycin A1, we discovered that: 1) the two drugs act on EVs generated indistinct subcellular compartmentsand 2) EVs released upon treatment with Homosalate, but not with Bafilomycin A1, conferred anti-anoikis properties to another recipient tumor cell line. In conclusion, we identified a new drug modifying EV release and demonstrated that under influence of different drugs, triple negative breast cancer cells release EV subpopulations from different subcellular origins harboring distinct functional properties. (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 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 Ultrafiltration qEV Protein markers EV: CD9/ CD63/ syntenin/ CD98/ 14-3-3 non-EV: None Proteomics no Show all info Study aim Function/Biogenesis/cargo sorting Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells MDAMB231 Nluc-CD9 EV-harvesting Medium Serum free medium Cell count 70000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Pelleting performed No Ultra filtration Cut-off size (kDa) 10 kDa Membrane type PES Commercial kit qEV Other Name other separation method qEV Characterization: Protein analysis Protein Concentration Method Not determined Protein Yield (µg) measured in all qEV fractions F7-24 peaked at around 300ug in F19-21 Western Blot Antibody details provided? Yes Antibody dilution provided? Yes Lysis buffer provided? Yes Detected EV-associated proteins CD9/ CD63/ syntenin/ CD98 Not detected EV-associated proteins 14-3-3 Characterization: Lipid analysis No Characterization: Particle analysis NTA EV concentration Yes Particle yield number of particles per million cells: 2.00e+9 EM EM-type Transmission-EM Image type Wide-field Report size (nm) 125

	EV210281	5/16	Homo sapiens	MDAMB231	(d)(U)C UF qEV	Grisard, Eleonora	2021	38%
Study summary Full title Homosalate boosts the release of tumor-derived Extracellular Vesicles with anti-anoikis properties All authors Eleonora Grisard, Aurianne Lescure, Nathalie Nevo, Maxime Corbé, Mabel Jouve, Gregory Lavieu, Alain Joliot, Elaine Del Nery, Lorena Martin-Jaular, Clotilde Théry Journal bioRxiv Abstract Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular (show more...)Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular vesicles (EVs). EVs could have different subcellular origin, composition and functional properties, but tools to distinguish between EV subtypes are scarce. Here, we tagged CD63-or CD9-positive EVs secreted by triple negative breast cancer cells with Nanoluciferase enzyme, to set-up a miniaturized method to quantify secretion of these two EV subtypes directly in the supernatant of cells. We performed a cell-based high-content screening to identify clinically-approved drugs able to affect EV secretion. One of the identified hits is Homosalate, an anti-inflammatory drug found in sunscreens which robustly increased EVs’release. Comparing EVs induced by Homosalate with those induced by Bafilomycin A1, we discovered that: 1) the two drugs act on EVs generated indistinct subcellular compartmentsand 2) EVs released upon treatment with Homosalate, but not with Bafilomycin A1, conferred anti-anoikis properties to another recipient tumor cell line. In conclusion, we identified a new drug modifying EV release and demonstrated that under influence of different drugs, triple negative breast cancer cells release EV subpopulations from different subcellular origins harboring distinct functional properties. (hide) EV-METRIC 38% (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 DMSO treated 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 Ultrafiltration qEV Protein markers EV: CD9/ CD63/ CD81/ Lamp1/ syntenin/ CD98 non-EV: None Proteomics no Show all info Study aim Function/Biogenesis/cargo sorting Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells MDAMB231 EV-harvesting Medium Serum free medium Cell viability (%) 80 Cell count 27000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Pelleting performed No Ultra filtration Cut-off size (kDa) 10 kDa Membrane type PES Commercial kit qEV Other Name other separation method qEV Characterization: Protein analysis Protein Concentration Method Not determined Western Blot Antibody details provided? Yes Antibody dilution provided? Yes Lysis buffer provided? Yes Detected EV-associated proteins CD9/ CD63/ CD81/ Lamp1/ syntenin/ CD98 Characterization: Lipid analysis No Characterization: Particle analysis NTA Report type Size range/distribution Reported size (nm) 100 EV concentration Yes Particle yield number of particles per million cells: 5.00e+9

	EV210281	6/16	Homo sapiens	MDAMB231	(d)(U)C UF qEV	Grisard, Eleonora	2021	38%
Study summary Full title Homosalate boosts the release of tumor-derived Extracellular Vesicles with anti-anoikis properties All authors Eleonora Grisard, Aurianne Lescure, Nathalie Nevo, Maxime Corbé, Mabel Jouve, Gregory Lavieu, Alain Joliot, Elaine Del Nery, Lorena Martin-Jaular, Clotilde Théry Journal bioRxiv Abstract Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular (show more...)Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular vesicles (EVs). EVs could have different subcellular origin, composition and functional properties, but tools to distinguish between EV subtypes are scarce. Here, we tagged CD63-or CD9-positive EVs secreted by triple negative breast cancer cells with Nanoluciferase enzyme, to set-up a miniaturized method to quantify secretion of these two EV subtypes directly in the supernatant of cells. We performed a cell-based high-content screening to identify clinically-approved drugs able to affect EV secretion. One of the identified hits is Homosalate, an anti-inflammatory drug found in sunscreens which robustly increased EVs’release. Comparing EVs induced by Homosalate with those induced by Bafilomycin A1, we discovered that: 1) the two drugs act on EVs generated indistinct subcellular compartmentsand 2) EVs released upon treatment with Homosalate, but not with Bafilomycin A1, conferred anti-anoikis properties to another recipient tumor cell line. In conclusion, we identified a new drug modifying EV release and demonstrated that under influence of different drugs, triple negative breast cancer cells release EV subpopulations from different subcellular origins harboring distinct functional properties. (hide) EV-METRIC 38% (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 Homosalate treated 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 Ultrafiltration qEV Protein markers EV: CD9/ CD63/ CD81/ Lamp1/ syntenin/ CD98 non-EV: None Proteomics no Show all info Study aim Function/Biogenesis/cargo sorting Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells MDAMB231 EV-harvesting Medium Serum free medium Cell viability (%) 80 Cell count 27000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Pelleting performed No Ultra filtration Cut-off size (kDa) 10 kDa Membrane type PES Commercial kit qEV Other Name other separation method qEV Characterization: Protein analysis Protein Concentration Method Not determined Western Blot Antibody details provided? Yes Antibody dilution provided? Yes Lysis buffer provided? Yes Detected EV-associated proteins CD9/ CD63/ CD81/ Lamp1/ syntenin/ CD98 Characterization: Lipid analysis No Characterization: Particle analysis NTA Report type Size range/distribution Reported size (nm) 100 EV concentration Yes Particle yield number of particles per million cells: 3.00e+9

	EV210281	7/16	Homo sapiens	MDAMB231	(d)(U)C UF qEV	Grisard, Eleonora	2021	38%
Study summary Full title Homosalate boosts the release of tumor-derived Extracellular Vesicles with anti-anoikis properties All authors Eleonora Grisard, Aurianne Lescure, Nathalie Nevo, Maxime Corbé, Mabel Jouve, Gregory Lavieu, Alain Joliot, Elaine Del Nery, Lorena Martin-Jaular, Clotilde Théry Journal bioRxiv Abstract Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular (show more...)Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular vesicles (EVs). EVs could have different subcellular origin, composition and functional properties, but tools to distinguish between EV subtypes are scarce. Here, we tagged CD63-or CD9-positive EVs secreted by triple negative breast cancer cells with Nanoluciferase enzyme, to set-up a miniaturized method to quantify secretion of these two EV subtypes directly in the supernatant of cells. We performed a cell-based high-content screening to identify clinically-approved drugs able to affect EV secretion. One of the identified hits is Homosalate, an anti-inflammatory drug found in sunscreens which robustly increased EVs’release. Comparing EVs induced by Homosalate with those induced by Bafilomycin A1, we discovered that: 1) the two drugs act on EVs generated indistinct subcellular compartmentsand 2) EVs released upon treatment with Homosalate, but not with Bafilomycin A1, conferred anti-anoikis properties to another recipient tumor cell line. In conclusion, we identified a new drug modifying EV release and demonstrated that under influence of different drugs, triple negative breast cancer cells release EV subpopulations from different subcellular origins harboring distinct functional properties. (hide) EV-METRIC 38% (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 BafilomycinA1 treated 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 Ultrafiltration qEV Protein markers EV: CD9/ CD63/ CD81/ Lamp1/ syntenin/ CD98 non-EV: None Proteomics no Show all info Study aim Function/Biogenesis/cargo sorting Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells MDAMB231 EV-harvesting Medium Serum free medium Cell viability (%) 80 Cell count 27000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Pelleting performed No Ultra filtration Cut-off size (kDa) 10 kDa Membrane type PES Commercial kit qEV Other Name other separation method qEV Characterization: Protein analysis Protein Concentration Method Not determined Western Blot Antibody details provided? Yes Antibody dilution provided? Yes Lysis buffer provided? Yes Detected EV-associated proteins CD9/ CD63/ CD81/ Lamp1/ syntenin/ CD98 Characterization: Lipid analysis No Characterization: Particle analysis NTA Report type Size range/distribution Reported size (nm) 100 EV concentration Yes Particle yield number of particles per million cells: 5.00e+9

	EV210281	8/16	Homo sapiens	MCF7	(d)(U)C UF	Grisard, Eleonora	2021	0%
Study summary Full title Homosalate boosts the release of tumor-derived Extracellular Vesicles with anti-anoikis properties All authors Eleonora Grisard, Aurianne Lescure, Nathalie Nevo, Maxime Corbé, Mabel Jouve, Gregory Lavieu, Alain Joliot, Elaine Del Nery, Lorena Martin-Jaular, Clotilde Théry Journal bioRxiv Abstract Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular (show more...)Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular vesicles (EVs). EVs could have different subcellular origin, composition and functional properties, but tools to distinguish between EV subtypes are scarce. Here, we tagged CD63-or CD9-positive EVs secreted by triple negative breast cancer cells with Nanoluciferase enzyme, to set-up a miniaturized method to quantify secretion of these two EV subtypes directly in the supernatant of cells. We performed a cell-based high-content screening to identify clinically-approved drugs able to affect EV secretion. One of the identified hits is Homosalate, an anti-inflammatory drug found in sunscreens which robustly increased EVs’release. Comparing EVs induced by Homosalate with those induced by Bafilomycin A1, we discovered that: 1) the two drugs act on EVs generated indistinct subcellular compartmentsand 2) EVs released upon treatment with Homosalate, but not with Bafilomycin A1, conferred anti-anoikis properties to another recipient tumor cell line. In conclusion, we identified a new drug modifying EV release and demonstrated that under influence of different drugs, triple negative breast cancer cells release EV subpopulations from different subcellular origins harboring distinct functional properties. (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 DMSO treated 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 Ultrafiltration Protein markers EV: None non-EV: None Proteomics no Show all info Study aim Function/Biogenesis/cargo sorting Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells MCF7 EV-harvesting Medium Serum free medium Cell viability (%) 95 Cell count 3000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Pelleting performed No Ultra filtration Cut-off size (kDa) 10 kDa Membrane type PES Characterization: Protein analysis None Protein Concentration Method Not determined Western Blot Antibody details provided? Yes Antibody dilution provided? Yes Lysis buffer provided? Yes Characterization: Lipid analysis No Characterization: Particle analysis NTA EV concentration Yes Particle yield number of particles per million cells: 1.00e+7

	EV210281	9/16	Homo sapiens	MCF7	(d)(U)C UF	Grisard, Eleonora	2021	0%
Study summary Full title Homosalate boosts the release of tumor-derived Extracellular Vesicles with anti-anoikis properties All authors Eleonora Grisard, Aurianne Lescure, Nathalie Nevo, Maxime Corbé, Mabel Jouve, Gregory Lavieu, Alain Joliot, Elaine Del Nery, Lorena Martin-Jaular, Clotilde Théry Journal bioRxiv Abstract Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular (show more...)Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular vesicles (EVs). EVs could have different subcellular origin, composition and functional properties, but tools to distinguish between EV subtypes are scarce. Here, we tagged CD63-or CD9-positive EVs secreted by triple negative breast cancer cells with Nanoluciferase enzyme, to set-up a miniaturized method to quantify secretion of these two EV subtypes directly in the supernatant of cells. We performed a cell-based high-content screening to identify clinically-approved drugs able to affect EV secretion. One of the identified hits is Homosalate, an anti-inflammatory drug found in sunscreens which robustly increased EVs’release. Comparing EVs induced by Homosalate with those induced by Bafilomycin A1, we discovered that: 1) the two drugs act on EVs generated indistinct subcellular compartmentsand 2) EVs released upon treatment with Homosalate, but not with Bafilomycin A1, conferred anti-anoikis properties to another recipient tumor cell line. In conclusion, we identified a new drug modifying EV release and demonstrated that under influence of different drugs, triple negative breast cancer cells release EV subpopulations from different subcellular origins harboring distinct functional properties. (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 Homosalate treated 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 Ultrafiltration Protein markers EV: None non-EV: None Proteomics no Show all info Study aim Function/Biogenesis/cargo sorting Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells MCF7 EV-harvesting Medium Serum free medium Cell viability (%) 95 Cell count 3000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Pelleting performed No Ultra filtration Cut-off size (kDa) 10 kDa Membrane type PES Characterization: Protein analysis None Protein Concentration Method Not determined Western Blot Antibody details provided? Yes Antibody dilution provided? Yes Lysis buffer provided? Yes Characterization: Lipid analysis No Characterization: Particle analysis NTA EV concentration Yes Particle yield number of particles per million cells: 1.00e+8

	EV210281	10/16	Homo sapiens	MCF7	(d)(U)C UF	Grisard, Eleonora	2021	0%
Study summary Full title Homosalate boosts the release of tumor-derived Extracellular Vesicles with anti-anoikis properties All authors Eleonora Grisard, Aurianne Lescure, Nathalie Nevo, Maxime Corbé, Mabel Jouve, Gregory Lavieu, Alain Joliot, Elaine Del Nery, Lorena Martin-Jaular, Clotilde Théry Journal bioRxiv Abstract Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular (show more...)Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular vesicles (EVs). EVs could have different subcellular origin, composition and functional properties, but tools to distinguish between EV subtypes are scarce. Here, we tagged CD63-or CD9-positive EVs secreted by triple negative breast cancer cells with Nanoluciferase enzyme, to set-up a miniaturized method to quantify secretion of these two EV subtypes directly in the supernatant of cells. We performed a cell-based high-content screening to identify clinically-approved drugs able to affect EV secretion. One of the identified hits is Homosalate, an anti-inflammatory drug found in sunscreens which robustly increased EVs’release. Comparing EVs induced by Homosalate with those induced by Bafilomycin A1, we discovered that: 1) the two drugs act on EVs generated indistinct subcellular compartmentsand 2) EVs released upon treatment with Homosalate, but not with Bafilomycin A1, conferred anti-anoikis properties to another recipient tumor cell line. In conclusion, we identified a new drug modifying EV release and demonstrated that under influence of different drugs, triple negative breast cancer cells release EV subpopulations from different subcellular origins harboring distinct functional properties. (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 BafilomycinA1 treated 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 Ultrafiltration Protein markers EV: None non-EV: None Proteomics no Show all info Study aim Function/Biogenesis/cargo sorting Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells MCF7 EV-harvesting Medium Serum free medium Cell viability (%) 95 Cell count 3000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Pelleting performed No Ultra filtration Cut-off size (kDa) 10 kDa Membrane type PES Characterization: Protein analysis None Protein Concentration Method Not determined Western Blot Antibody details provided? Yes Antibody dilution provided? Yes Lysis buffer provided? Yes Characterization: Lipid analysis No Characterization: Particle analysis NTA EV concentration Yes Particle yield number of particles per million cells: 1.00e+8

	EV210281	11/16	Homo sapiens	Hela	(d)(U)C UF	Grisard, Eleonora	2021	0%
Study summary Full title Homosalate boosts the release of tumor-derived Extracellular Vesicles with anti-anoikis properties All authors Eleonora Grisard, Aurianne Lescure, Nathalie Nevo, Maxime Corbé, Mabel Jouve, Gregory Lavieu, Alain Joliot, Elaine Del Nery, Lorena Martin-Jaular, Clotilde Théry Journal bioRxiv Abstract Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular (show more...)Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular vesicles (EVs). EVs could have different subcellular origin, composition and functional properties, but tools to distinguish between EV subtypes are scarce. Here, we tagged CD63-or CD9-positive EVs secreted by triple negative breast cancer cells with Nanoluciferase enzyme, to set-up a miniaturized method to quantify secretion of these two EV subtypes directly in the supernatant of cells. We performed a cell-based high-content screening to identify clinically-approved drugs able to affect EV secretion. One of the identified hits is Homosalate, an anti-inflammatory drug found in sunscreens which robustly increased EVs’release. Comparing EVs induced by Homosalate with those induced by Bafilomycin A1, we discovered that: 1) the two drugs act on EVs generated indistinct subcellular compartmentsand 2) EVs released upon treatment with Homosalate, but not with Bafilomycin A1, conferred anti-anoikis properties to another recipient tumor cell line. In conclusion, we identified a new drug modifying EV release and demonstrated that under influence of different drugs, triple negative breast cancer cells release EV subpopulations from different subcellular origins harboring distinct functional properties. (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 DMSO treated 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 Ultrafiltration Protein markers EV: None non-EV: None Proteomics no Show all info Study aim Function/Biogenesis/cargo sorting Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells Hela EV-harvesting Medium Serum free medium Cell viability (%) 93 Cell count 3000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Pelleting performed No Ultra filtration Cut-off size (kDa) 10 kDa Membrane type PES Characterization: Protein analysis None Protein Concentration Method BCA Western Blot Antibody details provided? Yes Antibody dilution provided? Yes Lysis buffer provided? Yes Characterization: Lipid analysis No Characterization: Particle analysis NTA EV concentration Yes Particle yield number of particles per million cells: 1.00e+7

	EV210281	12/16	Homo sapiens	Hela	(d)(U)C UF	Grisard, Eleonora	2021	0%
Study summary Full title Homosalate boosts the release of tumor-derived Extracellular Vesicles with anti-anoikis properties All authors Eleonora Grisard, Aurianne Lescure, Nathalie Nevo, Maxime Corbé, Mabel Jouve, Gregory Lavieu, Alain Joliot, Elaine Del Nery, Lorena Martin-Jaular, Clotilde Théry Journal bioRxiv Abstract Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular (show more...)Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular vesicles (EVs). EVs could have different subcellular origin, composition and functional properties, but tools to distinguish between EV subtypes are scarce. Here, we tagged CD63-or CD9-positive EVs secreted by triple negative breast cancer cells with Nanoluciferase enzyme, to set-up a miniaturized method to quantify secretion of these two EV subtypes directly in the supernatant of cells. We performed a cell-based high-content screening to identify clinically-approved drugs able to affect EV secretion. One of the identified hits is Homosalate, an anti-inflammatory drug found in sunscreens which robustly increased EVs’release. Comparing EVs induced by Homosalate with those induced by Bafilomycin A1, we discovered that: 1) the two drugs act on EVs generated indistinct subcellular compartmentsand 2) EVs released upon treatment with Homosalate, but not with Bafilomycin A1, conferred anti-anoikis properties to another recipient tumor cell line. In conclusion, we identified a new drug modifying EV release and demonstrated that under influence of different drugs, triple negative breast cancer cells release EV subpopulations from different subcellular origins harboring distinct functional properties. (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 Homosalate treated 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 Ultrafiltration Protein markers EV: None non-EV: None Proteomics no Show all info Study aim Function/Biogenesis/cargo sorting Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells Hela EV-harvesting Medium Serum free medium Cell viability (%) 93 Cell count 3000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Pelleting performed No Ultra filtration Cut-off size (kDa) 10 kDa Membrane type PES Characterization: Protein analysis None Protein Concentration Method Not determined Western Blot Antibody details provided? Yes Antibody dilution provided? Yes Lysis buffer provided? Yes Characterization: Lipid analysis No Characterization: Particle analysis NTA EV concentration Yes Particle yield number of particles per million cells: 1.00e+8

	EV210281	13/16	Homo sapiens	Hela	(d)(U)C UF	Grisard, Eleonora	2021	0%
Study summary Full title Homosalate boosts the release of tumor-derived Extracellular Vesicles with anti-anoikis properties All authors Eleonora Grisard, Aurianne Lescure, Nathalie Nevo, Maxime Corbé, Mabel Jouve, Gregory Lavieu, Alain Joliot, Elaine Del Nery, Lorena Martin-Jaular, Clotilde Théry Journal bioRxiv Abstract Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular (show more...)Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular vesicles (EVs). EVs could have different subcellular origin, composition and functional properties, but tools to distinguish between EV subtypes are scarce. Here, we tagged CD63-or CD9-positive EVs secreted by triple negative breast cancer cells with Nanoluciferase enzyme, to set-up a miniaturized method to quantify secretion of these two EV subtypes directly in the supernatant of cells. We performed a cell-based high-content screening to identify clinically-approved drugs able to affect EV secretion. One of the identified hits is Homosalate, an anti-inflammatory drug found in sunscreens which robustly increased EVs’release. Comparing EVs induced by Homosalate with those induced by Bafilomycin A1, we discovered that: 1) the two drugs act on EVs generated indistinct subcellular compartmentsand 2) EVs released upon treatment with Homosalate, but not with Bafilomycin A1, conferred anti-anoikis properties to another recipient tumor cell line. In conclusion, we identified a new drug modifying EV release and demonstrated that under influence of different drugs, triple negative breast cancer cells release EV subpopulations from different subcellular origins harboring distinct functional properties. (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 BafilomycinA1 treated 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 Ultrafiltration Protein markers EV: None non-EV: None Proteomics no Show all info Study aim Function/Biogenesis/cargo sorting Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells Hela EV-harvesting Medium Serum free medium Cell viability (%) 93 Cell count 3000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Pelleting performed No Ultra filtration Cut-off size (kDa) 10 kDa Membrane type PES Characterization: Protein analysis None Protein Concentration Method Not determined Western Blot Antibody details provided? Yes Antibody dilution provided? Yes Lysis buffer provided? Yes Characterization: Lipid analysis No Characterization: Particle analysis NTA EV concentration Yes Particle yield number of particles per million cells: 1.00e+8

	EV210281	14/16	Homo sapiens	Jurkat	(d)(U)C UF	Grisard, Eleonora	2021	0%
Study summary Full title Homosalate boosts the release of tumor-derived Extracellular Vesicles with anti-anoikis properties All authors Eleonora Grisard, Aurianne Lescure, Nathalie Nevo, Maxime Corbé, Mabel Jouve, Gregory Lavieu, Alain Joliot, Elaine Del Nery, Lorena Martin-Jaular, Clotilde Théry Journal bioRxiv Abstract Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular (show more...)Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular vesicles (EVs). EVs could have different subcellular origin, composition and functional properties, but tools to distinguish between EV subtypes are scarce. Here, we tagged CD63-or CD9-positive EVs secreted by triple negative breast cancer cells with Nanoluciferase enzyme, to set-up a miniaturized method to quantify secretion of these two EV subtypes directly in the supernatant of cells. We performed a cell-based high-content screening to identify clinically-approved drugs able to affect EV secretion. One of the identified hits is Homosalate, an anti-inflammatory drug found in sunscreens which robustly increased EVs’release. Comparing EVs induced by Homosalate with those induced by Bafilomycin A1, we discovered that: 1) the two drugs act on EVs generated indistinct subcellular compartmentsand 2) EVs released upon treatment with Homosalate, but not with Bafilomycin A1, conferred anti-anoikis properties to another recipient tumor cell line. In conclusion, we identified a new drug modifying EV release and demonstrated that under influence of different drugs, triple negative breast cancer cells release EV subpopulations from different subcellular origins harboring distinct functional properties. (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 DMSO treated 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 Ultrafiltration Protein markers EV: None non-EV: None Proteomics no Show all info Study aim Function/Biogenesis/cargo sorting Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells Jurkat EV-harvesting Medium Serum free medium Cell viability (%) 92 Cell count 3000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Pelleting performed No Ultra filtration Cut-off size (kDa) 10 kDa Membrane type PES Characterization: Protein analysis None Protein Concentration Method Not determined Western Blot Antibody details provided? Yes Antibody dilution provided? Yes Lysis buffer provided? Yes Characterization: Lipid analysis No Characterization: Particle analysis NTA EV concentration Yes Particle yield number of particles per million cells: 1.00e+7

	EV210281	15/16	Homo sapiens	Jurkat	(d)(U)C UF	Grisard, Eleonora	2021	0%
Study summary Full title Homosalate boosts the release of tumor-derived Extracellular Vesicles with anti-anoikis properties All authors Eleonora Grisard, Aurianne Lescure, Nathalie Nevo, Maxime Corbé, Mabel Jouve, Gregory Lavieu, Alain Joliot, Elaine Del Nery, Lorena Martin-Jaular, Clotilde Théry Journal bioRxiv Abstract Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular (show more...)Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular vesicles (EVs). EVs could have different subcellular origin, composition and functional properties, but tools to distinguish between EV subtypes are scarce. Here, we tagged CD63-or CD9-positive EVs secreted by triple negative breast cancer cells with Nanoluciferase enzyme, to set-up a miniaturized method to quantify secretion of these two EV subtypes directly in the supernatant of cells. We performed a cell-based high-content screening to identify clinically-approved drugs able to affect EV secretion. One of the identified hits is Homosalate, an anti-inflammatory drug found in sunscreens which robustly increased EVs’release. Comparing EVs induced by Homosalate with those induced by Bafilomycin A1, we discovered that: 1) the two drugs act on EVs generated indistinct subcellular compartmentsand 2) EVs released upon treatment with Homosalate, but not with Bafilomycin A1, conferred anti-anoikis properties to another recipient tumor cell line. In conclusion, we identified a new drug modifying EV release and demonstrated that under influence of different drugs, triple negative breast cancer cells release EV subpopulations from different subcellular origins harboring distinct functional properties. (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 Homosalate treated 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 Ultrafiltration Protein markers EV: None non-EV: None Proteomics no Show all info Study aim Function/Biogenesis/cargo sorting Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells Jurkat EV-harvesting Medium Serum free medium Cell viability (%) 92 Cell count 3000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Pelleting performed No Ultra filtration Cut-off size (kDa) 10 kDa Membrane type PES Characterization: Protein analysis None Protein Concentration Method Not determined Western Blot Antibody details provided? Yes Antibody dilution provided? Yes Lysis buffer provided? Yes Characterization: Lipid analysis No Characterization: Particle analysis NTA EV concentration Yes Particle yield number of particles per million cells: 1.00e+8

	EV210281	16/16	Homo sapiens	Jurkat	(d)(U)C UF	Grisard, Eleonora	2021	0%
Study summary Full title Homosalate boosts the release of tumor-derived Extracellular Vesicles with anti-anoikis properties All authors Eleonora Grisard, Aurianne Lescure, Nathalie Nevo, Maxime Corbé, Mabel Jouve, Gregory Lavieu, Alain Joliot, Elaine Del Nery, Lorena Martin-Jaular, Clotilde Théry Journal bioRxiv Abstract Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular (show more...)Eukaryotic cells, including cancer cells, secrete highly heterogeneous populations of extracellular vesicles (EVs). EVs could have different subcellular origin, composition and functional properties, but tools to distinguish between EV subtypes are scarce. Here, we tagged CD63-or CD9-positive EVs secreted by triple negative breast cancer cells with Nanoluciferase enzyme, to set-up a miniaturized method to quantify secretion of these two EV subtypes directly in the supernatant of cells. We performed a cell-based high-content screening to identify clinically-approved drugs able to affect EV secretion. One of the identified hits is Homosalate, an anti-inflammatory drug found in sunscreens which robustly increased EVs’release. Comparing EVs induced by Homosalate with those induced by Bafilomycin A1, we discovered that: 1) the two drugs act on EVs generated indistinct subcellular compartmentsand 2) EVs released upon treatment with Homosalate, but not with Bafilomycin A1, conferred anti-anoikis properties to another recipient tumor cell line. In conclusion, we identified a new drug modifying EV release and demonstrated that under influence of different drugs, triple negative breast cancer cells release EV subpopulations from different subcellular origins harboring distinct functional properties. (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 BafilomycinA1 treated 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 Ultrafiltration Protein markers EV: None non-EV: None Proteomics no Show all info Study aim Function/Biogenesis/cargo sorting Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells Jurkat EV-harvesting Medium Serum free medium Cell viability (%) 92 Cell count 3000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Pelleting performed No Ultra filtration Cut-off size (kDa) 10 kDa Membrane type PES Characterization: Protein analysis None Protein Concentration Method Not determined Western Blot Antibody details provided? Yes Antibody dilution provided? Yes Lysis buffer provided? Yes Characterization: Lipid analysis No Characterization: Particle analysis NTA EV concentration Yes Particle yield number of particles per million cells: 1.00e+8

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EV-TRACK ID	EV210281
species	Homo sapiens
sample type	Cell culture
cell type	MDAMB231	MDAMB231	MDAMB231 Nanoluc-CD63	MDAMB231 Nluc-CD9	MDAMB231	MDAMB231	MDAMB231	MCF7	MCF7	MCF7	Hela	Hela	Hela	Jurkat	Jurkat	Jurkat
condition	DMSO treated	Homosalate treated	Control condition	Control condition	DMSO treated	Homosalate treated	BafilomycinA1 treated	DMSO treated	Homosalate treated	BafilomycinA1 treated	DMSO treated	Homosalate treated	BafilomycinA1 treated	DMSO treated	Homosalate treated	BafilomycinA1 treated
separation protocol	dUC/ Ultrafiltration/ qEV/ IAF capture	dUC/ Ultrafiltration/ qEV/ IAF capture	dUC/ Ultrafiltration/ qEV	dUC/ Ultrafiltration/ qEV	dUC/ Ultrafiltration/ qEV	dUC/ Ultrafiltration/ qEV	dUC/ Ultrafiltration/ qEV	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration
Exp. nr.	3	4	1	2	5	6	7	8	9	10	11	12	13	14	15	16
EV-METRIC %	75	75	50	50	38	38	38	0	0	0	0	0	0	0	0	0

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