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Details	EV-TRACK ID	Experiment nr.	Species	Sample type	Separation protocol	First author	Year	EV-METRIC
	EV230008	4/42	Mus musculus	EO771	(d)(U)C DG UF	Cocozza F	2023	89%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 89% (99th 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 sEV 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 Density gradient Ultrafiltration Protein markers EV: Alix/ CD9/ CD63/ HSP90/ MHC1/ MFGE8 non-EV: Argonaute-2 Proteomics yes EV density (g/ml) 1.015-1.085 Show all info Study aim Function Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells EO771 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 200000 Density gradient Type Discontinuous Number of initial discontinuous layers 4 Lowest density fraction 6% Highest density fraction 22% Total gradient volume, incl. sample (mL) 16 Sample volume (mL) 1 Orientation Top-down Speed (g) 187000 Duration (min) 90 Fraction volume (mL) 2 Fraction processing Centrifugation Pelleting: volume per fraction 6 Pelleting: speed (g) 200000 Pelleting: adjusted k-factor 2.29E Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose EV-subtype Distinction between multiple subtypes Density Used subtypes 1.015-1.035 Characterization: Protein analysis Protein Concentration Method microBCA Western Blot Antibody details provided? No Detected EV-associated proteins Alix/ CD9/ CD63/ HSP90/ MHC1/ MFGE8 Detected contaminants Argonaute-2 Proteomics database PRIDE Characterization: Lipid analysis No Characterization: Particle analysis NTA Report type Median Reported size (nm) 135 EV concentration Yes EM EM-type Cryo-EM Image type Wide-field

	EV230008	5/42	Mus musculus	EO771	(d)(U)C DG UF	Cocozza F	2023	89%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 89% (99th 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 VLP 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 Density gradient Ultrafiltration Protein markers EV: Alix/ CD9/ CD63/ HSP90/ MHC1/ MFGE8 non-EV: Argonaute-2 Proteomics yes EV density (g/ml) 1.015-1.085 Show all info Study aim Function Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells EO771 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 200000 Density gradient Type Discontinuous Number of initial discontinuous layers 4 Lowest density fraction 6% Highest density fraction 22% Total gradient volume, incl. sample (mL) 16 Sample volume (mL) 1 Orientation Top-down Speed (g) 187000 Duration (min) 90 Fraction volume (mL) 2 Fraction processing Centrifugation Pelleting: volume per fraction 6 Pelleting: speed (g) 200000 Pelleting: adjusted k-factor 2.29E Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose EV-subtype Distinction between multiple subtypes Density Used subtypes 1.065-1.085 Characterization: Protein analysis Protein Concentration Method microBCA Western Blot Antibody details provided? No Detected EV-associated proteins Alix/ CD9/ CD63/ HSP90/ MHC1/ MFGE8 Not detected contaminants Argonaute-2 Proteomics database PRIDE Characterization: Lipid analysis No Characterization: Particle analysis NTA Report type Median Reported size (nm) 135 EV concentration Yes EM EM-type Cryo-EM Image type Wide-field

	EV230008	1/42	Mus musculus	EO771	(d)(U)C UF	Cocozza F	2023	67%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 67% (94th 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 10k 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: CD9/ CD63/ HSP90/ MFGE8 non-EV: Argonaute-2 Proteomics yes Show all info Study aim Function Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells EO771 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 10000 Wash: volume per pellet (ml) 6 Wash: time (min) 16 Wash: Rotor Type MLA-80 Wash: speed (g) 10000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose Characterization: Protein analysis Protein Concentration Method Not determined Western Blot Antibody details provided? No Detected EV-associated proteins CD9/ CD63/ HSP90/ MFGE8 Not detected contaminants Argonaute-2 Proteomics database PRIDE Characterization: Lipid analysis No Characterization: Particle analysis NTA Report type Median Reported size (nm) 150 EV concentration Yes EM EM-type Cryo-EM Image type Wide-field

	EV230008	2/42	Mus musculus	EO771	(d)(U)C UF	Cocozza F	2023	67%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 67% (94th 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 200k 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: Alix/ CD9/ CD63/ HSP90/ MFGE8 non-EV: Argonaute-2 Proteomics yes Show all info Study aim Function Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells EO771 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 200000 Wash: volume per pellet (ml) 6 Wash: time (min) 50 Wash: Rotor Type MLA-80 Wash: speed (g) 200000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose Characterization: Protein analysis Protein Concentration Method Not determined Western Blot Antibody details provided? No Detected EV-associated proteins Alix/ CD9/ CD63/ HSP90/ MFGE8 Not detected contaminants Argonaute-2 Proteomics database PRIDE Characterization: Lipid analysis No Characterization: Particle analysis NTA Report type Distribution of sizes (%)/ Distribution of sizes (%) Reported size (nm) 20% (0-100), 70% (100-200), 10% (>200) EV concentration Yes EM EM-type Cryo-EM Image type Wide-field Report size (nm) 25-200

	EV230008	6/42	Mus musculus	EO771	(d)(U)C DG UF	Cocozza F	2023	56%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 56% (90th 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 10k light 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 Density gradient Ultrafiltration Protein markers EV: CD9/ CD63 non-EV: None Proteomics no EV density (g/ml) 1.015-1.085 Show all info Study aim Function Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells EO771 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 10000 Density gradient Type Discontinuous Number of initial discontinuous layers 4 Lowest density fraction 6% Highest density fraction 22% Total gradient volume, incl. sample (mL) 16 Sample volume (mL) 1 Orientation Top-down Speed (g) 187000 Duration (min) 90 Fraction volume (mL) 2 Fraction processing Centrifugation Pelleting: volume per fraction 6 Pelleting: speed (g) 200000 Pelleting: adjusted k-factor 5.73E Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose EV-subtype Distinction between multiple subtypes Density Used subtypes 1.015-1.035 Characterization: Protein analysis Protein Concentration Method microBCA Western Blot Antibody details provided? No Detected EV-associated proteins CD9/ CD63 Characterization: Lipid analysis No Characterization: Particle analysis None

	EV230008	7/42	Mus musculus	EO771	(d)(U)C DG UF	Cocozza F	2023	56%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 56% (90th 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 10k dense 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 Density gradient Ultrafiltration Protein markers EV: CD9/ CD63 non-EV: None Proteomics no EV density (g/ml) 1.015-1.085 Show all info Study aim Function Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells EO771 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 10000 Density gradient Type Discontinuous Number of initial discontinuous layers 4 Lowest density fraction 6% Highest density fraction 22% Total gradient volume, incl. sample (mL) 16 Sample volume (mL) 1 Orientation Top-down Speed (g) 187000 Duration (min) 90 Fraction volume (mL) 2 Fraction processing Centrifugation Pelleting: volume per fraction 6 Pelleting: speed (g) 200000 Pelleting: adjusted k-factor 5.73E Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose EV-subtype Distinction between multiple subtypes Density Used subtypes 1.065-1.085 Characterization: Protein analysis Protein Concentration Method microBCA Western Blot Antibody details provided? No Detected EV-associated proteins CD9/ CD63 Characterization: Lipid analysis No Characterization: Particle analysis None

	EV230008	41/42	Mus musculus	myr/palm-OVA-EO771	(d)(U)C DG UF	Cocozza F	2023	56%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 56% (90th 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 myr/palm-OVA Focus vesicles sEV 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 Density gradient Ultrafiltration Protein markers EV: None non-EV: None Proteomics no EV density (g/ml) 1.015-1.085 Show all info Study aim Function Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells myr/palm-OVA-EO771 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 200000 Density gradient Type Discontinuous Number of initial discontinuous layers 4 Lowest density fraction 6% Highest density fraction 22% Total gradient volume, incl. sample (mL) 16 Sample volume (mL) 1 Orientation Top-down Speed (g) 187000 Duration (min) 90 Fraction volume (mL) 2 Fraction processing Centrifugation Pelleting: volume per fraction 6 Pelleting: speed (g) 200000 Pelleting: adjusted k-factor 2.29E Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose EV-subtype Distinction between multiple subtypes Density Used subtypes 1.015-1.035 Characterization: Protein analysis Protein Concentration Method microBCA Characterization: Lipid analysis No Characterization: Particle analysis None

	EV230008	42/42	Mus musculus	myr/palm-OVA-EO771	(d)(U)C DG UF	Cocozza F	2023	56%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 56% (90th 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 myr/palm-OVA Focus vesicles VLP 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 Density gradient Ultrafiltration Protein markers EV: None non-EV: None Proteomics no EV density (g/ml) 1.015-1.085 Show all info Study aim Function Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells myr/palm-OVA-EO771 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 200000 Density gradient Type Discontinuous Number of initial discontinuous layers 4 Lowest density fraction 6% Highest density fraction 22% Total gradient volume, incl. sample (mL) 16 Sample volume (mL) 1 Orientation Top-down Speed (g) 187000 Duration (min) 90 Fraction volume (mL) 2 Fraction processing Centrifugation Pelleting: volume per fraction 6 Pelleting: speed (g) 200000 Pelleting: adjusted k-factor 2.29E Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose EV-subtype Distinction between multiple subtypes Density Used subtypes 1.065-1.085 Characterization: Protein analysis Protein Concentration Method Not determined Characterization: Lipid analysis No Characterization: Particle analysis None

	EV230008	12/42	Mus musculus	4T1	(d)(U)C UF	Cocozza F	2023	44%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 44% (84th percentile of all experiments on the same sample type) Reported Not reported Not applicable EV-enriched proteins Protein analysis: analysis of three or more EV-enriched proteins non EV-enriched protein Protein analysis: assessment of a non-EV-enriched protein qualitative and quantitative analysis Particle analysis: implementation of both qualitative and quantitative methods. For the quantitative method, the reporting of measured EV concentration is expected. electron microscopy images Particle analysis: inclusion of a widefield and close-up electron microscopy image density gradient Separation method: density gradient, at least as validation of results attributed to EVs EV density Separation method: reporting of obtained EV density ultracentrifugation specifics Separation method: reporting of g-forces, duration and rotor type of ultracentrifugation steps antibody specifics Protein analysis: antibody clone/reference number and dilution lysate preparation Protein analysis: lysis buffer composition Study data Sample type Cell culture supernatant Sample origin Control condition Focus vesicles 200k 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 yes Show all info Study aim Function Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells 4T1 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 200000 Wash: volume per pellet (ml) 6 Wash: time (min) 50 Wash: Rotor Type MLA-80 Wash: speed (g) 200000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose Characterization: Protein analysis Protein Concentration Method microBCA Proteomics database PRIDE Characterization: Lipid analysis No Characterization: Particle analysis None

	EV230008	14/42	Mus musculus	mutuDC	(d)(U)C UF	Cocozza F	2023	44%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 44% (84th percentile of all experiments on the same sample type) Reported Not reported Not applicable EV-enriched proteins Protein analysis: analysis of three or more EV-enriched proteins non EV-enriched protein Protein analysis: assessment of a non-EV-enriched protein qualitative and quantitative analysis Particle analysis: implementation of both qualitative and quantitative methods. For the quantitative method, the reporting of measured EV concentration is expected. electron microscopy images Particle analysis: inclusion of a widefield and close-up electron microscopy image density gradient Separation method: density gradient, at least as validation of results attributed to EVs EV density Separation method: reporting of obtained EV density ultracentrifugation specifics Separation method: reporting of g-forces, duration and rotor type of ultracentrifugation steps antibody specifics Protein analysis: antibody clone/reference number and dilution lysate preparation Protein analysis: lysis buffer composition Study data Sample type Cell culture supernatant Sample origin Control condition Focus vesicles 200k 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 yes Show all info Study aim Function Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells mutuDC EV-harvesting Medium EV-depleted medium Preparation of EDS overnight (16h) at >=100,000g Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 200000 Wash: volume per pellet (ml) 6 Wash: time (min) 50 Wash: Rotor Type MLA-80 Wash: speed (g) 200000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose Characterization: Protein analysis Protein Concentration Method Not determined Proteomics database PRIDE Characterization: Lipid analysis No Characterization: Particle analysis EM EM-type Cryo-EM Image type Wide-field

	EV230008	16/42	Mus musculus	Pfa1	(d)(U)C UF	Cocozza F	2023	44%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 44% (84th percentile of all experiments on the same sample type) Reported Not reported Not applicable EV-enriched proteins Protein analysis: analysis of three or more EV-enriched proteins non EV-enriched protein Protein analysis: assessment of a non-EV-enriched protein qualitative and quantitative analysis Particle analysis: implementation of both qualitative and quantitative methods. For the quantitative method, the reporting of measured EV concentration is expected. electron microscopy images Particle analysis: inclusion of a widefield and close-up electron microscopy image density gradient Separation method: density gradient, at least as validation of results attributed to EVs EV density Separation method: reporting of obtained EV density ultracentrifugation specifics Separation method: reporting of g-forces, duration and rotor type of ultracentrifugation steps antibody specifics Protein analysis: antibody clone/reference number and dilution lysate preparation Protein analysis: lysis buffer composition Study data Sample type Cell culture supernatant Sample origin Control condition Focus vesicles 200k 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: Alix/ CD9/ CD63 non-EV: None Proteomics no Show all info Study aim Function Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells Pfa1 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 200000 Wash: volume per pellet (ml) 6 Wash: time (min) 50 Wash: Rotor Type MLA-80 Wash: speed (g) 200000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose Characterization: Protein analysis Protein Concentration Method Not determined Western Blot Antibody details provided? No Detected EV-associated proteins Alix/ CD9/ CD63 Characterization: Lipid analysis No Characterization: Particle analysis None

	EV230008	17/42	Mus musculus	Pfa1	(d)(U)C UF	Cocozza F	2023	44%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 44% (84th percentile of all experiments on the same sample type) Reported Not reported Not applicable EV-enriched proteins Protein analysis: analysis of three or more EV-enriched proteins non EV-enriched protein Protein analysis: assessment of a non-EV-enriched protein qualitative and quantitative analysis Particle analysis: implementation of both qualitative and quantitative methods. For the quantitative method, the reporting of measured EV concentration is expected. electron microscopy images Particle analysis: inclusion of a widefield and close-up electron microscopy image density gradient Separation method: density gradient, at least as validation of results attributed to EVs EV density Separation method: reporting of obtained EV density ultracentrifugation specifics Separation method: reporting of g-forces, duration and rotor type of ultracentrifugation steps antibody specifics Protein analysis: antibody clone/reference number and dilution lysate preparation Protein analysis: lysis buffer composition Study data Sample type Cell culture supernatant Sample origin Control condition Focus vesicles Mix 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: Alix/ CD9/ CD63 non-EV: None Proteomics no Show all info Study aim Function Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells Pfa1 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 200000 Wash: volume per pellet (ml) 6 Wash: time (min) 720-960 Wash: Rotor Type MLA-80 Wash: speed (g) 200000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose Characterization: Protein analysis Protein Concentration Method Not determined Western Blot Antibody details provided? No Detected EV-associated proteins CD9/ CD63 Not detected EV-associated proteins Alix Characterization: Lipid analysis No Characterization: Particle analysis None

	EV230008	34/42	Mus musculus	myr/ palm-mCherry-EO771	(d)(U)C UF	Cocozza F	2023	44%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 44% (84th percentile of all experiments on the same sample type) Reported Not reported Not applicable EV-enriched proteins Protein analysis: analysis of three or more EV-enriched proteins non EV-enriched protein Protein analysis: assessment of a non-EV-enriched protein qualitative and quantitative analysis Particle analysis: implementation of both qualitative and quantitative methods. For the quantitative method, the reporting of measured EV concentration is expected. electron microscopy images Particle analysis: inclusion of a widefield and close-up electron microscopy image density gradient Separation method: density gradient, at least as validation of results attributed to EVs EV density Separation method: reporting of obtained EV density ultracentrifugation specifics Separation method: reporting of g-forces, duration and rotor type of ultracentrifugation steps antibody specifics Protein analysis: antibody clone/reference number and dilution lysate preparation Protein analysis: lysis buffer composition Study data Sample type Cell culture supernatant Sample origin myr/palm-mCherry Focus vesicles 10k 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: / non-EV: None Proteomics no Show all info Study aim Function Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells myr/ palm-mCherry-EO771 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 10000 Wash: volume per pellet (ml) 6 Wash: time (min) 16 Wash: Rotor Type MLA-80 Wash: speed (g) 10000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose Characterization: Protein analysis Protein Concentration Method Not determined Characterization: Lipid analysis No Characterization: Particle analysis NTA EV concentration Yes EM EM-type Transmission-EM Image type Wide-field

	EV230008	35/42	Mus musculus	myr/ palm-mCherry-EO771	(d)(U)C UF	Cocozza F	2023	44%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 44% (84th percentile of all experiments on the same sample type) Reported Not reported Not applicable EV-enriched proteins Protein analysis: analysis of three or more EV-enriched proteins non EV-enriched protein Protein analysis: assessment of a non-EV-enriched protein qualitative and quantitative analysis Particle analysis: implementation of both qualitative and quantitative methods. For the quantitative method, the reporting of measured EV concentration is expected. electron microscopy images Particle analysis: inclusion of a widefield and close-up electron microscopy image density gradient Separation method: density gradient, at least as validation of results attributed to EVs EV density Separation method: reporting of obtained EV density ultracentrifugation specifics Separation method: reporting of g-forces, duration and rotor type of ultracentrifugation steps antibody specifics Protein analysis: antibody clone/reference number and dilution lysate preparation Protein analysis: lysis buffer composition Study data Sample type Cell culture supernatant Sample origin myr/palm-mCherry Focus vesicles 200k 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: / non-EV: None Proteomics no Show all info Study aim Function Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells myr/ palm-mCherry-EO771 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 200000 Wash: volume per pellet (ml) 6 Wash: time (min) 50 Wash: Rotor Type MLA-80 Wash: speed (g) 200000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose Characterization: Protein analysis Protein Concentration Method Not determined Characterization: Lipid analysis No Characterization: Particle analysis NTA EV concentration Yes EM EM-type Transmission-EM Image type Wide-field

	EV230008	37/42	Mus musculus	myr/ palm-mCherry-EO771	(d)(U)C DG UF	Cocozza F	2023	43%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 43% (81st 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 myr/palm-mCherry Focus vesicles sEV 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 Density gradient Ultrafiltration Protein markers EV: None non-EV: None Proteomics no EV density (g/ml) 1.015-1.085 Show all info Study aim Function Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells myr/ palm-mCherry-EO771 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 200000 Density gradient Type Discontinuous Number of initial discontinuous layers 4 Lowest density fraction 6% Highest density fraction 22% Total gradient volume, incl. sample (mL) 16 Sample volume (mL) 1 Orientation Top-down Speed (g) 187000 Duration (min) 90 Fraction volume (mL) 2 Fraction processing Centrifugation Pelleting: volume per fraction 6 Pelleting: speed (g) 200000 Pelleting: adjusted k-factor 2.29E Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose EV-subtype Distinction between multiple subtypes Density Used subtypes 1.015-1.035 Characterization: Protein analysis None Protein Concentration Method microBCA Characterization: Lipid analysis No Characterization: Particle analysis None

	EV230008	38/42	Mus musculus	myr/ palm-mCherry-EO771	(d)(U)C DG UF	Cocozza F	2023	43%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 43% (81st 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 myr/palm-mCherry Focus vesicles VLP 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 Density gradient Ultrafiltration Protein markers EV: None non-EV: None Proteomics no EV density (g/ml) 1.015-1.085 Show all info Study aim Function Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells myr/ palm-mCherry-EO771 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 200000 Density gradient Type Discontinuous Number of initial discontinuous layers 4 Lowest density fraction 6% Highest density fraction 22% Total gradient volume, incl. sample (mL) 16 Sample volume (mL) 1 Orientation Top-down Speed (g) 187000 Duration (min) 90 Fraction volume (mL) 2 Fraction processing Centrifugation Pelleting: volume per fraction 6 Pelleting: speed (g) 200000 Pelleting: adjusted k-factor 2.29E Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose EV-subtype Distinction between multiple subtypes Density Used subtypes 1.065-1.085 Characterization: Protein analysis None Protein Concentration Method Not determined Characterization: Lipid analysis No Characterization: Particle analysis None

	EV230008	11/42	Mus musculus	4T1	(d)(U)C UF	Cocozza F	2023	33%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 33% (75th 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 10k 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 Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells 4T1 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 10000 Wash: volume per pellet (ml) 6 Wash: time (min) 16 Wash: Rotor Type MLA-80 Wash: speed (g) 10000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose Characterization: Protein analysis Protein Concentration Method microBCA Characterization: Lipid analysis No Characterization: Particle analysis None

	EV230008	13/42	Mus musculus	mutuDC	(d)(U)C UF	Cocozza F	2023	33%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 33% (75th 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 10k 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 Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells mutuDC EV-harvesting Medium EV-depleted medium Preparation of EDS overnight (16h) at >=100,000g Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 10000 Wash: volume per pellet (ml) 6 Wash: time (min) 16 Wash: Rotor Type MLA-80 Wash: speed (g) 10000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose Characterization: Protein analysis Protein Concentration Method microBCA Characterization: Lipid analysis No Characterization: Particle analysis None

	EV230008	15/42	Mus musculus	Pfa1	(d)(U)C UF	Cocozza F	2023	33%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 33% (75th 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 10k 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 Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells Pfa1 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 10000 Wash: volume per pellet (ml) 6 Wash: time (min) 16 Wash: Rotor Type MLA-80 Wash: speed (g) 10000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose Characterization: Protein analysis Protein Concentration Method Not determined Characterization: Lipid analysis No Characterization: Particle analysis None

	EV230008	18/42	Mus musculus	Mus Dunni	(d)(U)C UF	Cocozza F	2023	33%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 33% (75th 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 10k 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 Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells Mus Dunni EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 10000 Wash: volume per pellet (ml) 6 Wash: time (min) 16 Wash: Rotor Type MLA-80 Wash: speed (g) 10000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose Characterization: Protein analysis Protein Concentration Method Not determined Characterization: Lipid analysis No Characterization: Particle analysis None

	EV230008	19/42	Mus musculus	Mus Dunni	(d)(U)C UF	Cocozza F	2023	33%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 33% (75th 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 200k 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 Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells Mus Dunni EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 200000 Wash: volume per pellet (ml) 6 Wash: time (min) 50 Wash: Rotor Type MLA-80 Wash: speed (g) 200000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose Characterization: Protein analysis Protein Concentration Method Not determined Characterization: Lipid analysis No Characterization: Particle analysis None

	EV230008	20/42	Mus musculus	TS/A	(d)(U)C UF	Cocozza F	2023	33%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 33% (75th 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 10k 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 Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells TS/A EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 10000 Wash: volume per pellet (ml) 6 Wash: time (min) 16 Wash: Rotor Type MLA-80 Wash: speed (g) 10000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose Characterization: Protein analysis Protein Concentration Method Not determined Characterization: Lipid analysis No Characterization: Particle analysis None

	EV230008	21/42	Mus musculus	TS/A	(d)(U)C UF	Cocozza F	2023	33%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 33% (75th 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 200k 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 Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells TS/A EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 200000 Wash: volume per pellet (ml) 6 Wash: time (min) 50 Wash: Rotor Type MLA-80 Wash: speed (g) 200000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose Characterization: Protein analysis Protein Concentration Method Not determined Characterization: Lipid analysis No Characterization: Particle analysis None

	EV230008	22/42	Mus musculus	B16F10	(d)(U)C UF	Cocozza F	2023	33%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 33% (75th 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 10k 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 Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells B16F10 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 10000 Wash: volume per pellet (ml) 6 Wash: time (min) 16 Wash: Rotor Type MLA-80 Wash: speed (g) 10000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose Characterization: Protein analysis Protein Concentration Method Not determined Characterization: Lipid analysis No Characterization: Particle analysis NTA EV concentration Yes

	EV230008	23/42	Mus musculus	B16F10	(d)(U)C UF	Cocozza F	2023	33%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 33% (75th 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 200k 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 Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells B16F10 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 200000 Wash: volume per pellet (ml) 6 Wash: time (min) 50 Wash: Rotor Type MLA-80 Wash: speed (g) 200000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose Characterization: Protein analysis Protein Concentration Method Not determined Characterization: Lipid analysis No Characterization: Particle analysis NTA EV concentration Yes

	EV230008	24/42	Mus musculus	MCA101	(d)(U)C UF	Cocozza F	2023	33%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 33% (75th 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 10k 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 Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells MCA101 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 10000 Wash: volume per pellet (ml) 6 Wash: time (min) 16 Wash: Rotor Type MLA-80 Wash: speed (g) 10000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose Characterization: Protein analysis Protein Concentration Method Not determined Characterization: Lipid analysis No Characterization: Particle analysis NTA EV concentration Yes

	EV230008	25/42	Mus musculus	MCA101	(d)(U)C UF	Cocozza F	2023	33%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 33% (75th 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 200k 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 Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells MCA101 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 200000 Wash: volume per pellet (ml) 6 Wash: time (min) 50 Wash: Rotor Type MLA-80 Wash: speed (g) 200000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose Characterization: Protein analysis Protein Concentration Method Not determined Characterization: Lipid analysis No Characterization: Particle analysis NTA EV concentration Yes

	EV230008	26/42	Mus musculus	MB49	(d)(U)C UF	Cocozza F	2023	33%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 33% (75th 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 10k 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 Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells MB49 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 10000 Wash: volume per pellet (ml) 6 Wash: time (min) 16 Wash: Rotor Type MLA-80 Wash: speed (g) 10000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose Characterization: Protein analysis Protein Concentration Method Not determined Characterization: Lipid analysis No Characterization: Particle analysis None

	EV230008	27/42	Mus musculus	MB49	(d)(U)C UF	Cocozza F	2023	33%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 33% (75th 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 200k 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 Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells MB49 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 200000 Wash: volume per pellet (ml) 6 Wash: time (min) 50 Wash: Rotor Type MLA-80 Wash: speed (g) 200000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose Characterization: Protein analysis Protein Concentration Method Not determined Characterization: Lipid analysis No Characterization: Particle analysis None

	EV230008	28/42	Mus musculus	KP	(d)(U)C UF	Cocozza F	2023	33%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 33% (75th 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 10k 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 Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells KP EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 10000 Wash: volume per pellet (ml) 6 Wash: time (min) 16 Wash: Rotor Type MLA-80 Wash: speed (g) 10000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose Characterization: Protein analysis Protein Concentration Method Not determined Characterization: Lipid analysis No Characterization: Particle analysis NTA EV concentration Yes

	EV230008	29/42	Mus musculus	KP	(d)(U)C UF	Cocozza F	2023	33%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 33% (75th 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 200k 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 Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells KP EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 200000 Wash: volume per pellet (ml) 6 Wash: time (min) 50 Wash: Rotor Type MLA-80 Wash: speed (g) 200000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose Characterization: Protein analysis Protein Concentration Method Not determined Characterization: Lipid analysis No Characterization: Particle analysis NTA EV concentration Yes

	EV230008	30/42	Mus musculus	LLC1	(d)(U)C UF	Cocozza F	2023	33%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 33% (75th 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 10k 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 Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells LLC1 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 10000 Wash: volume per pellet (ml) 6 Wash: time (min) 16 Wash: Rotor Type MLA-80 Wash: speed (g) 10000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose EV-subtype Distinction between multiple subtypes Density Characterization: Protein analysis Protein Concentration Method Not determined Characterization: Lipid analysis No Characterization: Particle analysis NTA EV concentration Yes

	EV230008	31/42	Mus musculus	LLC1	(d)(U)C UF	Cocozza F	2023	33%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 33% (75th 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 200k 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: / non-EV: None Proteomics no Show all info Study aim Function Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells LLC1 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 200000 Wash: volume per pellet (ml) 6 Wash: time (min) 50 Wash: Rotor Type MLA-80 Wash: speed (g) 200000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose Characterization: Protein analysis Protein Concentration Method Not determined Characterization: Lipid analysis No Characterization: Particle analysis NTA EV concentration Yes

	EV230008	32/42	Mus musculus	Raw264.7	(d)(U)C UF	Cocozza F	2023	33%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 33% (75th 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 10k 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 Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells Raw264.7 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 10000 Wash: volume per pellet (ml) 6 Wash: time (min) 16 Wash: Rotor Type MLA-80 Wash: speed (g) 10000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose Characterization: Protein analysis Protein Concentration Method Not determined Characterization: Lipid analysis No Characterization: Particle analysis None

	EV230008	33/42	Mus musculus	Raw264.7	(d)(U)C UF	Cocozza F	2023	33%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 33% (75th 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 200k 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: / non-EV: None Proteomics no Show all info Study aim Function Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells Raw264.7 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 200000 Wash: volume per pellet (ml) 6 Wash: time (min) 50 Wash: Rotor Type MLA-80 Wash: speed (g) 200000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose EV-subtype Distinction between multiple subtypes Density Characterization: Protein analysis Protein Concentration Method Not determined Characterization: Lipid analysis No Characterization: Particle analysis None

	EV230008	39/42	Mus musculus	myr/palm-OVA-EO771	(d)(U)C UF	Cocozza F	2023	33%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 33% (75th 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 myr/palm-OVA Focus vesicles 10k 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 Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells myr/palm-OVA-EO771 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 10000 Wash: volume per pellet (ml) 6 Wash: time (min) 16 Wash: Rotor Type MLA-80 Wash: speed (g) 10000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose Characterization: Protein analysis Protein Concentration Method Not determined Characterization: Lipid analysis No Characterization: Particle analysis None

	EV230008	40/42	Mus musculus	myr/palm-OVA-EO771	(d)(U)C UF	Cocozza F	2023	33%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 33% (75th 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 myr/palm-OVA Focus vesicles Mix 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 Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells myr/palm-OVA-EO771 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 200000 Wash: volume per pellet (ml) 6 Wash: time (min) 720-960 Wash: Rotor Type MLA-80 Wash: speed (g) 200000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose Characterization: Protein analysis Protein Concentration Method Not determined Characterization: Lipid analysis No Characterization: Particle analysis None

	EV230008	3/42	Mus musculus	EO771	(d)(U)C UF	Cocozza F	2023	29%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 29% (68th 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 Mix 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 yes Show all info Study aim Function Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells EO771 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 200000 Wash: volume per pellet (ml) 6 Wash: time (min) 720-960 Wash: Rotor Type MLA-80 Wash: speed (g) 200000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose Characterization: Protein analysis Protein Concentration Method Not determined Proteomics database PRIDE Characterization: Lipid analysis No Characterization: Particle analysis NTA Report type Median Reported size (nm) 140 EV concentration Yes

	EV230008	8/42	Mus musculus	EO771	(d)(U)C UF AF4	Cocozza F	2023	14%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 14% (44th 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 AF4 P3 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 AF4 Protein markers EV: None non-EV: None Proteomics no Show all info Study aim Function Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells EO771 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 200000 Wash: volume per pellet (ml) 6 Wash: time (min) 50 Wash: Rotor Type MLA-80 Wash: speed (g) 200000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose Other Name other separation method AF4 EV-subtype Distinction between multiple subtypes Size Used subtypes 35-80 nm Characterization: Protein analysis None Protein Concentration Method microBCA Characterization: Lipid analysis No Characterization: Particle analysis EM EM-type Cryo-EM Image type Wide-field Other particle analysis name(1) MALS Report type Size range/distribution Report size 35-80 EV-concentration No

	EV230008	9/42	Mus musculus	EO771	(d)(U)C UF AF4	Cocozza F	2023	14%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 14% (44th 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 AF4 P4 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 AF4 Protein markers EV: None non-EV: None Proteomics no Show all info Study aim Function Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells EO771 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 200000 Wash: volume per pellet (ml) 6 Wash: time (min) 50 Wash: Rotor Type MLA-80 Wash: speed (g) 200000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose Other Name other separation method AF4 EV-subtype Distinction between multiple subtypes Size Used subtypes 80-180 nm Characterization: Protein analysis None Protein Concentration Method Not determined Characterization: Lipid analysis No Characterization: Particle analysis EM EM-type Cryo-EM Image type Wide-field Other particle analysis name(1) MALS Report type Size range/distribution Report size 80-180 EV-concentration No

	EV230008	10/42	Mus musculus	EO771	(d)(U)C UF AF4	Cocozza F	2023	14%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 14% (44th 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 AF4 P5 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 AF4 Protein markers EV: None non-EV: None Proteomics no Show all info Study aim Function Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells EO771 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 200000 Wash: volume per pellet (ml) 6 Wash: time (min) 50 Wash: Rotor Type MLA-80 Wash: speed (g) 200000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose Other Name other separation method AF4 EV-subtype Distinction between multiple subtypes Size Used subtypes 180-220 nm Characterization: Protein analysis None Protein Concentration Method Not determined Characterization: Lipid analysis No Characterization: Particle analysis EM EM-type Cryo-EM Image type Wide-field Other particle analysis name(1) MALS Report type Size range/distribution Report size 180-220 EV-concentration No

	EV230008	36/42	Mus musculus	myr/ palm-mCherry-EO771	(d)(U)C UF	Cocozza F	2023	14%
Study summary Full title Extracellular vesicles and co-isolated endogenous retroviruses from murine cancer cells differentially affect dendritic cells. All authors Cocozza F, Martin-Jaular L, Lippens L, Di Cicco A, Arribas YA, Ansart N, Dingli F, Richard M, Merle L, Jouve San Roman M, Poullet P, Loew D, Lévy D, Hendrix A, Kassiotis G, Joliot A, Tkach M, Théry C Journal EMBO J Abstract Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or (show more...)Cells secrete extracellular vesicles (EVs) and non-vesicular extracellular (nano)particles (NVEPs or ENPs) that may play a role in intercellular communication. Tumor-derived EVs have been proposed to induce immune priming of antigen presenting cells or to be immuno-suppressive agents. We suspect that such disparate functions are due to variable compositions in EV subtypes and ENPs. We aimed to characterize the array of secreted EVs and ENPs of murine tumor cell lines. Unexpectedly, we identified virus-like particles (VLPs) from endogenous murine leukemia virus in preparations of EVs produced by many tumor cells. We established a protocol to separate small EVs from VLPs and ENPs. We compared their protein composition and analyzed their functional interaction with target dendritic cells. ENPs were poorly captured and did not affect dendritic cells. Small EVs specifically induced dendritic cell death. A mixed large/dense EV/VLP preparation was most efficient to induce dendritic cell maturation and antigen presentation. Our results call for systematic re-evaluation of the respective proportions and functions of non-viral EVs and VLPs produced by murine tumors and their contribution to tumor progression. (hide) EV-METRIC 14% (44th 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 myr/palm-mCherry Focus vesicles Mix 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 Sample Species Mus musculus Sample Type Cell culture supernatant EV-producing cells myr/ palm-mCherry-EO771 EV-harvesting Medium Serum free medium Cell viability (%) 85 Cell count 100000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: rotor type MLA-80 Pelleting: speed (g) 200000 Wash: volume per pellet (ml) 6 Wash: time (min) 720-960 Wash: Rotor Type MLA-80 Wash: speed (g) 200000 Ultra filtration Cut-off size (kDa) 10 Membrane type Regenerated cellulose Characterization: Protein analysis None Protein Concentration Method Not determined Characterization: Lipid analysis No Characterization: Particle analysis None

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EV-TRACK ID	EV230008
species	Mus musculus
sample type	Cell culture
cell type	EO771	EO771	EO771	EO771	EO771	EO771	myr/palm-OVA-EO771	myr/palm-OVA-EO771	4T1	mutuDC	Pfa1	Pfa1	myr/ palm-mCherry-EO771	myr/ palm-mCherry-EO771	myr/ palm-mCherry-EO771	myr/ palm-mCherry-EO771	4T1	mutuDC	Pfa1	Mus Dunni	Mus Dunni	TS/A	TS/A	B16F10	B16F10	MCA101	MCA101	MB49	MB49	KP	KP	LLC1	LLC1	Raw264.7	Raw264.7	myr/palm-OVA-EO771	myr/palm-OVA-EO771	EO771	EO771	EO771	EO771	myr/ palm-mCherry-EO771
medium	Serum free medium	Serum free medium	Serum free medium	Serum free medium	Serum free medium	Serum free medium	Serum free medium	Serum free medium	Serum free medium	EV-depleted medium	Serum free medium	Serum free medium	Serum free medium	Serum free medium	Serum free medium	Serum free medium	Serum free medium	EV-depleted medium	Serum free medium	Serum free medium	Serum free medium	Serum free medium	Serum free medium	Serum free medium	Serum free medium	Serum free medium	Serum free medium	Serum free medium	Serum free medium	Serum free medium	Serum free medium	Serum free medium	Serum free medium	Serum free medium	Serum free medium	Serum free medium	Serum free medium	Serum free medium	Serum free medium	Serum free medium	Serum free medium	Serum free medium
condition	Control condition	Control condition	Control condition	Control condition	Control condition	Control condition	myr/palm-OVA	myr/palm-OVA	Control condition	Control condition	Control condition	Control condition	myr/palm-mCherry	myr/palm-mCherry	myr/palm-mCherry	myr/palm-mCherry	Control condition	Control condition	Control condition	Control condition	Control condition	Control condition	Control condition	Control condition	Control condition	Control condition	Control condition	Control condition	Control condition	Control condition	Control condition	Control condition	Control condition	Control condition	Control condition	myr/palm-OVA	myr/palm-OVA	Control condition	Control condition	Control condition	Control condition	myr/palm-mCherry
separation protocol	dUC/ Density gradient/ Ultrafiltration	dUC/ Density gradient/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Density gradient/ Ultrafiltration	dUC/ Density gradient/ Ultrafiltration	dUC/ Density gradient/ Ultrafiltration	dUC/ Density gradient/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Density gradient/ Ultrafiltration	dUC/ Density gradient/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration	dUC/ Ultrafiltration/ AF4	dUC/ Ultrafiltration/ AF4	dUC/ Ultrafiltration/ AF4	dUC/ Ultrafiltration
EV subtype	1.015-1.035	1.065-1.085	NA	NA	1.015-1.035	1.065-1.085	1.015-1.035	1.065-1.085	NA	NA	NA	NA	NA	NA	1.015-1.035	1.065-1.085	NA	NA	NA	NA	NA	NA	NA	NA	NA	NA	NA	NA	NA	NA	NA	NA	NA	NA	NA	NA	NA	NA	35-80 nm	80-180 nm	180-220 nm	NA
vesicle related term	sEV	VLP	10k	200k	10k light	10k dense	sEV	VLP	200k	200k	200k	Mix	10k	200k	sEV	VLP	10k	10k	10k	10k	200k	10k	200k	10k	200k	10k	200k	10k	200k	10k	200k	10k	200k	10k	200k	10k	Mix	Mix	AF4 P3	AF4 P4	AF4 P5	Mix
Exp. nr.	4	5	1	2	6	7	41	42	12	14	16	17	34	35	37	38	11	13	15	18	19	20	21	22	23	24	25	26	27	28	29	30	31	32	33	39	40	3	8	9	10	36
EV-METRIC %	89	89	67	67	56	56	56	56	44	44	44	44	44	44	43	43	33	33	33	33	33	33	33	33	33	33	33	33	33	33	33	33	33	33	33	33	33	29	14	14	14	14

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