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Details	EV-TRACK ID	Experiment nr.	Species	Sample type	Separation protocol	First author	Year	EV-METRIC
	EV210113	1/4	Homo sapiens	primary adipose tissue-derived mesenchymal stromal cells	(d)(U)C	Gorgun, Cansu	2021	56%
Study summary Full title Role of extracellular vesicles from adipose tissue- and bone marrow-mesenchymal stromal cells in endothelial proliferation and chondrogenesis All authors Cansu Gorgun, Maria Elisabetta Federica Palamà, Daniele Reverberi, Maria Cristina Gagliani, Katia Cortese, Roberta Tasso, Chiara Gentili Journal Stem Cells Transl Med Abstract The secretome of mesenchymal stromal cells (MSCs) derived from different tissue sources is considere (show more...)The secretome of mesenchymal stromal cells (MSCs) derived from different tissue sources is considered an innovative therapeutic tool for regenerative medicine. Although adipose tissue-and bone marrow-derived MSCs (ADSCs and BMSCs, respectively) share many biological features, the different tissue origins can be mirrored by variations in their secretory profile, and in particular in the secreted extracellular vesicles (EVs). In this study, we carried out a detailed and comparative characterization of middle- and small-sized EVs (mEVs and sEVs, respectively) released by either ADSCs or BMSCs. Their involvement in an endochondral ossification setting was investigated using ex vivo metatarsal culture models that allowed to explore both blood vessel sprouting and bone growth plate dynamics. Although EVs separated from both cell sources presented similar characteristics in terms of size, concentration, and marker expression, they exhibited different characteristics in terms of protein content and functional effects. ADSC-EVs overexpressed pro-angiogenic factors in comparison to the BMSC-counterpart, and, consequently, they were able to induce a significant increase in endothelial cord outgrowth. On the other hand, BMSC-EVs contained a higher amount of pro-differentiation and chemotactic proteins, and they were able to prompt growth plate organization. The present study highlights the importance of selecting the appropriate cell source of EVs for targeted therapeutic applications. (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 Other / middle sized extracellular vesicles Separation protocol Separation protocol Gives a short, non-chronological overview of the different steps of the separation protocol. dUC = (Differential) (ultra)centrifugation DG = density gradient UF = ultrafiltration SEC = size-exclusion chromatography IAF = immuno-affinity capture (d)(U)C Protein markers EV: CD81/ Flotillin1/ CD63/ CD9/ Syntenin non-EV: GRP94 Proteomics no Show all info Study aim Function Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells primary adipose tissue-derived mesenchymal stromal cells EV-harvesting Medium Serum free medium Cell viability (%) 92 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Below or equal to 800 g Between 800 g and 10,000 g Between 10,000 g and 50,000 g Pelleting performed Yes Pelleting: time(min) 40 Pelleting: rotor type SW 28 Pelleting: speed (g) 10000 Wash: volume per pellet (ml) 5 Wash: time (min) 40 Wash: Rotor Type SW 55 Ti Wash: speed (g) 10000 Characterization: Protein analysis Protein Concentration Method microBCA Western Blot Antibody details provided? No Detected EV-associated proteins Flotillin1/ CD63/ Syntenin/ CD81 Not detected EV-associated proteins CD9 Not detected contaminants GRP94 Flow cytometry Type of Flow cytometry BD FACSAria II (BD Biosciences) Hardware adaptation to ~100nm EV's doi: 10.1002/cpsc.76. PMID: 30624011 Calibration bead size 0.1;0.16;0.2;0.24;0.3;0.5;0.9 Antibody details provided? No Detected EV-associated proteins CD63/ CD9/ CD81 Characterization: Lipid analysis No Characterization: Particle analysis TRPS Report type Mean Reported size (nm) 388+/-32.7 EM EM-type Transmission-EM Image type Close-up

	EV210113	2/4	Homo sapiens	primary adipose tissue-derived mesenchymal stromal cells	(d)(U)C	Gorgun, Cansu	2021	56%
Study summary Full title Role of extracellular vesicles from adipose tissue- and bone marrow-mesenchymal stromal cells in endothelial proliferation and chondrogenesis All authors Cansu Gorgun, Maria Elisabetta Federica Palamà, Daniele Reverberi, Maria Cristina Gagliani, Katia Cortese, Roberta Tasso, Chiara Gentili Journal Stem Cells Transl Med Abstract The secretome of mesenchymal stromal cells (MSCs) derived from different tissue sources is considere (show more...)The secretome of mesenchymal stromal cells (MSCs) derived from different tissue sources is considered an innovative therapeutic tool for regenerative medicine. Although adipose tissue-and bone marrow-derived MSCs (ADSCs and BMSCs, respectively) share many biological features, the different tissue origins can be mirrored by variations in their secretory profile, and in particular in the secreted extracellular vesicles (EVs). In this study, we carried out a detailed and comparative characterization of middle- and small-sized EVs (mEVs and sEVs, respectively) released by either ADSCs or BMSCs. Their involvement in an endochondral ossification setting was investigated using ex vivo metatarsal culture models that allowed to explore both blood vessel sprouting and bone growth plate dynamics. Although EVs separated from both cell sources presented similar characteristics in terms of size, concentration, and marker expression, they exhibited different characteristics in terms of protein content and functional effects. ADSC-EVs overexpressed pro-angiogenic factors in comparison to the BMSC-counterpart, and, consequently, they were able to induce a significant increase in endothelial cord outgrowth. On the other hand, BMSC-EVs contained a higher amount of pro-differentiation and chemotactic proteins, and they were able to prompt growth plate organization. The present study highlights the importance of selecting the appropriate cell source of EVs for targeted therapeutic applications. (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 Other / small sized extracellular vesicles Separation protocol Separation protocol Gives a short, non-chronological overview of the different steps of the separation protocol. dUC = (Differential) (ultra)centrifugation DG = density gradient UF = ultrafiltration SEC = size-exclusion chromatography IAF = immuno-affinity capture (d)(U)C Protein markers EV: CD81/ Flotillin1/ CD63/ CD9/ Syntenin non-EV: GRP94 Proteomics no Show all info Study aim Function Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells primary adipose tissue-derived mesenchymal stromal cells EV-harvesting Medium Serum free medium Cell viability (%) 92 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Below or equal to 800 g Between 800 g and 10,000 g Between 100,000 g and 150,000 g Pelleting performed Yes Pelleting: time(min) 120 Pelleting: rotor type SW 28 Pelleting: speed (g) 100000 Wash: volume per pellet (ml) 5 Wash: time (min) 120 Wash: Rotor Type SW 55 Ti Wash: speed (g) 100000 Characterization: Protein analysis Protein Concentration Method microBCA Western Blot Antibody details provided? No Detected EV-associated proteins Flotillin1/ CD63/ Syntenin/ CD81 Not detected EV-associated proteins CD9 Not detected contaminants GRP94 Flow cytometry Type of Flow cytometry BD FACSAria II (BD Biosciences) Hardware adaptation to ~100nm EV's doi: 10.1002/cpsc.76. PMID: 30624011 Calibration bead size 0.1;0.16;0.2;0.24;0.3;0.5;0.9 Antibody details provided? No Detected EV-associated proteins CD63/ CD9/ CD81 Characterization: Lipid analysis No Characterization: Particle analysis TRPS Report type Mean Reported size (nm) 138+/-12 EM EM-type Transmission-EM Image type Close-up

	EV210113	3/4	Homo sapiens	primary bone marrow-derived mesenchymal stromal cells	(d)(U)C	Gorgun, Cansu	2021	56%
Study summary Full title Role of extracellular vesicles from adipose tissue- and bone marrow-mesenchymal stromal cells in endothelial proliferation and chondrogenesis All authors Cansu Gorgun, Maria Elisabetta Federica Palamà, Daniele Reverberi, Maria Cristina Gagliani, Katia Cortese, Roberta Tasso, Chiara Gentili Journal Stem Cells Transl Med Abstract The secretome of mesenchymal stromal cells (MSCs) derived from different tissue sources is considere (show more...)The secretome of mesenchymal stromal cells (MSCs) derived from different tissue sources is considered an innovative therapeutic tool for regenerative medicine. Although adipose tissue-and bone marrow-derived MSCs (ADSCs and BMSCs, respectively) share many biological features, the different tissue origins can be mirrored by variations in their secretory profile, and in particular in the secreted extracellular vesicles (EVs). In this study, we carried out a detailed and comparative characterization of middle- and small-sized EVs (mEVs and sEVs, respectively) released by either ADSCs or BMSCs. Their involvement in an endochondral ossification setting was investigated using ex vivo metatarsal culture models that allowed to explore both blood vessel sprouting and bone growth plate dynamics. Although EVs separated from both cell sources presented similar characteristics in terms of size, concentration, and marker expression, they exhibited different characteristics in terms of protein content and functional effects. ADSC-EVs overexpressed pro-angiogenic factors in comparison to the BMSC-counterpart, and, consequently, they were able to induce a significant increase in endothelial cord outgrowth. On the other hand, BMSC-EVs contained a higher amount of pro-differentiation and chemotactic proteins, and they were able to prompt growth plate organization. The present study highlights the importance of selecting the appropriate cell source of EVs for targeted therapeutic applications. (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 Other / middle sized extracellular vesicles Separation protocol Separation protocol Gives a short, non-chronological overview of the different steps of the separation protocol. dUC = (Differential) (ultra)centrifugation DG = density gradient UF = ultrafiltration SEC = size-exclusion chromatography IAF = immuno-affinity capture (d)(U)C Protein markers EV: CD81/ Flotillin1/ CD63/ CD9/ Syntenin non-EV: GRP94 Proteomics no Show all info Study aim Function Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells primary bone marrow-derived mesenchymal stromal cells EV-harvesting Medium Serum free medium Cell viability (%) 86 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Below or equal to 800 g Between 800 g and 10,000 g Between 10,000 g and 50,000 g Pelleting performed Yes Pelleting: time(min) 40 Pelleting: rotor type SW 28 Pelleting: speed (g) 10000 Wash: volume per pellet (ml) 5 Wash: time (min) 40 Wash: Rotor Type SW 55 Ti Wash: speed (g) 10000 Characterization: Protein analysis Protein Concentration Method microBCA Western Blot Antibody details provided? No Detected EV-associated proteins Flotillin1/ CD63/ Syntenin/ CD81 Not detected EV-associated proteins CD9 Not detected contaminants GRP94 Flow cytometry Type of Flow cytometry BD FACSAria II (BD Biosciences) Hardware adaptation to ~100nm EV's doi: 10.1002/cpsc.76. PMID: 30624011 Calibration bead size 0.1;0.16;0.2;0.24;0.3;0.5;0.9 Antibody details provided? No Detected EV-associated proteins CD63/ CD9/ CD81 Characterization: Lipid analysis No Characterization: Particle analysis TRPS Report type Mean Reported size (nm) 346+/-42.1 EM EM-type Transmission-EM Image type Close-up

	EV210113	4/4	Homo sapiens	primary bone marrow-derived mesenchymal stromal cells	(d)(U)C	Gorgun, Cansu	2021	56%
Study summary Full title Role of extracellular vesicles from adipose tissue- and bone marrow-mesenchymal stromal cells in endothelial proliferation and chondrogenesis All authors Cansu Gorgun, Maria Elisabetta Federica Palamà, Daniele Reverberi, Maria Cristina Gagliani, Katia Cortese, Roberta Tasso, Chiara Gentili Journal Stem Cells Transl Med Abstract The secretome of mesenchymal stromal cells (MSCs) derived from different tissue sources is considere (show more...)The secretome of mesenchymal stromal cells (MSCs) derived from different tissue sources is considered an innovative therapeutic tool for regenerative medicine. Although adipose tissue-and bone marrow-derived MSCs (ADSCs and BMSCs, respectively) share many biological features, the different tissue origins can be mirrored by variations in their secretory profile, and in particular in the secreted extracellular vesicles (EVs). In this study, we carried out a detailed and comparative characterization of middle- and small-sized EVs (mEVs and sEVs, respectively) released by either ADSCs or BMSCs. Their involvement in an endochondral ossification setting was investigated using ex vivo metatarsal culture models that allowed to explore both blood vessel sprouting and bone growth plate dynamics. Although EVs separated from both cell sources presented similar characteristics in terms of size, concentration, and marker expression, they exhibited different characteristics in terms of protein content and functional effects. ADSC-EVs overexpressed pro-angiogenic factors in comparison to the BMSC-counterpart, and, consequently, they were able to induce a significant increase in endothelial cord outgrowth. On the other hand, BMSC-EVs contained a higher amount of pro-differentiation and chemotactic proteins, and they were able to prompt growth plate organization. The present study highlights the importance of selecting the appropriate cell source of EVs for targeted therapeutic applications. (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 Other / small sized extracellular vesicles Separation protocol Separation protocol Gives a short, non-chronological overview of the different steps of the separation protocol. dUC = (Differential) (ultra)centrifugation DG = density gradient UF = ultrafiltration SEC = size-exclusion chromatography IAF = immuno-affinity capture (d)(U)C Protein markers EV: CD81/ Flotillin1/ CD63/ CD9/ Syntenin non-EV: GRP94 Proteomics no Show all info Study aim Function Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells primary bone marrow-derived mesenchymal stromal cells EV-harvesting Medium Serum free medium Cell viability (%) 86 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Below or equal to 800 g Between 800 g and 10,000 g Between 100,000 g and 150,000 g Pelleting performed Yes Pelleting: time(min) 120 Pelleting: rotor type SW 28 Pelleting: speed (g) 100000 Wash: volume per pellet (ml) 5 Wash: time (min) 120 Wash: Rotor Type SW 55 Ti Wash: speed (g) 100000 Characterization: Protein analysis Protein Concentration Method microBCA Western Blot Antibody details provided? No Detected EV-associated proteins Syntenin/ CD63/ CD81 Not detected EV-associated proteins Flotillin1/ CD9 Not detected contaminants GRP94 Flow cytometry Type of Flow cytometry BD FACSAria II (BD Biosciences) Hardware adaptation to ~100nm EV's doi: 10.1002/cpsc.76. PMID: 30624011 Calibration bead size 0.1;0.16;0.2;0.24;0.3;0.5;0.9 Antibody details provided? No Detected EV-associated proteins CD63/ CD9/ CD81 Characterization: Lipid analysis No Characterization: Particle analysis TRPS Report type Mean Reported size (nm) 152+/-33.2 EM EM-type Transmission-EM Image type Close-up

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EV-TRACK ID	EV210113
species	Homo sapiens
sample type	Cell culture
cell type	primary adipose tissue-derived mesenchymal stromal cells	primary adipose tissue-derived mesenchymal stromal cells	primary bone marrow-derived mesenchymal stromal cells	primary bone marrow-derived mesenchymal stromal cells
condition	Control condition	Control condition	Control condition	Control condition
separation protocol	(d)(U)C	(d)(U)C	(d)(U)C	(d)(U)C
vesicle related term	Other middle sized EVs	Other small sized EVs	Other middle sized EVs	Other small sized EVs
Exp. nr.	1	2	3	4
EV-METRIC %	56	56	56	56