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Details	EV-TRACK ID	Experiment nr.	Species	Sample type	Separation protocol	First author	Year	EV-METRIC
	EV180022	1/4	Homo sapiens	pluripotent stem cells	(d)(U)C	Kaur S	2018	67%
Study summary Full title Small non-coding RNA landscape of extracellular vesicles from human stem cells. All authors Kaur S, Abu-Shahba AG, Paananen RO, Hongisto H, Hiidenmaa H, Skottman H, Seppänen-Kaijansinkko R, Mannerström B Journal J Cell Sci Abstract Extracellular vesicles (EVs) are reported to be involved in stem cell maintenance, self-renewal, and (show more...)Extracellular vesicles (EVs) are reported to be involved in stem cell maintenance, self-renewal, and differentiation. Due to their bioactive cargoes influencing cell fate and function, interest in EVs in regenerative medicine has rapidly increased. EV-derived small non-coding RNA mimic the functions of the parent stem cells, regulating the maintenance and differentiation of stem cells, controlling the intercellular regulation of gene expression, and eventually affecting the cell fate. In this study, we used RNA sequencing to provide a comprehensive overview of the expression profiles of small non-coding transcripts carried by the EVs derived from human adipose tissue stromal/stem cells (AT-MSCs) and human pluripotent stem cells (hPSCs), both human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSC). Both hPSCs and AT-MSCs were characterized and their EVs were extracted using standard protocols. Small non-coding RNA sequencing from EVs showed that hPSCs and AT-MSCs showed distinct profiles, unique for each stem cell source. Interestingly, in hPSCs, most abundant miRNAs were from specific miRNA families regulating pluripotency, reprogramming and differentiation (miR-17-92, mir-200, miR-302/367, miR-371/373, CM19 microRNA cluster). For the AT-MSCs, the highly expressed miRNAs were found to be regulating osteogenesis (let-7/98, miR-10/100, miR-125, miR-196, miR-199, miR-615-3p, mir-22-3p, mir-24-3p, mir-27a-3p, mir-193b-5p, mir-195-3p). Additionally, abundant small nuclear and nucleolar RNA were detected in hPSCs, whereas Y- and tRNA were found in AT-MSCs. Identification of EV-miRNA and non-coding RNA signatures released by these stem cells will provide clues towards understanding their role in intracellular communication, and well as their roles in maintaining the stem cell niche. (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 extracellular vesicle Separation protocol Separation protocol Gives a short, non-chronological overview of the different steps of the separation protocol. dUC = (Differential) (ultra)centrifugation DG = density gradient UF = ultrafiltration SEC = size-exclusion chromatography IAF = immuno-affinity capture (d)(U)C Adj. k-factor 208.3 (pelleting) / 208.3 (washing) Protein markers EV: TSG101/ HSP70/ CD63/ CD90 non-EV: Calnexin Proteomics no Show all info Study aim Identification of content (omics approaches), Technical analysis comparing/optimizing EV-related methods Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells pluripotent stem cells EV-harvesting Medium Serum free medium Cell viability (%) NA Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps 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) 121896 Pelleting: adjusted k-factor 208.3 Wash: time (min) 120 Wash: Rotor Type SW 28 Wash: speed (g) 121896 Wash: adjusted k-factor 208.3 Characterization: Protein analysis Protein Concentration Method Not determined Western Blot Antibody details provided? Yes Lysis buffer provided? Yes Detected EV-associated proteins CD63, HSP70, TSG101, CD90 Not detected contaminants Calnexin Characterization: Lipid analysis No Characterization: Particle analysis NTA Report type Size range/distribution Reported size (nm) 101-500 EV concentration Yes EM EM-type Transmission-EM Image type Wide-field

	EV180022	4/4	Homo sapiens	pluripotent stem cells	miRCURY Exosome Isolation Kit (Exiqon A/S, Vedbaek, Denmark)	Kaur S	2018	67%
Study summary Full title Small non-coding RNA landscape of extracellular vesicles from human stem cells. All authors Kaur S, Abu-Shahba AG, Paananen RO, Hongisto H, Hiidenmaa H, Skottman H, Seppänen-Kaijansinkko R, Mannerström B Journal J Cell Sci Abstract Extracellular vesicles (EVs) are reported to be involved in stem cell maintenance, self-renewal, and (show more...)Extracellular vesicles (EVs) are reported to be involved in stem cell maintenance, self-renewal, and differentiation. Due to their bioactive cargoes influencing cell fate and function, interest in EVs in regenerative medicine has rapidly increased. EV-derived small non-coding RNA mimic the functions of the parent stem cells, regulating the maintenance and differentiation of stem cells, controlling the intercellular regulation of gene expression, and eventually affecting the cell fate. In this study, we used RNA sequencing to provide a comprehensive overview of the expression profiles of small non-coding transcripts carried by the EVs derived from human adipose tissue stromal/stem cells (AT-MSCs) and human pluripotent stem cells (hPSCs), both human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSC). Both hPSCs and AT-MSCs were characterized and their EVs were extracted using standard protocols. Small non-coding RNA sequencing from EVs showed that hPSCs and AT-MSCs showed distinct profiles, unique for each stem cell source. Interestingly, in hPSCs, most abundant miRNAs were from specific miRNA families regulating pluripotency, reprogramming and differentiation (miR-17-92, mir-200, miR-302/367, miR-371/373, CM19 microRNA cluster). For the AT-MSCs, the highly expressed miRNAs were found to be regulating osteogenesis (let-7/98, miR-10/100, miR-125, miR-196, miR-199, miR-615-3p, mir-22-3p, mir-24-3p, mir-27a-3p, mir-193b-5p, mir-195-3p). Additionally, abundant small nuclear and nucleolar RNA were detected in hPSCs, whereas Y- and tRNA were found in AT-MSCs. Identification of EV-miRNA and non-coding RNA signatures released by these stem cells will provide clues towards understanding their role in intracellular communication, and well as their roles in maintaining the stem cell niche. (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 extracellular vesicle Separation protocol Separation protocol Gives a short, non-chronological overview of the different steps of the separation protocol. dUC = (Differential) (ultra)centrifugation DG = density gradient UF = ultrafiltration SEC = size-exclusion chromatography IAF = immuno-affinity capture miRCURY Exosome Isolation Kit (Exiqon A/S, Vedbaek, Denmark) Protein markers EV: None non-EV: None Proteomics no Show all info Study aim Identification of content (omics approaches), Technical analysis comparing/optimizing EV-related methods Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells pluripotent stem cells EV-harvesting Medium Serum free medium Cell viability (%) NA Separation Method Commercial kit miRCURY Exosome Isolation Kit (Exiqon A/S, Vedbaek, Denmark) Other Name other separation method miRCURY Exosome Isolation Kit (Exiqon A/S, Vedbaek, Denmark) Characterization: Protein analysis None Protein Concentration Method Not determined Characterization: RNA analysis Database No Proteinase treatment No RNAse treatment No Characterization: Lipid analysis No

	EV180022	3/4	Homo sapiens	adipose tissue-derived mesenchymal stem cells	(d)(U)C Filtration	Kaur S	2018	66%
Study summary Full title Small non-coding RNA landscape of extracellular vesicles from human stem cells. All authors Kaur S, Abu-Shahba AG, Paananen RO, Hongisto H, Hiidenmaa H, Skottman H, Seppänen-Kaijansinkko R, Mannerström B Journal J Cell Sci Abstract Extracellular vesicles (EVs) are reported to be involved in stem cell maintenance, self-renewal, and (show more...)Extracellular vesicles (EVs) are reported to be involved in stem cell maintenance, self-renewal, and differentiation. Due to their bioactive cargoes influencing cell fate and function, interest in EVs in regenerative medicine has rapidly increased. EV-derived small non-coding RNA mimic the functions of the parent stem cells, regulating the maintenance and differentiation of stem cells, controlling the intercellular regulation of gene expression, and eventually affecting the cell fate. In this study, we used RNA sequencing to provide a comprehensive overview of the expression profiles of small non-coding transcripts carried by the EVs derived from human adipose tissue stromal/stem cells (AT-MSCs) and human pluripotent stem cells (hPSCs), both human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSC). Both hPSCs and AT-MSCs were characterized and their EVs were extracted using standard protocols. Small non-coding RNA sequencing from EVs showed that hPSCs and AT-MSCs showed distinct profiles, unique for each stem cell source. Interestingly, in hPSCs, most abundant miRNAs were from specific miRNA families regulating pluripotency, reprogramming and differentiation (miR-17-92, mir-200, miR-302/367, miR-371/373, CM19 microRNA cluster). For the AT-MSCs, the highly expressed miRNAs were found to be regulating osteogenesis (let-7/98, miR-10/100, miR-125, miR-196, miR-199, miR-615-3p, mir-22-3p, mir-24-3p, mir-27a-3p, mir-193b-5p, mir-195-3p). Additionally, abundant small nuclear and nucleolar RNA were detected in hPSCs, whereas Y- and tRNA were found in AT-MSCs. Identification of EV-miRNA and non-coding RNA signatures released by these stem cells will provide clues towards understanding their role in intracellular communication, and well as their roles in maintaining the stem cell niche. (hide) EV-METRIC 66% (93rd percentile of all experiments on the same sample type) Reported Not reported Not applicable EV-enriched proteins Protein analysis: analysis of three or more EV-enriched proteins non EV-enriched protein Protein analysis: assessment of a non-EV-enriched protein qualitative and quantitative analysis Particle analysis: implementation of both qualitative and quantitative methods. For the quantitative method, the reporting of measured EV concentration is expected. electron microscopy images Particle analysis: inclusion of a widefield and close-up electron microscopy image density gradient Separation method: density gradient, at least as validation of results attributed to EVs EV density Separation method: reporting of obtained EV density ultracentrifugation specifics Separation method: reporting of g-forces, duration and rotor type of ultracentrifugation steps antibody specifics Protein analysis: antibody clone/reference number and dilution lysate preparation Protein analysis: lysis buffer composition Study data Sample type Cell culture supernatant Sample origin Control condition Focus vesicles extracellular vesicle Separation protocol Separation protocol Gives a short, non-chronological overview of the different steps of the separation protocol. dUC = (Differential) (ultra)centrifugation DG = density gradient UF = ultrafiltration SEC = size-exclusion chromatography IAF = immuno-affinity capture (d)(U)C Filtration Adj. k-factor 208.3 (pelleting) / 208.3 (washing) Protein markers EV: TSG101/ CD63/ CD90 non-EV: Calnexin Proteomics no Show all info Study aim Identification of content (omics approaches), Technical analysis comparing/optimizing EV-related methods Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells adipose tissue-derived mesenchymal stem cells EV-harvesting Medium EV-depleted serum Preparation of EDS >=18h at >= 100,000g Cell viability (%) NA Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps 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) 121896 Pelleting: adjusted k-factor 208.3 Wash: time (min) 120 Wash: Rotor Type SW 28 Wash: speed (g) 121896 Wash: adjusted k-factor 208.3 Characterization: Protein analysis Protein Concentration Method Not determined Western Blot Antibody details provided? Yes Lysis buffer provided? Yes Detected EV-associated proteins CD63, TSG101, CD90 Not detected contaminants Calnexin Characterization: Lipid analysis No Characterization: Particle analysis NTA Report type Size range/distribution Reported size (nm) 101-500 EV concentration Yes EM EM-type Transmission-EM Image type Wide-field

	EV180022	2/4	Homo sapiens	adipose tissue-derived mesenchymal stem cells	miRCURY Exosome Isolation Kit (Exiqon A/S, Vedbaek, Denmark)	Kaur S	2018	0%
Study summary Full title Small non-coding RNA landscape of extracellular vesicles from human stem cells. All authors Kaur S, Abu-Shahba AG, Paananen RO, Hongisto H, Hiidenmaa H, Skottman H, Seppänen-Kaijansinkko R, Mannerström B Journal J Cell Sci Abstract Extracellular vesicles (EVs) are reported to be involved in stem cell maintenance, self-renewal, and (show more...)Extracellular vesicles (EVs) are reported to be involved in stem cell maintenance, self-renewal, and differentiation. Due to their bioactive cargoes influencing cell fate and function, interest in EVs in regenerative medicine has rapidly increased. EV-derived small non-coding RNA mimic the functions of the parent stem cells, regulating the maintenance and differentiation of stem cells, controlling the intercellular regulation of gene expression, and eventually affecting the cell fate. In this study, we used RNA sequencing to provide a comprehensive overview of the expression profiles of small non-coding transcripts carried by the EVs derived from human adipose tissue stromal/stem cells (AT-MSCs) and human pluripotent stem cells (hPSCs), both human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSC). Both hPSCs and AT-MSCs were characterized and their EVs were extracted using standard protocols. Small non-coding RNA sequencing from EVs showed that hPSCs and AT-MSCs showed distinct profiles, unique for each stem cell source. Interestingly, in hPSCs, most abundant miRNAs were from specific miRNA families regulating pluripotency, reprogramming and differentiation (miR-17-92, mir-200, miR-302/367, miR-371/373, CM19 microRNA cluster). For the AT-MSCs, the highly expressed miRNAs were found to be regulating osteogenesis (let-7/98, miR-10/100, miR-125, miR-196, miR-199, miR-615-3p, mir-22-3p, mir-24-3p, mir-27a-3p, mir-193b-5p, mir-195-3p). Additionally, abundant small nuclear and nucleolar RNA were detected in hPSCs, whereas Y- and tRNA were found in AT-MSCs. Identification of EV-miRNA and non-coding RNA signatures released by these stem cells will provide clues towards understanding their role in intracellular communication, and well as their roles in maintaining the stem cell niche. (hide) EV-METRIC 0% (median: 14% of all experiments on the same sample type) Reported Not reported Not applicable EV-enriched proteins Protein analysis: analysis of three or more EV-enriched proteins non EV-enriched protein Protein analysis: assessment of a non-EV-enriched protein qualitative and quantitative analysis Particle analysis: implementation of both qualitative and quantitative methods. For the quantitative method, the reporting of measured EV concentration is expected. electron microscopy images Particle analysis: inclusion of a widefield and close-up electron microscopy image density gradient Separation method: density gradient, at least as validation of results attributed to EVs EV density Separation method: reporting of obtained EV density ultracentrifugation specifics Separation method: reporting of g-forces, duration and rotor type of ultracentrifugation steps antibody specifics Protein analysis: antibody clone/reference number and dilution lysate preparation Protein analysis: lysis buffer composition Study data Sample type Cell culture supernatant Sample origin Control condition Focus vesicles extracellular vesicle Separation protocol Separation protocol Gives a short, non-chronological overview of the different steps of the separation protocol. dUC = (Differential) (ultra)centrifugation DG = density gradient UF = ultrafiltration SEC = size-exclusion chromatography IAF = immuno-affinity capture miRCURY Exosome Isolation Kit (Exiqon A/S, Vedbaek, Denmark) Protein markers EV: None non-EV: None Proteomics no Show all info Study aim Identification of content (omics approaches), Technical analysis comparing/optimizing EV-related methods Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells adipose tissue-derived mesenchymal stem cells EV-harvesting Medium EV-depleted serum Preparation of EDS >=18h at >= 100,000g Cell viability (%) NA Separation Method Commercial kit miRCURY Exosome Isolation Kit (Exiqon A/S, Vedbaek, Denmark) Other Name other separation method miRCURY Exosome Isolation Kit (Exiqon A/S, Vedbaek, Denmark) Characterization: Protein analysis None Protein Concentration Method Not determined Characterization: RNA analysis Database No Proteinase treatment No RNAse treatment No Characterization: Lipid analysis No

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EV-TRACK ID	EV180022
species	Homo sapiens
sample type	Cell culture
cell type	pluripotent stem cells	pluripotent stem cells	adipose tissue-derived mesenchymal stem cells	adipose tissue-derived mesenchymal stem cells
medium	Serum free medium	Serum free medium	EV-depleted serum	EV-depleted serum
condition	Control condition	Control condition	Control condition	Control condition
separation protocol	(d)(U)C	miRCURY Exosome Isolation Kit (Exiqon A/S Vedbaek Denmark)	(d)(U)C Filtration	miRCURY Exosome Isolation Kit (Exiqon A/S Vedbaek Denmark)
Exp. nr.	1	4	3	2
EV-METRIC %	67	67	66	0