EV-TRACK

Search > Results

You searched for: EV200014 (EV-TRACK ID)

Showing 1 - 4 of 4

Results list Study Tree

Details	EV-TRACK ID	Experiment nr.	Species	Sample type	Separation protocol	First author	Year	EV-METRIC
	EV200014	1/4	Homo sapiens	IMR90 ER:STOP	(d)(U)C qEV	Wallis, Ryan	2021	56%
Study summary Full title Isolation methodology is essential to the evaluation of the extracellular vesicle component of the senescence‐associated secretory phenotype All authors Ryan Wallis, Natasa Josipovic, Hannah Mizen, Arturo Robles‐Tenorio, Eleanor J. Tyler, Argyris Papantonis, Cleo L. Bishop Journal J Extracell Vesicles Abstract A hallmark of senescence is the acquisition of an enhanced secretome comprising inflammatory mediato (show more...)A hallmark of senescence is the acquisition of an enhanced secretome comprising inflammatory mediators and tissue remodelling agents – the senescence‐associated secretory phenotype (SASP). Through the SASP, senescent cells are hypothesised to contribute to both ageing and pathologies associated with age. Whilst soluble factors have been the most widely investigated components of the SASP, there is growing evidence that small extracellular vesicles (EVs) comprise functionally important constituents. Thus, dissecting the contribution of the soluble SASP from the vesicular component is crucial to elucidating the functional significance of senescent cell derived EVs. Here, we take advantage of a systematic proteomics based approach to determine that soluble SASP factors co‐isolate with EVs following differential ultracentrifugation (dUC). We present size‐exclusion chromatography (SEC) as a method for separation of the soluble and vesicular components of the senescent secretome and thus EV purification. Furthermore, we demonstrate that SEC EVs isolated from senescent cells contribute to non‐cell autonomous paracrine senescence. Therefore, this work emphasises the requirement for methodological rigor due to the propensity of SASP components to co‐isolate during dUC and provides a framework for future investigations of the vesicular component of the SASP. (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 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 Commercial method Protein markers EV: TSG101/ CD9 non-EV: Calnexin Proteomics no Show all info Study aim Function/Identification of content (omics approaches)/Technical analysis comparing/optimizing EV-related methods Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells IMR90 ER:STOP EV-harvesting Medium EV-depleted medium Preparation of EDS Commercial EDS Cell count 6.20E+06 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Between 100,000 g and 150,000 g Pelleting performed Yes Pelleting: time(min) 90 Pelleting: rotor type T-865 Pelleting: speed (g) 99582 Commercial kit qEV Characterization: Protein analysis PMID previous EV protein analysis Protein Concentration Method microBCA Western Blot Antibody details provided? Yes Antibody dilution provided? Yes Lysis buffer provided? Yes Not detected EV-associated proteins TSG101/ CD9 Not detected contaminants Calnexin Characterization: Lipid analysis No Characterization: Particle analysis PMID previous EV particle analysis Extra particle analysis NTA Report type Modus Reported size (nm) 92 EV concentration Yes Particle yield total concentration;Yes, other: 1.60E+10

	EV200014	2/4	Homo sapiens	IMR90 ER:STOP	(d)(U)C	Wallis, Ryan	2021	56%
Study summary Full title Isolation methodology is essential to the evaluation of the extracellular vesicle component of the senescence‐associated secretory phenotype All authors Ryan Wallis, Natasa Josipovic, Hannah Mizen, Arturo Robles‐Tenorio, Eleanor J. Tyler, Argyris Papantonis, Cleo L. Bishop Journal J Extracell Vesicles Abstract A hallmark of senescence is the acquisition of an enhanced secretome comprising inflammatory mediato (show more...)A hallmark of senescence is the acquisition of an enhanced secretome comprising inflammatory mediators and tissue remodelling agents – the senescence‐associated secretory phenotype (SASP). Through the SASP, senescent cells are hypothesised to contribute to both ageing and pathologies associated with age. Whilst soluble factors have been the most widely investigated components of the SASP, there is growing evidence that small extracellular vesicles (EVs) comprise functionally important constituents. Thus, dissecting the contribution of the soluble SASP from the vesicular component is crucial to elucidating the functional significance of senescent cell derived EVs. Here, we take advantage of a systematic proteomics based approach to determine that soluble SASP factors co‐isolate with EVs following differential ultracentrifugation (dUC). We present size‐exclusion chromatography (SEC) as a method for separation of the soluble and vesicular components of the senescent secretome and thus EV purification. Furthermore, we demonstrate that SEC EVs isolated from senescent cells contribute to non‐cell autonomous paracrine senescence. Therefore, this work emphasises the requirement for methodological rigor due to the propensity of SASP components to co‐isolate during dUC and provides a framework for future investigations of the vesicular component of the SASP. (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 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 Protein markers EV: TSG101/ IL-8/ CD9 non-EV: Calnexin Proteomics yes Show all info Study aim Function/Identification of content (omics approaches)/Technical analysis comparing/optimizing EV-related methods Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells IMR90 ER:STOP EV-harvesting Medium EV-depleted medium Preparation of EDS Commercial EDS Cell count 6.20E+06 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Between 100,000 g and 150,000 g Pelleting performed Yes Pelleting: time(min) 90 Pelleting: rotor type T-865 Pelleting: speed (g) 99582 Characterization: Protein analysis PMID previous EV protein analysis Protein Concentration Method Not determined Western Blot Antibody details provided? Yes Antibody dilution provided? Yes Lysis buffer provided? Yes Detected EV-associated proteins CD9/ TSG101 Detected contaminants Calnexin ELISA Antibody details provided? No Detected EV-associated proteins IL-8 Proteomics database No Characterization: Lipid analysis No Characterization: Particle analysis PMID previous EV particle analysis Extra particle analysis NTA Report type Modus Reported size (nm) 85.2 EV concentration Yes Particle yield total concentration;Yes, other: 2.20E+11

	EV200014	3/4	Homo sapiens	IMR90 ER:RAS	(d)(U)C qEV	Wallis, Ryan	2021	56%
Study summary Full title Isolation methodology is essential to the evaluation of the extracellular vesicle component of the senescence‐associated secretory phenotype All authors Ryan Wallis, Natasa Josipovic, Hannah Mizen, Arturo Robles‐Tenorio, Eleanor J. Tyler, Argyris Papantonis, Cleo L. Bishop Journal J Extracell Vesicles Abstract A hallmark of senescence is the acquisition of an enhanced secretome comprising inflammatory mediato (show more...)A hallmark of senescence is the acquisition of an enhanced secretome comprising inflammatory mediators and tissue remodelling agents – the senescence‐associated secretory phenotype (SASP). Through the SASP, senescent cells are hypothesised to contribute to both ageing and pathologies associated with age. Whilst soluble factors have been the most widely investigated components of the SASP, there is growing evidence that small extracellular vesicles (EVs) comprise functionally important constituents. Thus, dissecting the contribution of the soluble SASP from the vesicular component is crucial to elucidating the functional significance of senescent cell derived EVs. Here, we take advantage of a systematic proteomics based approach to determine that soluble SASP factors co‐isolate with EVs following differential ultracentrifugation (dUC). We present size‐exclusion chromatography (SEC) as a method for separation of the soluble and vesicular components of the senescent secretome and thus EV purification. Furthermore, we demonstrate that SEC EVs isolated from senescent cells contribute to non‐cell autonomous paracrine senescence. Therefore, this work emphasises the requirement for methodological rigor due to the propensity of SASP components to co‐isolate during dUC and provides a framework for future investigations of the vesicular component of the SASP. (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 HRas:ER Oncogene Induced Senescence Model 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 Commercial method Protein markers EV: TSG101/ IL-8/ CD9 non-EV: Calnexin Proteomics yes Show all info Study aim Function/Identification of content (omics approaches)/Technical analysis comparing/optimizing EV-related methods Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells IMR90 ER:RAS EV-harvesting Medium EV-depleted medium Preparation of EDS Commercial EDS Cell count 4.40E+06 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Between 100,000 g and 150,000 g Pelleting performed Yes Pelleting: time(min) 90 Pelleting: rotor type T-865 Pelleting: speed (g) 99582 Commercial kit qEV Characterization: Protein analysis PMID previous EV protein analysis Protein Concentration Method microBCA Western Blot Antibody details provided? Yes Antibody dilution provided? Yes Lysis buffer provided? Yes Detected EV-associated proteins CD9/ TSG101 Not detected contaminants Calnexin ELISA Antibody details provided? No Detected EV-associated proteins IL-8 Proteomics database No Characterization: Lipid analysis No Characterization: Particle analysis PMID previous EV particle analysis Extra particle analysis NTA Report type Modus Reported size (nm) 107 EV concentration Yes Particle yield total concentration;Yes, other: 8.80E+10

	EV200014	4/4	Homo sapiens	IMR90 ER:RAS	(d)(U)C	Wallis, Ryan	2021	56%
Study summary Full title Isolation methodology is essential to the evaluation of the extracellular vesicle component of the senescence‐associated secretory phenotype All authors Ryan Wallis, Natasa Josipovic, Hannah Mizen, Arturo Robles‐Tenorio, Eleanor J. Tyler, Argyris Papantonis, Cleo L. Bishop Journal J Extracell Vesicles Abstract A hallmark of senescence is the acquisition of an enhanced secretome comprising inflammatory mediato (show more...)A hallmark of senescence is the acquisition of an enhanced secretome comprising inflammatory mediators and tissue remodelling agents – the senescence‐associated secretory phenotype (SASP). Through the SASP, senescent cells are hypothesised to contribute to both ageing and pathologies associated with age. Whilst soluble factors have been the most widely investigated components of the SASP, there is growing evidence that small extracellular vesicles (EVs) comprise functionally important constituents. Thus, dissecting the contribution of the soluble SASP from the vesicular component is crucial to elucidating the functional significance of senescent cell derived EVs. Here, we take advantage of a systematic proteomics based approach to determine that soluble SASP factors co‐isolate with EVs following differential ultracentrifugation (dUC). We present size‐exclusion chromatography (SEC) as a method for separation of the soluble and vesicular components of the senescent secretome and thus EV purification. Furthermore, we demonstrate that SEC EVs isolated from senescent cells contribute to non‐cell autonomous paracrine senescence. Therefore, this work emphasises the requirement for methodological rigor due to the propensity of SASP components to co‐isolate during dUC and provides a framework for future investigations of the vesicular component of the SASP. (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 HRas:ER Oncogene Induced Senescence Model 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 Protein markers EV: TSG101/ IL-8/ CD9 non-EV: Calnexin Proteomics yes Show all info Study aim Function/Identification of content (omics approaches)/Technical analysis comparing/optimizing EV-related methods Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells IMR90 ER:RAS EV-harvesting Medium EV-depleted medium Preparation of EDS Commercial EDS Cell count 4.40E+06 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Between 100,000 g and 150,000 g Pelleting performed Yes Pelleting: time(min) 90 Pelleting: rotor type T-865 Pelleting: speed (g) 99582 Characterization: Protein analysis PMID previous EV protein analysis Protein Concentration Method Not determined Western Blot Antibody details provided? Yes Antibody dilution provided? Yes Lysis buffer provided? Yes Detected EV-associated proteins CD9/ TSG101 Detected contaminants Calnexin ELISA Antibody details provided? No Detected EV-associated proteins IL-8 Proteomics database No Characterization: Lipid analysis No Characterization: Particle analysis PMID previous EV particle analysis Extra particle analysis NTA Report type Modus Reported size (nm) 102 EV concentration Yes Particle yield total concentration;Yes, other: 4.50E+11

				1 - 4 of 4

EV-TRACK ID	EV200014
species	Homo sapiens
sample type	Cell culture
cell type	IMR90 ER:STOP	IMR90 ER:STOP	IMR90 ER:RAS	IMR90 ER:RAS
condition	Control condition	Control condition	HRas:ER Oncogene Induced Senescence Model	HRas:ER Oncogene Induced Senescence Model
separation protocol	(d)(U)C qEV	(d)(U)C	(d)(U)C qEV	(d)(U)C
Exp. nr.	1	2	3	4
EV-METRIC %	56	56	56	56