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Details	EV-TRACK ID	Experiment nr.	Species	Sample type	Separation protocol	First author	Year	EV-METRIC
	EV160016	1/3	Homo sapiens	DKO-1	(d)(U)C UF Filtration	Cha DJ	2015	44%
Study summary Full title KRAS-dependent sorting of miRNA to exosomes All authors Cha DJ, Franklin JL, Dou Y, Liu Q, Higginbotham JN, Demory Beckler M, Weaver AM, Vickers K, Prasad N, Levy S, Zhang B, Coffey RJ, Patton JG. Journal Elife Abstract Mutant KRAS colorectal cancer (CRC) cells release protein-laden exosomes that can alter the tumor mi (show more...)Mutant KRAS colorectal cancer (CRC) cells release protein-laden exosomes that can alter the tumor microenvironment. To test whether exosomal RNAs also contribute to changes in gene expression in recipient cells, and whether mutant KRAS might regulate the... (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 exosome 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 UF Filtration Protein markers EV: "Flotillin1/ TSG101/ HSP70/ KRAS/ EGFR/ RAP1/ SRC/ LYN/ ITGB1/ ITGA2/ ITGAV/ ITGA6/ ITGB4/ EPHA2/ EPS8/ CTTN" non-EV: VDAC Proteomics yes Show all info Study aim Identification of content (omics approaches) Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells DKO-1 EV-harvesting Medium Serum free medium Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Below or equal to 800 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: time(min) 120 Pelleting: rotor type Not specified Pelleting: speed (g) 150000 Wash: time (min) 180 Wash: Rotor Type Not specified Wash: speed (g) 150000 Filtration steps 0.22µm or 0.2µm Ultra filtration Cut-off size (kDa) 100 Membrane type Not specified Characterization: Protein analysis Protein Concentration Method microBCA Western Blot Detected EV-associated proteins "Flotillin1/ TSG101/ HSP70/ KRAS/ EGFR/ RAP1/ SRC/ LYN/ ITGB1/ ITGA2/ ITGAV/ ITGA6/ ITGB4/ EPHA2/ EPS8/ CTTN" Not detected contaminants VDAC Proteomics database No Characterization: RNA analysis RNA analysis Type "(RT)(q)PCR;RNA sequencing" Database No Proteinase treatment No RNAse treatment No Characterization: Lipid analysis No Characterization: Particle analysis NTA Report type Not Reported EV concentration Yes EM EM-type Transmission-EM Image type Wide-field Report size (nm) 56.3

	EV160016	3/3	Homo sapiens	DKs-8	(d)(U)C UF Filtration	Cha DJ	2015	44%
Study summary Full title KRAS-dependent sorting of miRNA to exosomes All authors Cha DJ, Franklin JL, Dou Y, Liu Q, Higginbotham JN, Demory Beckler M, Weaver AM, Vickers K, Prasad N, Levy S, Zhang B, Coffey RJ, Patton JG. Journal Elife Abstract Mutant KRAS colorectal cancer (CRC) cells release protein-laden exosomes that can alter the tumor mi (show more...)Mutant KRAS colorectal cancer (CRC) cells release protein-laden exosomes that can alter the tumor microenvironment. To test whether exosomal RNAs also contribute to changes in gene expression in recipient cells, and whether mutant KRAS might regulate the... (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 exosome 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 UF Filtration Protein markers EV: "Flotillin1/ TSG101/ HSP70/ EGFR/ RAP1/ SRC/ ITGB1/ ITGA2/ ITGAV/ CTNNA" non-EV: VDAC Proteomics yes Show all info Study aim Identification of content (omics approaches) Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells DKs-8 EV-harvesting Medium Serum free medium Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Below or equal to 800 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: time(min) 120 Pelleting: rotor type Not specified Pelleting: speed (g) 150000 Wash: time (min) 180 Wash: Rotor Type Not specified Wash: speed (g) 150000 Filtration steps 0.22µm or 0.2µm Ultra filtration Cut-off size (kDa) 100 Membrane type Not specified Characterization: Protein analysis Protein Concentration Method microBCA Western Blot Detected EV-associated proteins "Flotillin1/ TSG101/ HSP70/ EGFR/ RAP1/ SRC/ ITGB1/ ITGA2/ ITGAV/ CTNNA" Not detected contaminants VDAC Proteomics database No Characterization: RNA analysis RNA analysis Type "(RT)(q)PCR;RNA sequencing" Database No Proteinase treatment No RNAse treatment No Characterization: Lipid analysis No Characterization: Particle analysis NTA Report type Not Reported EV concentration Yes EM EM-type Transmission-EM Image type Wide-field Report size (nm) 59.2

	EV160016	2/3	Homo sapiens	DLD-1	(d)(U)C UF Filtration	Cha DJ	2015	33%
Study summary Full title KRAS-dependent sorting of miRNA to exosomes All authors Cha DJ, Franklin JL, Dou Y, Liu Q, Higginbotham JN, Demory Beckler M, Weaver AM, Vickers K, Prasad N, Levy S, Zhang B, Coffey RJ, Patton JG. Journal Elife Abstract Mutant KRAS colorectal cancer (CRC) cells release protein-laden exosomes that can alter the tumor mi (show more...)Mutant KRAS colorectal cancer (CRC) cells release protein-laden exosomes that can alter the tumor microenvironment. To test whether exosomal RNAs also contribute to changes in gene expression in recipient cells, and whether mutant KRAS might regulate the... (hide) EV-METRIC 33% (74th 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 exosome 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 UF Filtration Protein markers EV: "Flotillin1/ TSG101/ HSP70/ KRAS/ EGFR/ RAP1/ SRC/ LYN/ ITGB1/ ITGA2/ ITGAV/ ITGB4/ EPHA2/ EPS8/ CTNNA" non-EV: VDAC Proteomics yes Show all info Study aim Identification of content (omics approaches) Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells DLD-1 EV-harvesting Medium Serum free medium Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Below or equal to 800 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: time(min) 120 Pelleting: rotor type Not specified Pelleting: speed (g) 150000 Wash: time (min) 180 Wash: Rotor Type Not specified Wash: speed (g) 150000 Filtration steps 0.22µm or 0.2µm Ultra filtration Cut-off size (kDa) 100 Membrane type Not specified Characterization: Protein analysis Protein Concentration Method microBCA Western Blot Detected EV-associated proteins "Flotillin1/ TSG101/ HSP70/ KRAS/ EGFR/ RAP1/ SRC/ LYN/ ITGB1/ ITGA2/ ITGAV/ ITGB4/ EPHA2/ EPS8/ CTNNA" Not detected contaminants VDAC Proteomics database No Characterization: RNA analysis RNA analysis Type "(RT)(q)PCR;RNA sequencing" Database No Proteinase treatment No RNAse treatment No Characterization: Lipid analysis No Characterization: Particle analysis NTA Report type Not Reported EV concentration Yes

	EV160016	1/3	Homo sapiens	K562	(d)(U)C Filtration	Zhang HM	2016	0%
Study summary Full title miR-146b-5p within BCR-ABL1-Positive Microvesicles Promotes Leukemic Transformation of Hematopoietic Cells. All authors Zhang HM, Li Q, Zhu X, Liu W, Hu H, Liu T, Cheng F, You Y, Zhong Z, Zou P, Li Q, Chen Z, Guo AY. Journal Cancer Res Abstract Evidence is accumulating that extracellular microvesicles (MV) facilitate progression and relapse in (show more...)Evidence is accumulating that extracellular microvesicles (MV) facilitate progression and relapse in cancer. Using a model in which MVs derived from K562 chronic myelogenous leukemia (CML) cells transform normal hematopoietic transplants into leukemia-like cells, we defined the underlying mechanisms of this process through gene-expression studies and network analyses of transcription factors (TF) and miRNAs. We found that antitumor miRNAs were increased and several defense pathways were initiated during the early phases of oncogenic transformation. Later, oncomiRs and genes involved in cell cycle, DNA repair, and energy metabolism pathways were upregulated. Regulatory network analyses revealed that a number of TFs and miRNAs were responsible for the pathway dysregulation and the oncogenic transformation. In particular, we found that miR-146b-5p, which was highly expressed in MVs, coordinated the regulation of cancer-related genes to promote cell-transforming processes. Notably, treatment of recipient cells with MV derived from K562 cells expressing mimics of miR-146b-5p revealed that it accelerated the transformation process in large part by silencing the tumor-suppressor NUMB High levels of miR-146b-5p also enhanced reactive oxygen species levels and genome instability of recipient cells. Taken together, our finding showed how upregulation of oncogenic miRNAs in MVs promote hematopoetic cells to a leukemic state, as well as a demonstration for TF and miRNA coregulatory analysis in exploring the dysregulation of cancers and discovering key factors. (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 (shedding) microvesicle 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 Protein markers EV: non-EV: Proteomics no Show all info Study aim Function/Identification of content (omics approaches) Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells K562 EV-harvesting Medium Serum-containing medium 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: time(min) 60 Pelleting: rotor type Not specified Pelleting: speed (g) 13000 Filtration steps > 0.45 µm, Characterization: Protein analysis None Protein Concentration Method Not determined Characterization: RNA analysis RNA analysis Type (RT)(q)PCR;RNA sequencing Database No Proteinase treatment No RNAse treatment No Characterization: Lipid analysis No Characterization: Particle analysis None

	EV160016	2/3	Homo sapiens	K562	(d)(U)C Filtration	Zhang HM	2016	0%
Study summary Full title miR-146b-5p within BCR-ABL1-Positive Microvesicles Promotes Leukemic Transformation of Hematopoietic Cells. All authors Zhang HM, Li Q, Zhu X, Liu W, Hu H, Liu T, Cheng F, You Y, Zhong Z, Zou P, Li Q, Chen Z, Guo AY. Journal Cancer Res Abstract Evidence is accumulating that extracellular microvesicles (MV) facilitate progression and relapse in (show more...)Evidence is accumulating that extracellular microvesicles (MV) facilitate progression and relapse in cancer. Using a model in which MVs derived from K562 chronic myelogenous leukemia (CML) cells transform normal hematopoietic transplants into leukemia-like cells, we defined the underlying mechanisms of this process through gene-expression studies and network analyses of transcription factors (TF) and miRNAs. We found that antitumor miRNAs were increased and several defense pathways were initiated during the early phases of oncogenic transformation. Later, oncomiRs and genes involved in cell cycle, DNA repair, and energy metabolism pathways were upregulated. Regulatory network analyses revealed that a number of TFs and miRNAs were responsible for the pathway dysregulation and the oncogenic transformation. In particular, we found that miR-146b-5p, which was highly expressed in MVs, coordinated the regulation of cancer-related genes to promote cell-transforming processes. Notably, treatment of recipient cells with MV derived from K562 cells expressing mimics of miR-146b-5p revealed that it accelerated the transformation process in large part by silencing the tumor-suppressor NUMB High levels of miR-146b-5p also enhanced reactive oxygen species levels and genome instability of recipient cells. Taken together, our finding showed how upregulation of oncogenic miRNAs in MVs promote hematopoetic cells to a leukemic state, as well as a demonstration for TF and miRNA coregulatory analysis in exploring the dysregulation of cancers and discovering key factors. (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 miR-146b-5p mimics transfected Focus vesicles (shedding) microvesicle 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 Protein markers EV: non-EV: Proteomics no Show all info Study aim Function/Identification of content (omics approaches) Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells K562 EV-harvesting Medium Serum-containing medium 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: time(min) 60 Pelleting: rotor type Not specified Pelleting: speed (g) 13000 Filtration steps > 0.45 µm, Characterization: Protein analysis None Protein Concentration Method Not determined Characterization: RNA analysis RNA analysis Type (RT)(q)PCR Database No Proteinase treatment No RNAse treatment No Characterization: Lipid analysis No Characterization: Particle analysis None

	EV160016	3/3	Homo sapiens	K562	(d)(U)C Filtration	Zhang HM	2016	0%
Study summary Full title miR-146b-5p within BCR-ABL1-Positive Microvesicles Promotes Leukemic Transformation of Hematopoietic Cells. All authors Zhang HM, Li Q, Zhu X, Liu W, Hu H, Liu T, Cheng F, You Y, Zhong Z, Zou P, Li Q, Chen Z, Guo AY. Journal Cancer Res Abstract Evidence is accumulating that extracellular microvesicles (MV) facilitate progression and relapse in (show more...)Evidence is accumulating that extracellular microvesicles (MV) facilitate progression and relapse in cancer. Using a model in which MVs derived from K562 chronic myelogenous leukemia (CML) cells transform normal hematopoietic transplants into leukemia-like cells, we defined the underlying mechanisms of this process through gene-expression studies and network analyses of transcription factors (TF) and miRNAs. We found that antitumor miRNAs were increased and several defense pathways were initiated during the early phases of oncogenic transformation. Later, oncomiRs and genes involved in cell cycle, DNA repair, and energy metabolism pathways were upregulated. Regulatory network analyses revealed that a number of TFs and miRNAs were responsible for the pathway dysregulation and the oncogenic transformation. In particular, we found that miR-146b-5p, which was highly expressed in MVs, coordinated the regulation of cancer-related genes to promote cell-transforming processes. Notably, treatment of recipient cells with MV derived from K562 cells expressing mimics of miR-146b-5p revealed that it accelerated the transformation process in large part by silencing the tumor-suppressor NUMB High levels of miR-146b-5p also enhanced reactive oxygen species levels and genome instability of recipient cells. Taken together, our finding showed how upregulation of oncogenic miRNAs in MVs promote hematopoetic cells to a leukemic state, as well as a demonstration for TF and miRNA coregulatory analysis in exploring the dysregulation of cancers and discovering key factors. (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 miR-146b-5p inhibitor transfected Focus vesicles (shedding) microvesicle 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 Protein markers EV: non-EV: Proteomics no Show all info Study aim Function/Identification of content (omics approaches) Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells K562 EV-harvesting Medium Serum-containing medium 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: time(min) 60 Pelleting: rotor type Not specified Pelleting: speed (g) 13000 Filtration steps > 0.45 µm, Characterization: Protein analysis None Protein Concentration Method Not determined Characterization: RNA analysis RNA analysis Type (RT)(q)PCR Database No Proteinase treatment No RNAse treatment No Characterization: Lipid analysis No Characterization: Particle analysis None

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