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Details	EV-TRACK ID	Experiment nr.	Species	Sample type	Separation protocol	First author	Year	EV-METRIC
	EV120110	1/2	Rattus norvegicus/rattus	NAY	(d)(U)C DG	Zech D	2012	14%
Study summary Full title Tumor-exosomes and leukocyte activation: an ambivalent crosstalk All authors Zech D, Rana S, Büchler MW, Zöller M Journal Cell Commun Signal Abstract BACKGROUND: Tumor-exosomes being reported to suppress or promote a cancer-directed immune response, (show more...)BACKGROUND: Tumor-exosomes being reported to suppress or promote a cancer-directed immune response, we used exosomes of the rat pancreatic adenocarcinoma BSp73ASML (ASML) to evaluate, whether and which steps in immune response induction can be affected by tumor-exosomes and how the impaired responsiveness can be circumvented. RESULTS: ASML-exosomes bind to and are taken up by all leukocyte subpopulations in vivo and in vitro, uptake by CD11b+ leukocytes exceeding that by T and B cells. ASML-exosomes affect leukocyte proliferation via reduced CD44v6 up-regulation and lck, ZAP70 and ERK1,2 phosphorylation, which can be compensated by dendritic cells (DC). ASML-exosomes do not support Treg. Yet, impaired activation of anti-apoptotic signals is accompanied by slightly increased apoptosis susceptibility. IgM secretion is unaffected; NK and CTL activity are strengthened, ASML-exosomes co-operating with DC in CTL activation. ASML-exosomes transiently interfere with leukocyte migration by occupying migration-promoting receptors CD44, CD49d, CD62L and CD54 during binding/internalization. CONCLUSION: ASML-exosomes might well serve as adjuvant in immunotherapy as they support leukocyte effector functions and have only a minor impact on leukocyte activation, which can be overridden by DC. However, exosome-induced modulation of immune cells relies, at least in part, on exosome uptake and message transfer. This implies that depending on the individual tumor's exosome composition, exosomes may distinctly affect the immune system. Nonetheless, whether immunotherapy can profit from using tumor-exosomes as adjuvant can easily be settled beforehand in vitro. (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 NAY Focus vesicles exosomes 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 DG Protein markers EV: non-EV: Proteomics no Show all info Study aim Function Sample Species Rattus norvegicus/rattus Sample Type Cell culture supernatant EV-harvesting Medium serum free 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 Between 100,000 g and 150,000 g Pelleting performed Yes Pelleting: time(min) 90 Density gradient Only used for validation of main results Yes Lowest density fraction 0.25 Highest density fraction 2.5 Orientation Bottom-up

	EV120110	2/2	Rattus norvegicus/rattus	NAY	(d)(U)C Filtration	Zech D	2012	0%
Study summary Full title Tumor-exosomes and leukocyte activation: an ambivalent crosstalk All authors Zech D, Rana S, Büchler MW, Zöller M Journal Cell Commun Signal Abstract BACKGROUND: Tumor-exosomes being reported to suppress or promote a cancer-directed immune response, (show more...)BACKGROUND: Tumor-exosomes being reported to suppress or promote a cancer-directed immune response, we used exosomes of the rat pancreatic adenocarcinoma BSp73ASML (ASML) to evaluate, whether and which steps in immune response induction can be affected by tumor-exosomes and how the impaired responsiveness can be circumvented. RESULTS: ASML-exosomes bind to and are taken up by all leukocyte subpopulations in vivo and in vitro, uptake by CD11b+ leukocytes exceeding that by T and B cells. ASML-exosomes affect leukocyte proliferation via reduced CD44v6 up-regulation and lck, ZAP70 and ERK1,2 phosphorylation, which can be compensated by dendritic cells (DC). ASML-exosomes do not support Treg. Yet, impaired activation of anti-apoptotic signals is accompanied by slightly increased apoptosis susceptibility. IgM secretion is unaffected; NK and CTL activity are strengthened, ASML-exosomes co-operating with DC in CTL activation. ASML-exosomes transiently interfere with leukocyte migration by occupying migration-promoting receptors CD44, CD49d, CD62L and CD54 during binding/internalization. CONCLUSION: ASML-exosomes might well serve as adjuvant in immunotherapy as they support leukocyte effector functions and have only a minor impact on leukocyte activation, which can be overridden by DC. However, exosome-induced modulation of immune cells relies, at least in part, on exosome uptake and message transfer. This implies that depending on the individual tumor's exosome composition, exosomes may distinctly affect the immune system. Nonetheless, whether immunotherapy can profit from using tumor-exosomes as adjuvant can easily be settled beforehand in vitro. (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 NAY Focus vesicles exosomes 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 Sample Species Rattus norvegicus/rattus Sample Type Cell culture supernatant EV-harvesting Medium serum free 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 Between 100,000 g and 150,000 g Pelleting performed Yes Pelleting: time(min) 90 Filtration steps 0.22µm or 0.2µm

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EV-TRACK ID	EV120110
species	Rattus norvegicus/rattus
sample type	Cell culture
cell type	NAY
condition	NAY
separation protocol	(d)(U)C DG	(d)(U)C Filtration
Exp. nr.	1	2
EV-METRIC %	14	0