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Details	EV-TRACK ID	Experiment nr.	Species	Sample type	Separation protocol	First author	Year	EV-METRIC
	EV110038	3/3	Rattus norvegicus	NAY	(d)(U)C DG	Fitzner D	2011	44%
Study summary Full title Selective transfer of exosomes from oligodendrocytes to microglia by macropinocytosis All authors Fitzner D, Schnaars M, van Rossum D, Krishnamoorthy G, Dibaj P, Bakhti M, Regen T, Hanisch UK, Simons M Journal J Cell Sci Abstract The transfer of antigens from oligodendrocytes to immune cells has been implicated in the pathogenes (show more...)The transfer of antigens from oligodendrocytes to immune cells has been implicated in the pathogenesis of autoimmune diseases. Here, we show that oligodendrocytes secrete small membrane vesicles called exosomes, which are specifically and efficiently taken up by microglia both in vitro and in vivo. Internalisation of exosomes occurs by a macropinocytotic mechanism without inducing a concomitant inflammatory response. After stimulation of microglia with interferon-?, we observe an upregulation of MHC class II in a subpopulation of microglia. However, exosomes are preferentially internalised in microglia that do not seem to have antigen-presenting capacity. We propose that the constitutive macropinocytotic clearance of exosomes by a subset of microglia represents an important mechanism through which microglia participate in the degradation of oligodendroglial membrane in an immunologically 'silent' manner. By designating the capacity for macropinocytosis and antigen presentation to distinct cells, degradation and immune function might be assigned to different subtypes of microglia. (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 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: Alix/ TSG101/ PLP/ MOG/ Flotillin2 non-EV: Cell organelle protein Proteomics no EV density (g/ml) 1.150 Show all info Study aim Biogenesis/Sorting Sample Species Rattus norvegicus Sample Type Cell culture supernatant EV-harvesting Medium serum free 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) 60 Density gradient Only used for validation of main results Yes Lowest density fraction 1.03g/cm3 Highest density fraction 1.32g/cm3 Orientation Top-down Speed (g) 100000 Characterization: Protein analysis Western Blot Detected EV-associated proteins Alix/ TSG101/ Flotillin2/ PLP/ MOG Detected contaminants Cell organelle protein ELISA Detected EV-associated proteins Flotillin2/ PLP/ MOG

	EV110038	2/3	Rattus norvegicus	NAY	(d)(U)C	Fitzner D	2011	22%
Study summary Full title Selective transfer of exosomes from oligodendrocytes to microglia by macropinocytosis All authors Fitzner D, Schnaars M, van Rossum D, Krishnamoorthy G, Dibaj P, Bakhti M, Regen T, Hanisch UK, Simons M Journal J Cell Sci Abstract The transfer of antigens from oligodendrocytes to immune cells has been implicated in the pathogenes (show more...)The transfer of antigens from oligodendrocytes to immune cells has been implicated in the pathogenesis of autoimmune diseases. Here, we show that oligodendrocytes secrete small membrane vesicles called exosomes, which are specifically and efficiently taken up by microglia both in vitro and in vivo. Internalisation of exosomes occurs by a macropinocytotic mechanism without inducing a concomitant inflammatory response. After stimulation of microglia with interferon-?, we observe an upregulation of MHC class II in a subpopulation of microglia. However, exosomes are preferentially internalised in microglia that do not seem to have antigen-presenting capacity. We propose that the constitutive macropinocytotic clearance of exosomes by a subset of microglia represents an important mechanism through which microglia participate in the degradation of oligodendroglial membrane in an immunologically 'silent' manner. By designating the capacity for macropinocytosis and antigen presentation to distinct cells, degradation and immune function might be assigned to different subtypes of microglia. (hide) EV-METRIC 22% (58th 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 Protein markers EV: Alix/ MOG/ Flotillin2 non-EV: Cell organelle protein Proteomics no Show all info Study aim Biogenesis/Sorting Sample Species Rattus norvegicus Sample Type Cell culture supernatant EV-harvesting Medium serum free 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) 60 Characterization: Protein analysis Western Blot Detected EV-associated proteins Alix/ Flotillin2/ MOG Detected contaminants Cell organelle protein ELISA Detected EV-associated proteins Flotillin2/ MOG Characterization: Particle analysis EM EM-type immune EM EM protein Phosphatidylserine Image type Close-up

	EV110038	1/3	Rattus norvegicus	NAY	(d)(U)C	Fitzner D	2011	0%
Study summary Full title Selective transfer of exosomes from oligodendrocytes to microglia by macropinocytosis All authors Fitzner D, Schnaars M, van Rossum D, Krishnamoorthy G, Dibaj P, Bakhti M, Regen T, Hanisch UK, Simons M Journal J Cell Sci Abstract The transfer of antigens from oligodendrocytes to immune cells has been implicated in the pathogenes (show more...)The transfer of antigens from oligodendrocytes to immune cells has been implicated in the pathogenesis of autoimmune diseases. Here, we show that oligodendrocytes secrete small membrane vesicles called exosomes, which are specifically and efficiently taken up by microglia both in vitro and in vivo. Internalisation of exosomes occurs by a macropinocytotic mechanism without inducing a concomitant inflammatory response. After stimulation of microglia with interferon-?, we observe an upregulation of MHC class II in a subpopulation of microglia. However, exosomes are preferentially internalised in microglia that do not seem to have antigen-presenting capacity. We propose that the constitutive macropinocytotic clearance of exosomes by a subset of microglia represents an important mechanism through which microglia participate in the degradation of oligodendroglial membrane in an immunologically 'silent' manner. By designating the capacity for macropinocytosis and antigen presentation to distinct cells, degradation and immune function might be assigned to different subtypes of microglia. (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 Protein markers EV: Phosphatidylserine non-EV: Proteomics no Show all info Study aim Biogenesis/Sorting Sample Species Rattus norvegicus Sample Type Cell culture supernatant EV-harvesting Medium serum free 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) 60 Western Blot Detected EV-associated proteins Phosphatidylserine ELISA Detected EV-associated proteins Phosphatidylserine Flow cytometry specific beads Selected surface protein(s) Yes Characterization: Particle analysis EM EM-type immune EM EM protein Phosphatidylserine Image type Close-up

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EV-TRACK ID	EV110038
species	Rattus norvegicus
sample type	Cell culture
cell type	NAY
condition	NAY
separation protocol	(d)(U)C DG	(d)(U)C	(d)(U)C
Exp. nr.	3	2	1
EV-METRIC %	44	22	0