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Details	EV-TRACK ID	Experiment nr.	Species	Sample type	Separation protocol	First author	Year	EV-METRIC
	EV220053	1/7	Homo sapiens	HUVEC	DG Filtration dUC	Zhu X	2015	33%
Study summary Full title IFITM3-containing exosome as a novel mediator for anti-viral response in dengue virus infection. All authors Zhu X, He Z, Yuan J, Wen W, Huang X, Hu Y, Lin C, Pan J, Li R, Deng H, Liao S, Zhou R, Wu J, Li J, Li M Journal Cell Microbiol Abstract Interferon-inducible transmembrane proteins 1, 2 and 3 (IFITM1, IFITM2 and IFITM3) have recently bee (show more...)Interferon-inducible transmembrane proteins 1, 2 and 3 (IFITM1, IFITM2 and IFITM3) have recently been identified as potent antiviral effectors that function to suppress the entry of a broad range of enveloped viruses and modulate cellular tropism independent of viral receptor expression. However, the antiviral effect and mechanisms of IFITMs in response to viral infections remain incompletely understood and characterized. In this work, we focused our investigation on the function of the extracellular IFITM3 protein. In cell models of DENV-2 infection, we found that IFITM3 contributed to both the baseline and interferon-induced inhibition of DENV entry. Most importantly, our study for the first time demonstrated the presence of IFITM-containing exosome in the extracellular environment, and identified an ability of cellular exosome to intercellularly deliver IFITM3 and thus transmit its antiviral effect from infected to non-infected cells. Thus, our findings provide new insights in the basic mechanisms underlying the actions of IFITM3, which might lead to future development of exosome-mediated anti-viral strategies using IFITM3 as a therapeutic agent. Conceivably, variations in the basal and inducible levels of IFITMs, as well as in intracellular and extracellular levels of IFITMs, might predict the severity of dengue virus infections among individuals or across species. (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 DG Filtration dUC Protein markers EV: IFITM3/ CD63/ Flotillin2 non-EV: Calnexin/ GM130 Proteomics no Show all info Study aim Function Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells HUVEC EV-harvesting Medium EV-depleted medium Preparation of EDS overnight (16h) at >=100,000g Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: speed (g) 150000 Density gradient Type Discontinuous Number of initial discontinuous layers 3 Lowest density fraction 0.25M Highest density fraction 2.5M Sample volume (mL) 11 Orientation Bottom-up Speed (g) 150000 Duration (min) 960 Fraction processing Centrifugation Pelleting: duration (min) 90 Pelleting: speed (g) 150000 Pelleting-wash: duration (min) 90 Filtration steps 0.22µm or 0.2µm Other Name other separation method dUC Characterization: Protein analysis Protein Concentration Method BCA Western Blot Detected EV-associated proteins CD63/ IFITM3/ Flotillin2 Not detected contaminants Calnexin/ GM130 Characterization: Lipid analysis No EM EM-type Transmission-EM Image type Close-up

	EV220053	2/7	Homo sapiens	HUVEC	DG Filtration dUC	Zhu X	2015	33%
Study summary Full title IFITM3-containing exosome as a novel mediator for anti-viral response in dengue virus infection. All authors Zhu X, He Z, Yuan J, Wen W, Huang X, Hu Y, Lin C, Pan J, Li R, Deng H, Liao S, Zhou R, Wu J, Li J, Li M Journal Cell Microbiol Abstract Interferon-inducible transmembrane proteins 1, 2 and 3 (IFITM1, IFITM2 and IFITM3) have recently bee (show more...)Interferon-inducible transmembrane proteins 1, 2 and 3 (IFITM1, IFITM2 and IFITM3) have recently been identified as potent antiviral effectors that function to suppress the entry of a broad range of enveloped viruses and modulate cellular tropism independent of viral receptor expression. However, the antiviral effect and mechanisms of IFITMs in response to viral infections remain incompletely understood and characterized. In this work, we focused our investigation on the function of the extracellular IFITM3 protein. In cell models of DENV-2 infection, we found that IFITM3 contributed to both the baseline and interferon-induced inhibition of DENV entry. Most importantly, our study for the first time demonstrated the presence of IFITM-containing exosome in the extracellular environment, and identified an ability of cellular exosome to intercellularly deliver IFITM3 and thus transmit its antiviral effect from infected to non-infected cells. Thus, our findings provide new insights in the basic mechanisms underlying the actions of IFITM3, which might lead to future development of exosome-mediated anti-viral strategies using IFITM3 as a therapeutic agent. Conceivably, variations in the basal and inducible levels of IFITMs, as well as in intracellular and extracellular levels of IFITMs, might predict the severity of dengue virus infections among individuals or across species. (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 non-targeting control siRNA-transfected 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 DG Filtration dUC Protein markers EV: IFITM3/ CD63/ Flotillin2 non-EV: Calnexin/ GM130 Proteomics no Show all info Study aim Function Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells HUVEC EV-harvesting Medium EV-depleted medium Preparation of EDS overnight (16h) at >=100,000g Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: speed (g) 150000 Density gradient Type Discontinuous Number of initial discontinuous layers 3 Lowest density fraction 0.25M Highest density fraction 2.5M Sample volume (mL) 11 Orientation Bottom-up Speed (g) 150000 Duration (min) 960 Fraction processing Centrifugation Pelleting: duration (min) 90 Pelleting: speed (g) 150000 Pelleting-wash: duration (min) 90 Filtration steps 0.22µm or 0.2µm Other Name other separation method dUC Characterization: Protein analysis Protein Concentration Method BCA Western Blot Detected EV-associated proteins CD63/ Flotillin2/ IFITM3 Not detected contaminants Calnexin/ GM130 Characterization: Lipid analysis No

	EV220053	3/7	Homo sapiens	HUVEC	DG Filtration dUC	Zhu X	2015	33%
Study summary Full title IFITM3-containing exosome as a novel mediator for anti-viral response in dengue virus infection. All authors Zhu X, He Z, Yuan J, Wen W, Huang X, Hu Y, Lin C, Pan J, Li R, Deng H, Liao S, Zhou R, Wu J, Li J, Li M Journal Cell Microbiol Abstract Interferon-inducible transmembrane proteins 1, 2 and 3 (IFITM1, IFITM2 and IFITM3) have recently bee (show more...)Interferon-inducible transmembrane proteins 1, 2 and 3 (IFITM1, IFITM2 and IFITM3) have recently been identified as potent antiviral effectors that function to suppress the entry of a broad range of enveloped viruses and modulate cellular tropism independent of viral receptor expression. However, the antiviral effect and mechanisms of IFITMs in response to viral infections remain incompletely understood and characterized. In this work, we focused our investigation on the function of the extracellular IFITM3 protein. In cell models of DENV-2 infection, we found that IFITM3 contributed to both the baseline and interferon-induced inhibition of DENV entry. Most importantly, our study for the first time demonstrated the presence of IFITM-containing exosome in the extracellular environment, and identified an ability of cellular exosome to intercellularly deliver IFITM3 and thus transmit its antiviral effect from infected to non-infected cells. Thus, our findings provide new insights in the basic mechanisms underlying the actions of IFITM3, which might lead to future development of exosome-mediated anti-viral strategies using IFITM3 as a therapeutic agent. Conceivably, variations in the basal and inducible levels of IFITMs, as well as in intracellular and extracellular levels of IFITMs, might predict the severity of dengue virus infections among individuals or across species. (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 IFITM3 siRNA-1-transfected 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 DG Filtration dUC Protein markers EV: IFITM3/ CD63/ Flotillin2 non-EV: Calnexin/ GM130 Proteomics no Show all info Study aim Function Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells HUVEC EV-harvesting Medium EV-depleted medium Preparation of EDS overnight (16h) at >=100,000g Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: speed (g) 150000 Density gradient Type Discontinuous Number of initial discontinuous layers 3 Lowest density fraction 0.25M Highest density fraction 2.5M Sample volume (mL) 11 Orientation Bottom-up Speed (g) 150000 Duration (min) 960 Fraction processing Centrifugation Pelleting: duration (min) 90 Pelleting: speed (g) 150000 Pelleting-wash: duration (min) 90 Filtration steps 0.22µm or 0.2µm Other Name other separation method dUC Characterization: Protein analysis Protein Concentration Method BCA Western Blot Detected EV-associated proteins CD63/ Flotillin2 Not detected EV-associated proteins IFITM3 Not detected contaminants Calnexin/ GM130 Characterization: Lipid analysis No

	EV220053	4/7	Homo sapiens	293T	DG Filtration dUC	Zhu X	2015	33%
Study summary Full title IFITM3-containing exosome as a novel mediator for anti-viral response in dengue virus infection. All authors Zhu X, He Z, Yuan J, Wen W, Huang X, Hu Y, Lin C, Pan J, Li R, Deng H, Liao S, Zhou R, Wu J, Li J, Li M Journal Cell Microbiol Abstract Interferon-inducible transmembrane proteins 1, 2 and 3 (IFITM1, IFITM2 and IFITM3) have recently bee (show more...)Interferon-inducible transmembrane proteins 1, 2 and 3 (IFITM1, IFITM2 and IFITM3) have recently been identified as potent antiviral effectors that function to suppress the entry of a broad range of enveloped viruses and modulate cellular tropism independent of viral receptor expression. However, the antiviral effect and mechanisms of IFITMs in response to viral infections remain incompletely understood and characterized. In this work, we focused our investigation on the function of the extracellular IFITM3 protein. In cell models of DENV-2 infection, we found that IFITM3 contributed to both the baseline and interferon-induced inhibition of DENV entry. Most importantly, our study for the first time demonstrated the presence of IFITM-containing exosome in the extracellular environment, and identified an ability of cellular exosome to intercellularly deliver IFITM3 and thus transmit its antiviral effect from infected to non-infected cells. Thus, our findings provide new insights in the basic mechanisms underlying the actions of IFITM3, which might lead to future development of exosome-mediated anti-viral strategies using IFITM3 as a therapeutic agent. Conceivably, variations in the basal and inducible levels of IFITMs, as well as in intracellular and extracellular levels of IFITMs, might predict the severity of dengue virus infections among individuals or across species. (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 DG Filtration dUC Protein markers EV: IFITM3/ CD63/ Flotillin2 non-EV: Calnexin/ GM130 Proteomics no Show all info Study aim Function Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells 293T EV-harvesting Medium EV-depleted medium Preparation of EDS overnight (16h) at >=100,000g Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: speed (g) 150000 Density gradient Type Discontinuous Number of initial discontinuous layers 3 Lowest density fraction 0.25M Highest density fraction 2.5M Sample volume (mL) 11 Orientation Bottom-up Speed (g) 150000 Duration (min) 960 Fraction processing Centrifugation Pelleting: duration (min) 90 Pelleting: speed (g) 150000 Pelleting-wash: duration (min) 90 Filtration steps 0.22µm or 0.2µm Other Name other separation method dUC Characterization: Protein analysis Protein Concentration Method BCA Western Blot Detected EV-associated proteins CD63/ Flotillin2 Not detected EV-associated proteins IFITM3 Not detected contaminants Calnexin/ GM130 Characterization: Lipid analysis No

	EV220053	5/7	Homo sapiens	293T	DG Filtration dUC	Zhu X	2015	33%
Study summary Full title IFITM3-containing exosome as a novel mediator for anti-viral response in dengue virus infection. All authors Zhu X, He Z, Yuan J, Wen W, Huang X, Hu Y, Lin C, Pan J, Li R, Deng H, Liao S, Zhou R, Wu J, Li J, Li M Journal Cell Microbiol Abstract Interferon-inducible transmembrane proteins 1, 2 and 3 (IFITM1, IFITM2 and IFITM3) have recently bee (show more...)Interferon-inducible transmembrane proteins 1, 2 and 3 (IFITM1, IFITM2 and IFITM3) have recently been identified as potent antiviral effectors that function to suppress the entry of a broad range of enveloped viruses and modulate cellular tropism independent of viral receptor expression. However, the antiviral effect and mechanisms of IFITMs in response to viral infections remain incompletely understood and characterized. In this work, we focused our investigation on the function of the extracellular IFITM3 protein. In cell models of DENV-2 infection, we found that IFITM3 contributed to both the baseline and interferon-induced inhibition of DENV entry. Most importantly, our study for the first time demonstrated the presence of IFITM-containing exosome in the extracellular environment, and identified an ability of cellular exosome to intercellularly deliver IFITM3 and thus transmit its antiviral effect from infected to non-infected cells. Thus, our findings provide new insights in the basic mechanisms underlying the actions of IFITM3, which might lead to future development of exosome-mediated anti-viral strategies using IFITM3 as a therapeutic agent. Conceivably, variations in the basal and inducible levels of IFITMs, as well as in intracellular and extracellular levels of IFITMs, might predict the severity of dengue virus infections among individuals or across species. (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 IFITM3-transfected 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 DG Filtration dUC Protein markers EV: IFITM3/ CD63/ Flotillin2 non-EV: Calnexin/ GM130 Proteomics no Show all info Study aim Function Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells 293T EV-harvesting Medium EV-depleted medium Preparation of EDS overnight (16h) at >=100,000g Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: speed (g) 150000 Density gradient Type Discontinuous Number of initial discontinuous layers 3 Lowest density fraction 0.25M Highest density fraction 2.5M Sample volume (mL) 11 Orientation Bottom-up Speed (g) 150000 Duration (min) 960 Fraction processing Centrifugation Pelleting: duration (min) 90 Pelleting: speed (g) 150000 Pelleting-wash: duration (min) 90 Filtration steps 0.22µm or 0.2µm Other Name other separation method dUC Characterization: Protein analysis Protein Concentration Method BCA Western Blot Detected EV-associated proteins IFITM3/ CD63/ Flotillin2 Not detected contaminants Calnexin/ GM130 Characterization: Lipid analysis No

	EV220053	6/7	Homo sapiens	293T	DG Filtration dUC	Zhu X	2015	33%
Study summary Full title IFITM3-containing exosome as a novel mediator for anti-viral response in dengue virus infection. All authors Zhu X, He Z, Yuan J, Wen W, Huang X, Hu Y, Lin C, Pan J, Li R, Deng H, Liao S, Zhou R, Wu J, Li J, Li M Journal Cell Microbiol Abstract Interferon-inducible transmembrane proteins 1, 2 and 3 (IFITM1, IFITM2 and IFITM3) have recently bee (show more...)Interferon-inducible transmembrane proteins 1, 2 and 3 (IFITM1, IFITM2 and IFITM3) have recently been identified as potent antiviral effectors that function to suppress the entry of a broad range of enveloped viruses and modulate cellular tropism independent of viral receptor expression. However, the antiviral effect and mechanisms of IFITMs in response to viral infections remain incompletely understood and characterized. In this work, we focused our investigation on the function of the extracellular IFITM3 protein. In cell models of DENV-2 infection, we found that IFITM3 contributed to both the baseline and interferon-induced inhibition of DENV entry. Most importantly, our study for the first time demonstrated the presence of IFITM-containing exosome in the extracellular environment, and identified an ability of cellular exosome to intercellularly deliver IFITM3 and thus transmit its antiviral effect from infected to non-infected cells. Thus, our findings provide new insights in the basic mechanisms underlying the actions of IFITM3, which might lead to future development of exosome-mediated anti-viral strategies using IFITM3 as a therapeutic agent. Conceivably, variations in the basal and inducible levels of IFITMs, as well as in intracellular and extracellular levels of IFITMs, might predict the severity of dengue virus infections among individuals or across species. (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 vector-transfected 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 DG Filtration dUC Protein markers EV: IFITM3/ CD63/ Flotillin2 non-EV: Calnexin/ GM130 Proteomics no Show all info Study aim Function Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells 293T EV-harvesting Medium EV-depleted medium Preparation of EDS overnight (16h) at >=100,000g Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: speed (g) 150000 Density gradient Type Discontinuous Number of initial discontinuous layers 3 Lowest density fraction 0.25M Highest density fraction 2.5M Sample volume (mL) 11 Orientation Bottom-up Speed (g) 150000 Duration (min) 960 Fraction processing Centrifugation Pelleting: duration (min) 90 Pelleting: speed (g) 150000 Pelleting-wash: duration (min) 90 Filtration steps 0.22µm or 0.2µm Other Name other separation method dUC Characterization: Protein analysis Protein Concentration Method BCA Western Blot Detected EV-associated proteins CD63/ Flotillin2 Not detected EV-associated proteins IFITM3 Not detected contaminants Calnexin/ GM130 Characterization: Lipid analysis No

	EV220053	7/7	Homo sapiens	293T	DG Filtration dUC	Zhu X	2015	33%
Study summary Full title IFITM3-containing exosome as a novel mediator for anti-viral response in dengue virus infection. All authors Zhu X, He Z, Yuan J, Wen W, Huang X, Hu Y, Lin C, Pan J, Li R, Deng H, Liao S, Zhou R, Wu J, Li J, Li M Journal Cell Microbiol Abstract Interferon-inducible transmembrane proteins 1, 2 and 3 (IFITM1, IFITM2 and IFITM3) have recently bee (show more...)Interferon-inducible transmembrane proteins 1, 2 and 3 (IFITM1, IFITM2 and IFITM3) have recently been identified as potent antiviral effectors that function to suppress the entry of a broad range of enveloped viruses and modulate cellular tropism independent of viral receptor expression. However, the antiviral effect and mechanisms of IFITMs in response to viral infections remain incompletely understood and characterized. In this work, we focused our investigation on the function of the extracellular IFITM3 protein. In cell models of DENV-2 infection, we found that IFITM3 contributed to both the baseline and interferon-induced inhibition of DENV entry. Most importantly, our study for the first time demonstrated the presence of IFITM-containing exosome in the extracellular environment, and identified an ability of cellular exosome to intercellularly deliver IFITM3 and thus transmit its antiviral effect from infected to non-infected cells. Thus, our findings provide new insights in the basic mechanisms underlying the actions of IFITM3, which might lead to future development of exosome-mediated anti-viral strategies using IFITM3 as a therapeutic agent. Conceivably, variations in the basal and inducible levels of IFITMs, as well as in intracellular and extracellular levels of IFITMs, might predict the severity of dengue virus infections among individuals or across species. (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 IFN--treated 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 DG Filtration dUC Protein markers EV: IFITM3/ CD63/ Flotillin2 non-EV: Calnexin/ GM130 Proteomics no Show all info Study aim Function Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells 293T EV-harvesting Medium EV-depleted medium Preparation of EDS overnight (16h) at >=100,000g Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Equal to or above 150,000 g Pelleting performed Yes Pelleting: speed (g) 150000 Density gradient Type Discontinuous Number of initial discontinuous layers 3 Lowest density fraction 0.25M Highest density fraction 2.5M Sample volume (mL) 11 Orientation Bottom-up Speed (g) 150000 Duration (min) 960 Fraction processing Centrifugation Pelleting: duration (min) 90 Pelleting: speed (g) 150000 Pelleting-wash: duration (min) 90 Filtration steps 0.22µm or 0.2µm Other Name other separation method dUC Characterization: Protein analysis Protein Concentration Method BCA Western Blot Detected EV-associated proteins IFITM3/ CD63/ Flotillin2 Not detected contaminants Calnexin/ GM130 Characterization: Lipid analysis No

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EV-TRACK ID	EV220053
species	Homo sapiens
sample type	Cell culture
cell type	HUVEC	HUVEC	HUVEC	293T	293T	293T	293T
condition	Control condition	non-targeting control siRNA-transfected	IFITM3 siRNA-1-transfected	Control condition	IFITM3-transfected	control vector-transfected	IFN--treated
separation protocol	DG Filtration dUC	DG Filtration dUC	DG Filtration dUC	DG Filtration dUC	DG Filtration dUC	DG Filtration dUC	DG Filtration dUC
Exp. nr.	1	2	3	4	5	6	7
EV-METRIC %	33	33	33	33	33	33	33

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