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Results list Most used isolation protocols Top EV-enriched proteins

Details	EV-TRACK ID	Experiment nr.	Species	Sample type	Isolation protocol	First author	Year	EV-METRIC
	EV110005	1/1	Rattus norvegicus/rattus	Cell culture supernatant	dUC Sucrose-DG (valid.)	Carayon K	2011	44%
Study summary Full title Proteolipidic composition of exosomes changes during reticulocyte maturation All authors Carayon K, Chaoui K, Ronzier E, Lazar I, Bertrand-Michel J, Roques V, Balor S, Terce F, Lopez A, Salomé L, Joly E Journal J Biol Chem Abstract During the orchestrated process leading to mature erythrocytes, reticulocytes must synthesize large (show more...)During the orchestrated process leading to mature erythrocytes, reticulocytes must synthesize large amounts of hemoglobin, while eliminating numerous cellular components. Exosomes are small secreted vesicles that play an important role in this process of specific elimination. To understand the mechanisms of proteolipidic sorting leading to their biogenesis, we have explored changes in the composition of exosomes released by reticulocytes during their differentiation, in parallel to their physical properties. By combining proteomic and lipidomic approaches, we found dramatic alterations in the composition of the exosomes retrieved over the course of a 7-day in vitro differentiation protocol. Our data support a previously proposed model, whereby in reticulocytes the biogenesis of exosomes involves several distinct mechanisms for the preferential recruitment of particular proteins and lipids and suggest that the respective prominence of those pathways changes over the course of the differentiation process. (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 electron microscopy images Particle analysis: inclusion of a widefield and close-up electron microscopy image density gradient Isolation method: density gradient, at least as validation of results attributed to EVs EV density Isolation method: reporting of obtained EV density ultracentrifugation specifics Isolation 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 Focus vesicles exosomes Isolation protocol Isolation protocol Gives a short, non-chronological overview of the different steps of the isolation protocol. dUC = differential ultracentrifugation DG = density gradient UF = ultrafiltration SEC = size-exclusion chromatography dUC + Sucrose-DG (valid.) Protein markers EV: HSP90/ TSG101/ HSC70/ CHMP4B/ CHMP4C/ MHC1 non-EV: None Proteomics yes EV density (g/ml) 1.16-1.21 Show all info Study aim Omics Sample Species Rattus norvegicus/rattus Sample Type Cell culture supernatant EV-harvesting Medium EV Depleted Isolation Method Differential ultracentrifugation dUC: centrifugation steps Between 10,000 g and 50,000 g Between 100,000 g and 150,000 g Pelleting: time(min) 120 Density gradient Only used for validation of main results 1 Density medium Sucrose Lowest density fraction 0.5 Highest density fraction 2.5 Orientation Top-down Speed (g) 100000 Characterization: Protein analysis Western Blot Detected EV-associated proteins HSP90/ TSG101/ HSC70/ CHMP4B/ CHMP4C/ MHC1 ELISA Detected EV-associated proteins HSC70/ CHMP4B/ CHMP4C/ MHC1 Fluorescent NTA Characterization: Particle analysis EM EM-type transmission EM Image type Wide-field

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