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Details	EV-TRACK ID	Experiment nr.	Species	Sample type	Separation protocol	First author	Year	EV-METRIC
	EV110024	1/1	Homo sapiens	NAY	(d)(U)C DG Filtration	Xiu F	2011	33%
Study summary Full title Cutting edge: HLA-DO impairs the incorporation of HLA-DM into exosomes All authors Xiu F, Côté MH, Bourgeois-Daigneault MC, Brunet A, Gauvreau MÉ, Shaw A, Thibodeau J Journal J Immunol Abstract In multivesicular bodies, HLA-DM (DM) assists the loading of antigenic peptides on classical MHC cla (show more...)In multivesicular bodies, HLA-DM (DM) assists the loading of antigenic peptides on classical MHC class II molecules such as HLA-DR. In cells expressing HLA-DO (DO), DM is redistributed from the internal vesicles to the limiting membrane of these organelles. This suggests that DO might reduce DM incorporation into exosomes, which are shed upon fusion of multivesicular bodies with the plasma membrane. To test this hypothesis, we used the 721.45 B lymphoblastoid cell line and different HeLa cell transfectants. We demonstrate that the poor recovery of DM in exosomes as compared with HLA-DR is not the mere reflection of differences in protein expression. Indeed, we found that DO contributes to the inefficient transfer of DM to exosomes. This negative regulation requires an intact di-leucine endosomal sorting motif in the cytoplasmic tail of HLA-DO?. These results demonstrate that canonical sorting signals and protein-protein interactions modulate the selection of MHC protein cargos. (hide) EV-METRIC 33% (75th 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 Filtration Protein markers EV: MHC2/ CD9 non-EV: Proteomics no EV density (g/ml) 1.11-1.27 Show all info Study aim Biogenesis/Sorting Sample Species Homo sapiens Sample Type Cell culture supernatant EV-harvesting Medium EV Depleted 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 0.25 Highest density fraction 2 Orientation Bottom-up Filtration steps 0.22µm or 0.2µm Characterization: Protein analysis Western Blot Antibody details provided? No Detected EV-associated proteins CD9/ MHC2 ELISA Antibody details provided? No Detected EV-associated proteins MHC2 Characterization: Particle analysis EM EM-type immune EM EM protein CD9; MHC2 Image type Close-up

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EV-TRACK ID	EV110024
species	Homo sapiens
sample type	Cell culture
cell type	NAY
condition	NAY
separation protocol	(d)(U)C DG Filtration
Exp. nr.	1
EV-METRIC %	33