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Details	EV-TRACK ID	Experiment nr.	Species	Sample type	Separation protocol	First author	Year	EV-METRIC
	EV110008	1/1	Mus musculus	NAY	(d)(U)C DG	Lee YS	2011	33%
Study summary Full title Introduction of the CIITA gene into tumor cells produces exosomes with enhanced anti-tumor effects All authors Lee YS, Kim SH, Cho JA, Kim CW Journal Exp Mol Med Abstract Exosomes are small membrane vesicles secreted from various types of cells. Tumor-derived exosomes co (show more...)Exosomes are small membrane vesicles secreted from various types of cells. Tumor-derived exosomes contain MHC class I molecules and tumor-specific antigens, receiving attention as a potential cancer vaccine. For induction of efficient anti-tumor immunity, CD4+ helper T cells are required, which recognize appropriate MHC class II-peptide complexes. In this study, we have established an MHC class II molecule-expressing B16F1 murine melanoma cell line (B16F1- CIITA) by transduction of the CIITA (Class II transactivator) gene. Exosomes from B16-CII cells (CIITA- Exo) contained a high amount of MHC class II as well as a tumor antigen TRP2. When loaded on dendritic cells (DCs), CIITA-Exo induced the increased expression of MHC class II molecules and CD86 than the exosomes from the parental cells (Exo). In vitro assays using co-culture of immunized splenocytes and exosome-loaded DCs demonstrated that CIITA-Exo enhanced the splenocyte proliferation and IL-2 secretion. Consistently, compared to B16-Exo, CIITA-Exo induced the increased mRNA levels of inflammatory cytokines such as TNF-?, chemokine receptor CCR7 and the production of Th1-polarizing cytokine IL-12. A tumor preventive model showed that CIITA-Exo significantly inhibited tumor growth in a dose-dependent manner. Ex vivo assays using immunized mice demonstrated that CIITA-Exo induced a higher amount of Th1-polarized immune responses such as Th1-type IgG2a antibodies and IFN-? cytokine as well as TRP2-specific CD8+ T cells. A tumor therapeutic model delayed effects of tumor growth by CIITA-Exo. These findings indicate that CIITA-Exo are more efficient as compared to parental Exo to induce anti-tumor immune responses, suggesting a potential role of MHC class II-containing tumor exosomes as an efficient cancer vaccine. (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 Protein markers EV: TSG101/ MHC2/ Hsc70/ MHC1 non-EV: Tubulin/ Cell organelle protein Proteomics no Show all info Study aim Function Sample Species Mus musculus Sample Type Cell culture supernatant EV-harvesting Medium EV Depleted 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) 60 Density gradient Only used for validation of main results Yes Lowest density fraction 0.25 Highest density fraction 2.5 Orientation Bottom-up Characterization: Protein analysis Western Blot Antibody details provided? No Detected EV-associated proteins TSG101/ MHC1/ MHC2/ Hsc70 Detected contaminants Cell organelle protein/ Tubulin ELISA Antibody details provided? No Detected EV-associated proteins MHC1/ MHC2/ Hsc70 Characterization: Particle analysis EM EM-type transmission EM Image type Wide-field

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EV-TRACK ID	EV110008
species	Mus musculus
sample type	Cell culture
cell type	NAY
condition	NAY
separation protocol	(d)(U)C DG
Exp. nr.	1
EV-METRIC %	33