- 12-0639-41 - Invitrogen Antibodies CD63 antibody

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M2-like tumor-associated macrophages transmit exosomal miR-27b-3p and maintain glioblastoma stem-like cell properties.

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Kuypers S, Smisdom N, Pintelon I, Timmermans JP, Ameloot M, Michiels L, Hendrix J, Hosseinkhani B
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Enrichment of CD44 in Exosomes From Breast Cancer Cells Treated With Doxorubicin Promotes Chemoresistance.

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Cheng L, Zhang K, Qing Y, Li D, Cui M, Jin P, Xu T
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Comprehensive Cell Surface Antigen Analysis Identifies Transferrin Receptor Protein-1 (CD71) as a Negative Selection Marker for Human Neuronal Cells.

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
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Experimental details: Staining of normal human peripheral blood cells with Anti-Human CD193 (CCR3) APC (Product # 17-1939-42), Anti-Human IgE FITC (Product # 11-6986-42) and Mouse IgG1 K Isotype Control PE (Product # 12-4714-81) (blue histogram) or Anti-Human CD63 PE (purple histogram). CD193+IgE+ cells (left) were used for analysis.
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Experimental details: NULL
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Supportive validation

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Experimental details: Figure 2 hWJMSC-MVs surface expressed molecules analysis. Flow cytometery analysis showed hWJMSC-MVs were positive for some surface expressed molecules typically expressed by MSCs, such as CD9, CD44, CD63, CD73, and negative for CD34, CD45.
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Experimental details: Figure 1 Characterization of endothelial colony-forming cells (ECFC)-derived extracellular vesicles (EVs). ( A ) Isolated ECFC-derived EVs were conjugated to CD63-coated latex beads to aid detection by flow cytometry. The gated bead-bound population (left panel) tested positive for EV markers CD9 and CD63. ( B ) Electron micrograph demonstrating the heterogeneity of the EV population (scale bar 1 mum). ( C ) Higher-magnification electron micrograph showing EVs of variable sizes and including a multivesicular body (scale bar 200 nm). ( D-F ) Electron micrographs of EVs bound to latex beads (LB) coated with anti-CD63 antibody. ( D ) White arrows indicate CD63-positive EVs of varying sizes (scale bar 1 mum). ( E ) Higher-magnification micrograph of EVs (scale bar 200 nm). In ( F ), a multivesicular body appears to be encapsulated within a CD63-positive membrane (scale bar 200 nm).
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Experimental details: Fig. 3 CTSS is produced in macrophage and transported by exosomes. Western blot analysis of CTSS in macrophage cell lysates and medium after PA (500 muM) stimulation for 24 h ( A ). Western blot analysis of exosome-specific proteins in exosomes and cell lysates detected 24 h after palmitic acid (PA, 500 muM) stimulation of macrophage ( B ). Electron microscopy of exosomes ( C ). (Scale bars = 500 nm) Exosome-derived CTSS from PA (500 muM) stimulated macrophage ( D ). QRT-PCR ( E ) and western blot ( F ) analysis of CTSS gene knockdown efficiency in macrophage. Data are shown as the mean +- SEM, n = 3, ANOVA. * P < 0.05. Detection of CTSS in exosomes from PA (500 muM) stimulated macrophage; macrophage were transfected with shCTSS, scr transfection as the control ( G )
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Experimental details: Fig. 2 M2-TAM-derived exosomes enhance the stemness of GSCs. A TEM images of M2-TAM-derived exosomes. The presence of CD63, CD206, and CD163 in M2-TAM-derived exosomes analyzed by flow cytometric ( B ) and Western blot ( C ) analyses. D Fluorescence images of GSCs co-cultured with Dil-labeled M2-TAM-derived exosomes with nuclei stained with DAPI (blue). E Representative images showing neurosphere formation of GSCs along with the statistics of GSC formation rate and diameter of spheres. F Expression of stem cell-related protein CD133, Nestin, Oct4, Sox2, and GFAP in GSCs measured by Western blot analysis. G HE staining images of xenograft tumors from mice. H The representative immunohistochemical images of Sox2 in xenograft tumors and the percentage of GSCs labeled by Sox2. I Kaplan-Meier survive curve of tumor-bearing mice. n = 10. * p < 0.05 vs. the control EXO group. Measurement data were depicted as mean +- standard deviation, comparison of that between two groups was conducted by unpaired t test. Cell experiments were repeated three times independently.
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Supportive validation

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Experimental details: Figure 5 ( A ) Flow cytometric data confirmed exosomes by identifying exosomal marker CD63 and 90 +- 8% of exosomes in suspension represented CD63 marker. ( B ) Representative micrograph from transmission electron microscopy showing the nano-sized exosomes (Scale bar 40 nm). Exo: exosomes.
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Supportive validation

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Experimental details: Fig. 1 Exosome isolation and analysis. A&C Representative transmission electron microscopy (TEM) images obtained for exosomes from HOSEPiC and SKOV-3. Scale bar: 100 nm. B&D flow cytometry analysis analyses show intensities for exosomal markers (CD9, CD63). (E) Particle size distribution by NanoSight analysis of exosomes from HOSEPiC and SKOV-3 (F) Western blot analyses show increased intensities for exosomal markers (Alix, TSG101)
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12-0639-41

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