Molecular and Functional Analysis of E0771 TAM-EVs (A) Schedule of subcutaneous E0771 cancer cell inoculation in C57BL/6 mice and drug administration. (B) Flow cytometry analysis of immune infiltrates in E0771 tumors. Data show percentage values (mean ± SD; n = 3 mice/condition). Statistics by unpaired two-tailed Student's t test (left and middle) or two-way ANOVA, using Sidak's multiple comparison test (right). (C) Yield of EVs recovered from E0771 tumors prior to and after sucrose gradient fractionation, determined by BCA (mean ± SD; n = 3 EV preparations/condition). Statistics by unpaired two-tailed Student's t test. (D) EV concentration and size distribution by NTA in the 6 sucrose fractions (mean ± SEM; n = 3 EV preparations/condition). (E) EV protein content and EV concentration in each sucrose fraction determined by BCA and NTA, respectively (mean ± SD; n = 3 EV preparations/condition). (F) Representative TEM images of EVs. One representative EV preparation per condition is shown. Scale bars, 200 nm. (G) WB analysis of cultured E0771 cells and matched EVs. One representative EV preparation is shown. (H and I) WB analysis of E0771 tumor-EVs (H). Relative signal quantification of MRC1, COX1, and TBXAS1 is shown in (I) as mean band intensity normalized to CD9 (n = 3 EV preparations/condition). Statistics as in (C). (J) Venn diagrams comparing proteins enriched in IgG-EVs versus anti-CSF1R-EVs from E0771 tumors (FC > 1.7; see Table S7 ) with those of the MC38 TAM-EV signature (see Table S5 ). (K) LC-MS/MS analysis of TBXAS1 in E0771 tumor-EVs (mean ± SD; n = 3 EV preparations/condition). Statistics as in (C). (L) ELISA-based quantification of TXB 2 in medium conditioned by E0771 cells (mean ± SD; n = 3 EV preparations/condition). Statistics by one-way ANOVA, using Tukey's multiple comparison test.
{"Image Orientation": "Horizontal (normal)", "Image XResolution": "500", "Image YResolution": "500", "Image ResolutionUnit": "Pixels/Inch", "Image Software": "Adobe Photoshop CC 2017 (Windows)", "Image DateTime": "2019:05:23 14:35:38", "Image ExifOffset": "168", "Thumbnail Compression": "JPEG (old-style)", "Thumbnail XResolution": "500", "Thumbnail YResolution": "500", "Thumbnail ResolutionUnit": "Pixels/Inch", "Thumbnail JPEGInterchangeFormat": "306", "Thumbnail JPEGInterchangeFormatLength": "6929", "EXIF ColorSpace": "Uncalibrated", "EXIF ExifImageWidth": "3068", "EXIF ExifImageLength": "4240"}