Supplementary Figure 6: An anti-HER2 EVIR enhances EV-uptake by MHCI-deficient DCs.
From: EVIR: chimeric receptors that enhance dendritic cell cross-dressing with tumor antigens
(a) Schematic representation of the proviral CRISPR LV, which contains a control gRNA or a gRNA targeting H2kb or B2m, along with puromycin (PURO) resistance and inducible CAS9 transgenes.
(b) Flow cytometry dot plots showing H-2Kb or B2M expression in MC38 cells transduced with a LV without gRNA (control) or containing a gRNA either targeting H2kb or B2m. The analysis shows cell clones that have been selected for subsequent studies.
(c) PCR analysis of genomic OVA in the indicated MC38 cell lines. The analysis shows cell clones that have been selected for subsequent studies.
(d) Size distribution of H2kb- and B2m-deficient MC38-EVs, as determined by NTA. Data show mean values ± SEM (n=3 technical measurements of one EV preparation per type). Data are representative of 2 independent EV preparations.
(e) Flow cytometry showing the MFI of PKH67 in DC-CtrlR and DC-EVIR treated with PKH67-labeled, wild-type (WT, H2kb-proficient), H2kb- or B2m-deficient EVs that had been isolated from either MC38 or MC38-HER2 cells, as indicated. DCs were analyzed 24 h after exposure to EVs. Data show mean values ± SEM (n=3 independent cell cultures per condition); statistical analysis by two-way ANOVA with Sidak's multiple comparison test.
Numerical values for the experiments with quantitative data are presented in Supplementary Table 2.
