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. 2023 Apr 23;31(8):2489–2506. doi: 10.1016/j.ymthe.2023.04.011

Figure 1.

Figure 1

Preparation and characterization of siAdar1-LNP@mPD1 nanovesicles

(A) Schematic illustration of siAdar1-LNP@mPD1 nanovesicle preparation and the principle of antitumor immune response. (B–D) CLSM images (B), flow cytometry analysis (C), and western blot (WB) analysis (D) of stably expressing PD1 on engineered CHO cell membrane. Scale bar, 5 μm. (E) The interaction between mPD1 NVs and anti-PD1 antibody detected by co-immunoprecipitation (coIP). For immunoprecipitation, anti-PD1 antibody was used to pull down mPD1 NVs, while IgG was used as the control. (F) CLSM images of mPD1 NVs binding to PDL1-EGFP that is intentionally overexpressed on 4T1 cells, the co-localization of mPD1 NVs and PDL1 proteins are represented by the yellow colored dots as indicated by white arrows. Scale bar, 20 μm). mPD1 NVs (40 μg/mL) were incubated with 4T1 cells at 4°C for 1 h. (G) CLSM images of siAdar1-LNP with Cy5-labeled siAdar1 (red fluorescence) and LNP loaded with DiO (green fluorescence). Scale bars, 20 μm. (H) Gel fluorescence imaging of free siAdar1 and siAdar1-LNP. (I and J) Transmission electron microscopy images (I) and dynamic light scattering (DLS) results (J) of siAdar1-LNP and siAdar1-LNP@mPD1. Scale bar, 100 nm. (K and L) SDS-PAGE electrophoresis patterns (K) and WB analysis (L) of siAdar1-LNP (I), mPD1 NVs (II), and siAdar1-LNP@mPD1 (III).