Defective lysosome docking and secretion at the IS of Exo70-silenced cells. (A) Confocal images of control and Exo70-silenced cells activated on BCR ligand⁺ coverslips for 60 min. LAMP-1 (green), Exo70 (red), and F-actin (blue). Scale bar: 3 µm. (B) Ratio between the central and peripheral lysosomes in the cell. **, P = 0.001; unpaired t test; n ≥ 44 cells from two independent experiments. (C) Top: TIRF images of LysoSensor Green–stained lysosomes at IS of control and Exo70-silenced B cells plated on BCR ligand⁺ coated slides. Bottom: Lysosome trajectories of the TIRF images are shown as yellow lines. Scale bar: 5 µm. (D) Quantification of dynamic parameters of lysosomes at the IS. ****, P < 0.0001; unpaired t test; n ≥ 500 lysosome trajectories from ≥14 cells per condition from two independent experiments. Error bars are mean ± SEM. (E) Representative epifluorescence images of Exo70-silenced or control cells activated with CypHer5E-coupled BCR ligand⁺ beads for 90 min. Cells are shown as Z-projections of a stack. Insets highlight the CypHer5E fluorescence in the bead. Scale bar: 3 µm. (F) Quantification of the bead CypHer5E intensity, normalized by the intensity of noninteracting beads. ****, P < 0.0001; two-way ANOVA with Sidak’s multiple comparison test; n ≥ 160 cells from two independent experiments. Error bars are mean ± SEM.