Fig 1: Antigen-induced permeabilization triggers lysosomal exocytosis.(A) Flow cytometry analysis of surface-exposed (no detergent permeabilization) and/or intracellular LIMP-2 (with detergent permeabilization) of bead-bound B-cells after incubation with αM- or Tf-beads for 30 min, showing 3000 cells per condition. (B and C) Percentages of cells with surface-exposed LIMP-2 (relative to values with secondary antibody alone) in bead-bound B-cells incubated with αM- or Tf-beads (B) or with HEL-, DEL-I- or Tf-beads (C) for 30 min. Data points represent independent experiments (mean ± SD). (D) Confocal images of surface-exposed LIMP-2 in B-cells incubated with αM- or Tf-beads (arrows, bead-binding sites). (E) FIR (bead-binding site:opposite PM) of surface-exposed LIMP-2 in individual cells over time. Data points represent individual cells (mean ± SD). (F) Total internal reflection microscopy (TIRF) images (left) and FI surface plots (right) of SiR-Lyso at the B-cell surface contacting αM-PLB (Video 10). (G) Representative MFI versus time plot of a SiR-Lyso-loaded lysosome undergoing exocytosis. (H) SiR-Lyso exocytosis events (circles) in individual B-cells during the first 0–15 min or 25–45 min of incubation with αM-PLB. (I) Timing of individual SiR-Lyso exocytosis events in B-cells incubated with αM-PLB for 45 min. Data points represent individual SiR-Lyso exocytosis events from three independent experiments (mean ± SD). (J) Numbers of SiR-Lyso exocytosis events per B-cell permeabilized (PI+) or not permeabilized (PI-) by αM-PLB during 45 min. Data points represent individual cells from three independent experiments (mean ± SD). *p ≤ 0.05, **p ≤ 0.01, unpaired Student’s t-test (B and J) or one-way ANOVA (C and E). Bars, 5 μm.