(A) AcGFP-actin–expressing Raji cells were treated with either control mAbs (OKT3), tositumomab, or L243 (all at 10 μg/ml) for 4 hours, then studied by FRAP, with flurorescence recovery assessed for 100 seconds. Typical image data is shown for control- and tositumomab-treated cells. Upper panels: virtually no recovery was observed in control mAb–treated cells. Lower panels (Tos): recovery of junctional actin within the cell-to-cell contact area was observed after approximately 80 seconds. Scale bars: 20 μm. Squares indicate regions targeted for photobleaching. (B) Typical fluorescence recovery pattern of AcGFP-actin after photobleaching. Control cells showed no recovery within 100 seconds (left). Recovery of junctional actin fluorescence was shown within 80 seconds after photobleaching in cells treated with anti-CD20 (Tos; center) and anti–HLA-DR (L243; right). (C) Transient cytoplasmic bridges between cells undergoing HA. Phase contrast time-lapse microscopy of Raji cells treated with tositumomab (10 μg/ml). Intervals between images are 5 minutes. Arrows indicate location of bridge formation. Scale bar: 20 μm. (D and E) Peripheral relocalization of mitochondria toward cell-to-cell contact area as assessed by TEM (D) and JC-1 staining (E). Arrows indicate localized mitochondria. Scale bar: 10 μm. (E) JC-1–labeled cells were treated with tositumomab or L243 (10 μg/ml). After that, images were captured detecting monomeric (green) and J-aggregate (red) forms of JC-1. Arrows indicate J-aggregate mitochondria at cell-cell junctions. Scale bars: 10 μm (D); 20 μm (E). (F) Time-lapse video microscopy of JC-1–labeled cells. Red mitochondria represent mitochondria with high membrane potential, migrating toward cell-to-cell contact areas. Interval between images is 1 hour. Scale bar: 20 μm.