Confocal microscopy showing membrane compartmentalization of rituximab-associated versus GA101-associated CD20 complexes. (A) After ≥ 1 hour of incubation at 37°C with 5 μg/mL of dye-labeled antibodies, only partial colocalization of rituximab (red) and GA101 (green) was seen in isolated Z138 cells (panels i-iii), whereas control stainings with differently labeled versions of the same antibody resulted in each case in 100% colocalization (panels iv-vi for GA101 and panels vii-ix for rituximab). (B) Different membrane compartmentalization of rituximab (green) and GA101 (red) was also observed in the Burkitt lymphoma cell line Ramos (panels i-iii) and in freshly isolated normal B cells (panels iv-vi). (C) After homotypic aggregation of Z138 cells, rituximab (green) remained concentrated in large membrane spots, whereas GA101 (red) accumulated in areas of cell-cell contact, resulting in almost complete separation of the 2 labels (panels i-iii).