Normal and malignant human B cells internalize type I anti-CD20 mAb. (A) An in vitro surface fluorescence quenching assay as described in “Surface fluoresence quenching assay” was used to analyze primary CLL cells treated with Alexa-488–labeled mAb (Tosit-488, GA101gly-488, Ritux-488 or Ofatum-488, all 5 μg/mL) for 2 or 6 hours (***P < .0001, Wilcoxon test). Internalization of anti-CD20 mAb is expressed as the percentage of anti-CD20 mAb detectable at the cell surface using secondary (quenching) anti–Alexa-488 mAb (surface accessible anti-CD20). Anti-CD20 mAb that has been internalized cannot be quenched by anti–Alexa-488 and so the cells remain fluorescent. (B) A variety of B-cell tumors and normal B cells from healthy volunteers were similarly examined after treatment with Tosit-488 or Ritux-488 (5 μg/mL), and the extent of accessible mAb shown. Each point represents a sample from a different patient and medians are shown.