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Unconjugated monoclonal antibodies (mAbs) are an important component of effective combination therapies for chronic lymphocytic leukaemia (CLL). Antibody-dependent phagocytosis (ADP) is a major mediator of mAb cytotoxicity, but there is limited knowledge of the determinants of ADP efficacy. We used macrophages derivedin vitrofrom autologous circulating monocytes to test the effects of mAb structure and concentration, target : effector cell ratio, duration of co-incubation and CLL cell CD20 expression on ADP. Next-generation anti-CD20 mAbs (ofatumumab, ublituximab, obinutuzumab, ocaratuzumab) were significantly more effective at inducing ADP compared to rituximab, but none were as effective as the anti-CD52 mAb alemtuzumab. Ofatumumab (10 μg/ml) used as a representative next-generation anti-CD20 mAb achieved an ADP plateau at 3 h co-incubation with a target : effector ratio of 10 : 1 (mean = 2·1 CLL cells/macrophage, range = 1·5–3·5). At 0·156 μg/ml (the lowest concentration tested) ofatumumab ADP was significantly higher than alemtuzumab. However, ofatumumab-induced ADP did not increase significantly at higher mAb concentrations. We show that anti-CD20 mAb ADP efficacy is determined by the mAb characteristics, target : effector ratio and incubation time. We suggest that preclinical evaluation of anti-CD20 mAbs to understand the determinants of ADP could be useful in designing future combination therapies for CLL.