CD20 monoclonal antibody (mAb) immunotherapy is effective for lymphoma and autoimmune disease. In a mouse model of immunotherapy using mouse anti–mouse CD20 mAbs, the innate monocyte network depletes B cells through immunoglobulin (Ig)G Fc receptor (FcγR)-dependent pathways with a hierarchy of IgG2a/c>IgG1/IgG2b>IgG3. To understand the molecular basis for these CD20 mAb subclass differences, B cell depletion was assessed in mice deficient or blocked for stimulatory FcγRI, FcγRIII, FcγRIV, or FcR common γ chain, or inhibitory FcγRIIB. IgG1 CD20 mAbs induced B cell depletion through preferential, if not exclusive, interactions with low-affinity FcγRIII. IgG2b CD20 mAbs interacted preferentially with intermediate affinity FcγRIV. The potency of IgG2a/c CD20 mAbs resulted from FcγRIV interactions, with potential contributions from high-affinity FcγRI. Regardless, FcγRIV could mediate IgG2a/b/c CD20 mAb–induced depletion in the absence of FcγRI and FcγRIII. In contrast, inhibitory FcγRIIB deficiency significantly increased CD20 mAb–induced B cell depletion by enhancing monocyte function. Although FcγR-dependent pathways regulated B cell depletion from lymphoid tissues, both FcγR-dependent and -independent pathways contributed to mature bone marrow and circulating B cell clearance by CD20 mAbs. Thus, isotype-specific mAb interactions with distinct FcγRs contribute significantly to the effectiveness of CD20 mAbs in vivo, which may have important clinical implications for CD20 and other mAb-based therapies.