The insulin-like growth factor I receptor (IGF-IR) plays a critical role in cell proliferation and survival. We previously reported that a recombinant anti-IGF-IR antibody, scFv-Fc, consisting of 1H7 monoclonal antibody (mAb)-derived single chain antibody (scFv) and human IgG1 Fc, significantly suppressed breast tumor growth, and we proposed IGFIR down-regulation as a mechanism for tumor growth inhibition (Horm Metab Res. 2003; 35:836, Cancer Res. 2003; 63:627). This study used MCF-7 breast cancer cells to investigate the effects of anti-IGF-IR mAbs with various epitope specificities on IGF-IR downregulation and signaling pathways. Despite their differing effects on IGF-IR signaling, all five mAbs used down-regulated IGF-IR. Inhibitor experiments indicated that anti-IGF-IR mAbs induced internalization of IGF-IR from clathrin coated-pits. Pretreatment of MCF-7 cells with methylamine substantially reduced the antibody-mediated IGF-IR down-regulation while MG115 did not. Ubiquitination of IGF-IR did not occur in MCF-7 cells after mAb treatment. These results suggest that anti-IGF-IR antibodies with different epitope-specificities can cause internalization of IGF-IR from clathrin-coated pits and down-regulation via a lysosome-dependent pathway in an IGF-IR activation-independent manner.