Cancer-specific monoclonal antibodies (CasMabs) that recognize cancer-specific antigens with in vivo antitumor efficacy are innovative therapeutic strategies for minimizing adverse effects. We previously established a cancer-specific anti-human epidermal growth factor receptor 2 (HER2) monoclonal antibody (mAb), HMab-250/HCasMab-2. In flow cytometry and immunohistochemistry, HMab-250 reacted with HER2-positive breast cancer cells but did not show reactivity to normal epithelial cells. In contrast, a clinically approved anti-HER2 mAb, trastuzumab, strongly recognizes both breast cancer and normal epithelial cells in flow cytometry. The human IgG version of HMab-250 (HMab-250-hG) possesses compatible in vivo antitumor effects against breast cancer xenografts to trastuzumab despite the lower affinity and effector activation than trastuzumab in vitro. This study compared the antibody-dependent cellular cytotoxicity (ADCC) and complement-dependent cellular cytotoxicity (CDC) between HMab-250-hG and trastuzumab. Both HMab-250-hG and trastuzumab showed ADCC activity against HER2-overexpressed Chinese hamster ovary -K1 and breast cancer cell lines (BT-474 and SK-BR-3) in the presence of human natural killer cells. Some tendency was observed where trastuzumab showed a more significant ADCC effect compared to HMab-250-hG. Importantly, HMab-250-hG exhibited superior CDC activity in these cells compared to trastuzumab. Similar results were obtained in the mouse IgG types of both HMab-250 and trastuzumab. These results suggest the different contributions of ADCC and CDC activities to the antitumor effects of HMab-250-hG and trastuzumab, and indicate a future direction for the clinical development of HMab-250-hG against HER2-positive tumors.