In the antibody-based targeted therapies of B-cell lymphomas, complement-mediated cytotoxicity (CMC) is an important mechanism. CMC is activated after the binding of drugs (monoclonal antibodies) to tumor cells. The activation of CMC ultimately leads to the lysis of tumor cells. However, it remains poorly understood how CMC alters the morphology and mechanics of single tumor cells at the nanoscale. In recent years, nanoscopic observations of cellular behaviors with the use of atomic force microscopy (AFM) have contributed much to the field of cell biology. In this work, by combining AFM with fluorescence microscopy, the detailed changes in cellular ultra-microstructures and mechanical properties during the process of CMC were quantitatively investigated on single tumor cells. AFM imaging distinctly showed that the CMC effect could lead to the formation of nano holes on the tumor cells. Quantitative analysis of AFM images indicated that cell surface became lower and rougher after the CMC process. The cellular mechanics measurements showed that during the process of CMC cells firstly softened and finally stiffened, which was validated by dynamically monitoring the mechanical changes of single living cells during CMC. The experimental results provide novel insights into the antibody-dependent CMC.