Abstract Scanning acoustic microscopy (SAM) is a useful observational tool in cellular study as non-invasive living observation is feasible, unlike in conventional optical microscopy. In a previous study, cell morphological changes were successfully visualized using acoustic impedance measurements. These acoustic impedance changes correspond to cell elasticity, mainly reflecting changes in the cytoskeleton. In this study, we evaluate the elastic changes in murine breast cancer cell C127I during mitosis. C127I cells were cultured to ∼75% confluency before measurement, using a transducer with a central frequency of 320 MHz. Dynamic changes during mitosis were successfully mapped using SAM and confirmed by laser confocal microscopy. Cells in prometaphase, anaphase, and telophase, which could previously only be confirmed through immunostaining, were successfully visualized using SAM. This suggests that SAM is capable of distinguishing cells in different mitotic phases based on the changes in acoustic impedance.