In this paper, a novel oscillating nano knife for single cell cutting at its native state was proposed. The nano knife with a small edge angle 5° was fabricated from a commercial AFM cantilever by focused ion beam (FIB) etching. The spring constant of the nano knife was calibrated by nanorobotic manipulation method. A piezo actuator was employed to generate the vibration of the nano knife. The oscillating amplitude response to the applied voltage was calculated based on the realtime image observation. Single yeast cell (W303) cutting using oscillating nano knife was performed based on the nanorobotic manipulation system inside an environment-SEM. The maximum cutting forces and the slices angle were measured from the deformation of the nano knife beam and the single cell. The result showed the oscillating nano knife was capable and effective for single cell cutting task.