Here we report an electrochemical sensor that can realize label-free cancer cell detection using the first clinical trial II used aptamer AS1411 and functionalized graphene. By taking advantages of AS1411 high binding affinity and specificity to the overexpressed nucleolin on the cancer cell surface, our developed electrochemical aptasensor can distinguish cancer cells and normal ones and detect as low as one thousand cells. With DNA hybridization technique, this E-DNA sensor can be regenerated and reusable for cancer cell detection. Our work gives a good example for label-free cancer cell detection based on aptamer and graphene-modified electrode.