Abstract: Objective: To construct a quantum dot-based electrochemical sensor amplified by terminal deoxynucleotidyl transferase (TdT) for the ultrasensitive detection of epithelial cell adhesion molecule (EpCAM) on the surface of tumor cells. Methods: A two-step hydrothermal method was adopted to synthesize DNAfunctionalized quantum dot (QD-DNA). TdT was used to construct extended long polymeric aptamers. The two components were hybridized to obtain conjugated probes, and an electrochemical sensor was further fabricated. The reproducibility, anti-interference capability and specificity of the sensor were systematically investigated; meanwhile, the phenotypic changes of MCF-7 cells during drug treatment were monitored. Results: The QDbased electrochemical sensor amplified by TdT was successfully fabricated, with a detection limit as low as 80 cells, showing favorable detection sensitivity, specificity, accuracy and anti-interference capability. The sensor could effectively detect the expression levels of EpCAM on different cell surfaces and dynamically monitor the phenotypic changes of tumor cells during drug treatment. Conclusion: The QD-based electrochemical sensor amplified by TdT can effectively distinguish tumor cells from normal cells and dynamically monitor the phenotypic changes of tumor cells during drug treatment, showing promising application potential in early tumor diagnosis and precise screening.