Objective: With the depending research in biology, nanoscience and other fields, atomic force microscopy (AFM) is increasingly used in the field of biomaterials. AFM can reveal the mechanical properties of biological tissues at the nanoscale, which are often used as indicators of the physiological state and function of tissues. Methods: In this paper, the porcine esophagus was chosen as experimental sample to study the hardness properties of esophageal tissues at different loading rates, deflection and dwell time with AFM. Results: The results showed that the hardness of esophageal tissues at the nanoscale was strongly correlated with the loading rate and the deflection, which increased with the increasing loading rate and decreased with the increasing deflection of cantilever. The difference in the hardness was associated with the viscoelasticity and viscoplasticity of esophageal tissues, including contact stress, energy transition and strain plastic gradient. Conclusions: The results have important significance for clinical diagnosis, surgical operation and artificial material development, and reveal the changing law of the mechanical properties of the esophageal tissues from the microscale.