Ochratoxin A (OTA) is a highly toxic food contaminant and is harmful to human beings. Herein, a ratiometric electrochemical aptasensor based on a DNA tetrahedral nanomaterial (NTH) was developed in combination with the signal tag of a zirconium metal-organic framework (UiO-66) for the detection of OTA. In the sensor, UiO-66 and a [Fe(CN)6]3-/4- electrolyte solution were used as the signal probe and the internal reference probe, respectively. In the presence of OTA, the OTA aptamer was released from the electrode due to the specific binding of OTA. Thus, signal probe P1 labeled-UiO-66 was captured on the electrode surface by hybridization with DNA NTH. Since signal probe P1 labeled-UiO-66 was close to the electrode, it leads to an increased signal current of UiO-66 at +0.9 V. As the conductivity of the modified electrode decreased, the current signal of [Fe(CN)6]3-/4- at +0.2 V also decreased. The proposed ratiometric electrochemical aptasensor could effectively eliminate external environmental influences and could avoid electrochemical background signals. The aptasensor demonstrated high specificity for OTA, and achieved a good linear range of 1 pg mL-1-100 ng mL-1 with a detection limit of 330 fg mL-1. The developed electrochemical aptamer biosensor effectively detected OTA in corn kernel samples, verifying its practical application for the determination of OTA in actual samples.