Temperature and pH tolerance ratiometric aptasensor: Efficiently self-calibrating electrochemical detection of aflatoxin B1.

Ratiometric electrochemical aptasensors based on the two different redox reporters can hardly achieve the efficient self-calibration, as a result of variations between reporters. In this work, we reported a ratiometric electrochemical aptasensor using ferrocene as a single redox reporter (Fc) to achieve self-calibrating electrochemical detection of aflatoxin B1 (AFB1). The hybrid duplex of Fc-labeled AFB1 aptamer (Fc-Apt) and Fc-labeled assistant DNA (Fc-aDNA) was designed to construct the sensing interface. Such DNA structure adopted the fixed molar ratio (1:1) and locations (the distances to electrode) of these two types of Fc (i.e., Fc-Apt and Fc-aDNA). In this way, the ratio of redox currents from Fc-Apt (I) and Fc-aDNA (I) kept stable under the varied temperatures (15-40 °C) and pH values (4-10). Moreover, values of I and I under the different conditions can be calculated simply by recording the total current of Fc (I). For analysis, the recognition of AFB1 by Fc-Apt led to the striping of their complex from the aptasensor, resulting in a decrease in I while I remained stable. Consequently, the developed aptasensor using the ratio of I/I as a yardstick offered a linear range of 0.1-10000 pg mL with a detection limit of 0.012 pg mL for the detection of AFB1. The applicability of aptasensor was validated by corn sample analysis, which exhibited comparable reliability and accuracy of gold standard method, i.e., HPLC-MS/MS. Our work provided an efficient strategy to fabricate high-performance ratiometric electrochemical aptasensors.