Label-free and amplified aptasensor for thrombin detection based on background reduction and direct electron transfer of hemin

In this work, we describe the development of a sensitive and label-free aptasensor for thrombin detection based on background noise reduction by exonuclease I (Exo I) and signal amplification by direct electron transfer (DET) of hemin. The thrombin binding aptamers (TBAs) are self-assembled on a sensing electrode. In the absence of the target thrombin, the TBAs are digested by Exo I, which avoids the association of hemin and significantly minimizes the background current noise. The presence of thrombin stabilizes the folded TBA G-quadruplex and prevents it from degrading by Exo I. The G-quadruplex bound hemin thus generates amplified signal output. In our sensing approach, the introduction of Exo I significantly enhances the signal to noise ratio of the sensor response and achieves sensitive detection of thrombin. Our new method is also coupled with good selectivity against other non-target proteins and thus holds great potential for the development of robust aptasensors for the detection of different types of targets.