Dual-Aptamer Drift Canceling Techniques to Improve Long-Term Stability of Real-Time Structure-Switching Aptasensors

This paper presents a dual-aptamer scheme to mitigate signal drifts caused by structure-switching aptamers during long-term monitoring. Electrochemical aptamer-based (E-AB) biosensors have recently shown great potential for continuous in vivo monitoring. However, the accuracy of detection is often limited by signaling drifts. Traditional approaches rely on kinetic differential measurements (KDM) coupled with square-wave voltammetry to eliminate these drifts. Yet, we have discovered that KDM does not apply universally to all aptamers, as their responses at different SWV frequencies heavily rely on their structure-switching characteristics and the electron transfer (ET) kinetics of the redox reporters. In light of this, we propose a "dual-aptamer" scheme that utilizes two aptamers, each responding differently to the same target molecule to cancel out drift. These paired aptamers are identified through (1) screening from an existing pool of aptamers and (2) engineering the signaling behavior of the redox reporters. We demonstrate the differential signaling of the aptamer pair in the presence of ampicillin and ATP molecules and show that the pair exhibits similar drifts in undiluted goat serum. By implementing drift cancelation, sensor drift is reduced by a factor of 370. Additionally, the differential signaling enables an increased recording throughput by leveraging differential readout electronics. The authors believe that the proposed technique holds significant benefits for long-term in vivo monitoring.