A novel label-free and signal-on electrochemical aptasensor based on the autonomous assembly of hemin/G-quadruplex and direct electron transfer of hemin

In this study, we developed a signal-on and label-free aptasensor based on the direct electron transfer of hemin and signal amplification by target-catalyzed hairpin assembly followed by a hybridization chain reaction (HCR). Upon the addition of a target protein, it facilitates the opening of the hairpin structure of A1 and thus accelerates the hybridization between A1 and A2; the target protein was displaced from hairpin A1 with hairpin A2 through a process similar to DNA branch migration. The released target participates in the next hybridization process with A1. Finally, each target passes through many cycles, confining numerous A1 close to the gold electrode. Subsequently, the single-strand fragment on the electrode surface initiated HCR, resulting in the hybridization reaction to form double-strand DNA concatemers on the electrode surface. Consequently, hemin stacked into the G-quadruplex-forming region, and the hemin/G-quadruplex was formed, generating an amplified electrochemical signal by differential pulse voltammetry. In our sensing approach, the introduction of HCR significantly enhanced the signal of the sensor response. Moreover, the approach is free of any label conjugation step for signal amplification and simple and thus has great potential for the development of robust aptasensors.