Ag⁺-Mediated DNA Nanomachine Cascade Nanomaterial Amplification Enable One-Pot Electrochemical Analysis of Circulating Tumor DNA

Circular tumor DNA (ctDNA) is a trace nucleic acid that functions as an essential tumor marker. In this context, the present study proposes a one-pot electrochemical analysis of ctDNA EGFR L858R in lung cancer leveraging a Ag⁺-mediated DNA nanosphere (I amplification) and cation exchange reaction (II amplification), and Cu²⁺ acts as a signal molecule. Once the target L858R exists, it specifically destroys the structure of DNA nanosphere@Ag⁺, and large amounts of Ag⁺ are released. After the addition of copper sulfide nanoparticles, Cu²⁺ can be replaced by a cation exchange reaction. Eventually, the electrochemical signal of Cu²⁺ is elevated. The analytical performance of the method is satisfactory, L858R can be detected in the linear range of 1 aM-1 fM with a detection limit of 0.3 aM. Furthermore, the system exhibits notable selectivity in differentiating base mismatch targets and other ctDNA sequences. The recovery rate of blood samples is between 95.5 and 105%. The electrochemical results from the analysis of 42 clinical blood samples are consistent with those of the quantitative real-time polymerase chain reaction, computed tomography, and pathology results. In summary, this novel strategy utilizes preprepared functional nucleic acid nanomaterials and cascade amplification, which is expected to contribute to the sensitive and expeditious detection of trace nucleic acids.