In this work, we constructed an exonuclease III cleavage reaction-based isothermal amplification of nucleic acids with CRISPR/Cas12a-mediated pH-induced regenerative Electrochemiluminescence (ECL) biosensor for ultrasensitive and specific detection of SARS-CoV-2 nucleic acids for SARS-CoV-2 diagnosis. The triple-stranded nucleic acid in this biosensor has an extreme dependence on pH, which makes our constructed biosensor reproducible. This is essential for effective large-scale screening of SARS-CoV-2 in areas where resources are currently relatively scarce. Using this pH-induced regenerative biosensor, we detected the SARS-CoV-2 RdRp gene with a detection limit of 43.70 aM. In addition, the detection system has good stability and reproducibility, and we expect that this method may provide a potential platform for the diagnosis of COVID-19.
In this work, we propose a Ti 3 C 2 T x -based electrochemiluminescence (ECL) sensing platform, combined with a programmable entropy-driven cycling strategy and inverted DNA tetrahedral nanomachines to detect the SARS-CoV-2 RdRp gene. The use of Ti 3 C 2 T x in the ECL sensing platform provides unique advantages that improve the assay's sensitivity. Firstly, the SARS-CoV-2 RdRp gene triggers an entropy-driven cyclic amplification reaction to obtain a large number of Output strands. Subsequently, the Output strand reacts with an inverted DNA tetrahedral swing arm strand to start the self-driven process. Finally, the ferrocene (Fc)-labeled hairpin DNA strand of the inverted DNA tetrahedron is severed to achieve the signal change. Under optimal conditions, the ECL platform integrated with entropy-driven cycle amplification reaction and inverted DNA tetrahedral nanomachines obtained a wide range of 100 aM to 10 nM and a limit of detection (LOD) of 48.3 aM for the SARS-CoV-2 RdRp gene. Moreover, the recovery rates of 97% to 103.3% and 98.34% to 103% were obtained in real environments and human pharyngeal swabs, respectively. This ECL sensing platform, which combines an entropy-driven cycle amplification reaction and inverted DNA tetrahedral nanomachines, opens up new opportunities for the detection of other substances.
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