A simple and enzyme-free method for sensitive p53 analysis based on DNAzyme-mediated signal amplification.

There is an urgent demand for a simple yet extremely accurate biosensor to analyze tumorigenesis. Herein, we present a novel fluorescent and enzyme-free approach for detecting p53 gene cascading proximity ligation-mediated catalytic hairpin assembly and DNAzyme-assisted signal reaction. When the target p53 gene is present, the interaction between p53 and L1 and L2 chains initiates catalytic hairpin assembly and subsequently exposes DNAzyme in the P3 probe. The exposed DNAzyme binds with the loop region of the P4 probe and generates a nicking site, resulting in the release of a significant amount of ATMND that is conjugated in the stem section of P4. This leads to an amplified fluorescence response, which serves as a fluorescence signal for the detection of the p53 gene. This method allows for the accurate and sensitive identification of the p53 gene, exhibiting a linear reaction range of 1 fM to 1 nM, with a limit of detection as low as 0.23 fM. Furthermore, this fluorescent method has been utilized for the examination of clinical samples with a favorable recovery rate. Crucially, this versatile platform may be expanded to analyze different targets by changing the corresponding recognition unit, showing great potential for point-of-care testing in tumorigenesis analysis.