Nano-Mechanical DNA Devices Coupled With CRISPR-Cas12a for CA15-3 Detection

Accurate monitoring of cancer markers is crucial for clinical treatment and prognosis. CA15-3 activity levels are strongly associated with clinical progression of breast cancer, but their monitoring often relies on large instruments and professionals, and the process is time-consuming and costly. To address these concerns, we proposed an electrochemical biosensing strategy that integrated nano-mechanical DNA devices coupled with the CRISPR-Cas12a to drive molecularly gated functionalized substrates for the ultrasensitive detection of CA15-3. Specifically, Triple helical molecular switch (THMS) as a signal input switch to ensure target recognition specificity and the diffusion-limited 3D DNA walking machine coupled with CRISPR-Cas12a technology as signal amplification means. Based on the bimolecular dynamics model, the rate constants k₁ (1.40 × 10⁵ M^-1sec^-1) and k₂ (2.5 × 10⁴ M^-1sec^-1) of the GNP-PEG(+)/T 3D orbitals modified with positively charged SH-PEG-NH₂ are larger than those of unmodified orbitals, proving that nanointerface diffusion restriction effect can accelerate the toehold-mediated chain displacement reaction (TMDR). With the assistance of Co-N/C modified screen-printed electrode (SPE-Co-N/C) sensing interface, the calculated detection limit of CA15-3 is as low as 7.14 × 10^-6 U mL^-1. The proposed assay, which demonstrated satisfactory selectivity and reproducibility, and correlated highly with ELISA kit results, offered a promising tool for breast cancer early detection and therapeutic monitoring.