Ultrasensitive sensing platform based on AuFeCoNiCu high-entropy alloy nanoparticle-DNA walker dual signal amplification for electrochemical detection of miRNA-21.

Overcoming the critical limitation of poor sensitivity in electrochemical biosensors for early breast cancer diagnosis, an innovative approach is presented. We developed a robust coordination strategy, utilizing serine-functionalized graphene quantum dots (SGQDs), to synthesize AuFeCoNiCu high-entropy alloy nanoparticles (HEA NPs). The resulting AuFeCoNiCu HEA NPs (32 ± 1.7 nm) exhibit single a face centered cubic (FCC) phase, good elemental homogeneity, and graphene surface modification. This unique structure confers a significantly enhanced catalytic activity (> 12-fold higher than pure Au NPs) and a superior affinity towards polar electrolytes. Furthermore, these HEA NPs were integrated with a DNA walker circuit to construct one electrochemical biosensor for ultrasensitive detection of miRNA-21. The target miRNA-21 triggers the DNA walker process, immobilizing ferrocene molecules on the electrode surface generating a measurable electrochemical signal. This dual-amplification strategy (HEA NP catalysis + DNA walker) achieved an unprecedented sensitivity: a linear current response (at 0.22 V) over an extraordinary range (1 × 10⁻⁰ M to 1 × 10⁻ M) and a record-low detection limit of 3.4 × 10⁻ M. This represents 2-3 orders of magnitude sensitivity improvement over the state-of-the-art sensors, successfully demonstrated in serum analysis.