Copper selenide nanomaterial mediated light-driven RPA-SDA cascade amplification combined with photothermal test...

The rapid and sensitive on-site detection of the methicillin resistance gene (mecA) is crucial for monitoring antimicrobial resistance in dairy products. Herein, we developed a novel, fully integrated detection platform by coupling light-driven (LD) recombinase polymerase amplification (RPA) with strand displacement amplification (SDA) and a photothermal lateral flow test strip (LFTS), mediated by copper selenide (CS) nanomaterials. This cascade system leverages the excellent photothermal conversion property of CS, which under 808 nm laser irradiation provide localized heating to drive both RPA and SDA isothermally, eliminating the need for conventional temperature control equipment. The dual-amplification strategy significantly enhanced sensitivity, with the LD system achieving a 1.9-fold higher efficiency than water-bathed amplification within 30 min. Amplicons were quantitatively detected using a gold-modified CS-based photothermal LFTS, where captured nanoparticles generated a measurable temperature increase (ΔT). The assay demonstrated a wide linear range from 3.9 to 3.9 × 10⁶ copies/μL with a limit of detection of 0.43 copies/μL, exhibited high specificity against non-target antibiotic resistance genes (sul1, sul2, nuc), and showed good storage stability. When applied to spiked raw milk samples, recovery rates ranged from 84.7% to 112.3%, correlating well with qPCR results. The entire workflow from amplification to readout was completed within 35 min in a closed-tube format, offering a powerful, rapid, and equipment-free strategy for on-site monitoring of drug-resistant genes in complex food matrices.