Magnetically Driven Dual-miRNA Framework Nucleic Acid Biosensing Platform for Precise Classification of Breast Cancer Subtypes

The significant variation in treatment strategies among breast cancer subtypes establishes precise early subtyping as a critical prerequisite for effective therapy. While molecular profiling offers accurate classification, developing a rapid and reliable detection method remains challenging. Hence, we constructed an integrated platform by integrating a DNA tetrahedral probe (DTP) and surface antiadhesive magnetic micro/nanorobots (MNRs), enabling rapid contact between the MNR-DTP system and target analytes. The probe enhances fluorescence via target-triggered strand displacement and CHA-mediated signal amplification, allowing breast cancer subtypes to be identified through distinct dual-color fluorescence patterns. Furthermore, magnetically driven MNRs improve detection efficiency by enhancing mass transfer, promoting mixing, and accelerating probe–target interactions. Experimental results demonstrate that this strategy markedly enhances fluorescence output, enables rapid detection of dual-miRNA signatures in different breast cell lines, and distinguishes expression heterogeneity at the single-cell level. The system shows high specificity and improved sensitivity, with limits of detection of 1.5 pM for miR-21 and 1.17 pM for miR-31. The biocompatibility and stability of the proposed MNR-DTP system make it a promising tool for early breast cancer subtype discrimination and multiplexed target recognition in complex biological settings.