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.