Fluorescence-enhanced lateral flow nanosensor for dual miRNA detection: a rapid point-of-care tool for early diagnosis and severity monitoring of ischemic stroke

Abstract

Early and accurate diagnosis of ischemic stroke (IS) remains a clinical challenge due to the lack of rapid and sensitive tools for quantifying disease-specific biomarkers. Here, we present an automated dual-color lateral flow immunoassay (LFIA) platform that enables the simultaneous detection of two key miRNA biomarkers, miR-155 and miR-125b-5p, with ultra-high sensitivity and specificity. By employing hairpin DNA-functionalized fluorescent polystyrene (PS) microspheres as signal amplifiers, the assay achieves detection limits as low as 3.6 fM and 6.8 fM for miR-155 and miR-125b-5p, respectively, in both blood and cerebrospinal fluid samples from 168 IS patients. The entire detection process is completed within 15 minutes, allowing for equipment-free, on-site quantification. Integrated into a portable and user-friendly device, the platform is suitable for point-of-care and at-home testing. Validation against qRT-PCR demonstrated strong concordance, confirming the reliability of the LFIA system. This novel nanosensor not only facilitates early IS diagnosis but also supports real-time severity assessment, offering a transformative approach to personalized stroke management.