Microfluidic fluorescence biosensor for quantitative detection of HER2 in serum using an antibody–aptamer sandwich assay

Early detection of cancer biomarkers requires analytical platforms capable of sensitive quantification in complex biological matrices. Here we report a microfluidic fluorescence biosensor for the quantification of the breast cancer biomarker Human epidermal growth factor receptor 2 in human serum. The system integrates a hybrid bead-based antibody–aptamer sandwich assay with thin-film hydrogenated amorphous silicon (a-Si:H) photodiodes and a dielectric interference filter, enabling compact on-chip fluorescence detection within a fully integrated microfluidic architecture. Protein G-functionalized microbeads enabled oriented immobilization of capture antibodies, while a fluorophore-labeled HER2 aptamer served as the detection probe, forming a hybrid antibody–aptamer recognition scheme that minimized non-specific interactions in serum matrices. The microfluidic architecture and washing direction were optimized to suppress residual unbound aptamer, significantly improving the signal-to-background ratio. Under optimized conditions, the biosensor exhibited a near-linear fluorescence response for HER2 concentrations in 90% human serum across clinically relevant levels, achieving a limit of detection of 7–8 ng mL -1 , below the clinical threshold of 15 ng mL -1 . Photodiode measurements were consistent with fluorescence microscopy, confirming reliable on-chip signal acquisition. These results demonstrate a scalable strategy that combines hybrid molecular recognition, microfluidic background suppression, and integrated thin-film photodetection within a compact fluorescence biosensing platform for quantitative biomarker analysis.