High-efficiency upconversion luminescent quenching via defect-engineered Cu_3−xP for dual-aptamers-mediated tri-modal H1N1 lateral flow immunoassay

Early diagnosis of Influenza A virus is critical for influenza prevention and control by reducing severe illness and mortality. However, conventional lateral flow immunoassay (LFIA) is limited by low sensitivity, and luminescent LFIA is short of significant signal changes at low target concentrations. Herein, a luminescence quenching LFIA platform is developed by integrating upconversion nanoparticles, defect engineered Cu_3−xP, and dual aptamers to address above bottleneck via a synergistic mechanism. Density functional theory calculations indicates that in the new type Cu_3−xP quencher, Cu vacancies induce new electronic states at the valence band edge and create extra electron transition pathway, which boost Förster Resonance Energy Transfer and upconversion luminescence quenching efficiency. Therefore, the test line luminescence signal is effectively suppressed to establish a sensitive luminescence quenching LFIA, which is sensitive to signal changes. Cu_3−xP also exhibits broad UV-vis absorption for colorimetric and photothermal signals. The synergy luminescence-colorimetry-photothermal tri-modal analytical platform achieves a limit of detection of 0.907 ng/mL for H1N1 hemagglutinin. Moreover, two selected aptamers with stable and non-overlapping binding sites verified by molecular docking are critical for forming a sandwich structure, enabling high sensitivity toward H1N1 over H7N9, H9N2, H6N1, H7N7, and H5N1, along with good anti-interference ability. This work provides a scalable point of care influenza detection strategy balancing sensitivity and specificity, optimizing LFIA for on site viral diagnostics.