Simultaneous Detection of Five Infectious Diseases in a Single Strip: Oligo dT-Utilized Lateral Flow Immunoassay

The need for accurate and simultaneous diagnosis of multiple respiratory infectious diseases has become increasingly critical due to ongoing viral mutations and the similarity of symptoms among various viruses. Here, we have advanced our detection capabilities by developing a multiplex lateral flow immunoassay (LFA) platform that integrates oligonucleotides and antibodies, enabling the simultaneous detection of five respiratory viruses: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), Influenza A (FluA), Influenza B (FluB), Respiratory syncytial virus (RSV), and Adenovirus (ADV), on a single membrane. By applying the oligonucleotide and antibody-conjugated AuNPs, the platform enables highly sensitive and specific detection. In addition, signal amplification using RPA70A-conjugated gold nanoparticles that developed in the previous study can further be applied optionally for low-concentration biomarkers. Our interferences and cross-reactivity tests confirmed that these complexes do not produce false positives, substantiating the assay's utility in clinical settings. This platform, therefore, provides a robust solution for the precise and rapid diagnosis of complex viral infections, positioning it as suitable for application in pandemic response scenarios.