Visualized Lateral Flow Assay for Dual Viral RNA Fragment Detection

In this technical note, we report an easy-to-produce,reverse-transcription-free,and protein-enzyme-free lateral flow assay for detection of viralRNA fragments by taking SARS-CoV-2 ORF1ab and N as target models.Catalytic hairpin assembly is utilized for dual RNA fragment orthogonalreaction to generate copious amounts of opened hairpin duplexes, whichbridge DNA-modified gold nanoparticles and capture strands on thestrip to induce coloration. The dual RNA fragments are simultaneouslyvisualized during one time of sample flow, and single-base-mismatchednontarget sequences can be differentiated. The test strip can be flexiblyadapted to detect evolutional SARS-CoV-2 variants such as Delta andOmicron. It also shows potential in visually detecting long-sequencevirus simulants and achieves a sensitivity comparable to that of RT-qPCRby incorporation with upstream sample amplification. The lateral flowassay should offer a convenient and reliable technique for viral nucleicacid detection.