Visualized Lateral Flow Assay for Dual Viral RNA Fragment Detection

Area of Science:

• Biotechnology
• Molecular Biology
• Assay Development

Background:

• Accurate and rapid detection of viral RNA is crucial for public health.
• Existing methods like RT-qPCR require complex laboratory setups and trained personnel.
• There is a need for simpler, field-deployable diagnostic tools for viral RNA detection.

Purpose of the Study:

• To develop a novel, reverse-transcription-free, and enzyme-free lateral flow assay for detecting viral RNA fragments.
• To demonstrate the assay's capability in detecting SARS-CoV-2 RNA fragments and its adaptability to different variants.
• To evaluate the assay's sensitivity and potential for integration with sample amplification techniques.

Main Methods:

• Utilized catalytic hairpin assembly for dual RNA fragment orthogonal reactions to amplify detection signals.
• Employed DNA-modified gold nanoparticles and capture strands on a lateral flow strip for visual coloration.
• Targeted SARS-CoV-2 ORF1ab and N gene fragments as model RNA sequences.

Main Results:

• Successfully developed an easy-to-produce lateral flow assay for viral RNA detection.
• Achieved simultaneous visualization of dual RNA fragments with single-base-mismatch differentiation.
• Demonstrated adaptability to detect SARS-CoV-2 variants (Delta, Omicron) and long viral sequence simulants.
• Showcased sensitivity comparable to RT-qPCR when coupled with upstream sample amplification.

Conclusions:

• The developed lateral flow assay is a convenient, reliable, and adaptable technique for viral nucleic acid detection.
• This method eliminates the need for reverse-transcription and protein enzymes, simplifying the detection process.
• The assay holds significant potential for rapid, point-of-care viral diagnostics, especially in resource-limited settings.