Chain hybridization-based CRISPR-lateral flow assay enables accurate gene visual detection

Area of Science:

• Biotechnology
• Molecular Biology
• Diagnostics

Background:

• CRISPR-Cas gene detection using lateral flow assays (CRISPR-LFA) shows promise for point-of-care testing.
• Conventional CRISPR-LFA methods often yield false-positive results due to reliance on immuno-based test strips.

Purpose of the Study:

• To develop a novel CRISPR-based lateral flow assay (CHLFA) that eliminates false positives.
• To establish a highly accurate and visualized gene detection platform for point-of-care applications.

Main Methods:

• Developed a Chain Hybridization-based Lateral Flow Assay (CHLFA) platform utilizing nucleic acid hybridization.
• Integrated CRISPR-Cas12a or CRISPR-Cas13a with GNP-probe hybridization, bypassing the need for immunoreactions.
• Assayed for target gene detection within 50 minutes, with sensitivity down to 1-10 gene copies.

Main Results:

• The CRISPR-CHLFA system successfully eliminated false-positive results common in conventional CRISPR-LFA.
• Achieved highly accurate visual detection of target-negative samples.
• Demonstrated 100% accuracy in detecting SARS-CoV-2 Omicron variant and Mycobacterium tuberculosis in clinical specimens.

Conclusions:

• The CRISPR-CHLFA platform provides a robust alternative for accurate, visualized gene detection.
• This technology is suitable for developing advanced point-of-care testing (POCT) biosensors.
• CHLFA offers a significant improvement over existing CRISPR-LFA methods for disease diagnostics.