Cas14a1-advanced LAMP for ultrasensitive and visual Pathogen diagnostic

Area of Science:

• Molecular Biology
• Biosensor Development
• Diagnostic Technologies

Background:

• Clustered regularly interspaced short palindromic repeats (CRISPR)/Cas systems are utilized in biosensors with isothermal amplification strategies (IAS) for enhanced sensitivity and specificity.
• Existing CRISPR-based biosensors often suffer from assay instability and complex manipulation, limiting their practical application in pathogen detection.

Purpose of the Study:

• To develop a stable and user-friendly assay for pathogen detection.
• To enhance the sensitivity and specificity of biosensors using CRISPR/Cas technology.
• To create a rapid and visual platform for bacterial genomic DNA detection.

Main Methods:

• A Cas14a1-advanced LAMP assay (CALA) was developed, integrating Rapid Extraction of Bacterial Genomic DNA (REBGD).
• Ultrasensitive and visual detection platforms were established using a single-stranded fluorescence reporter and ssDNA-gold nanoparticles (AuNPs) as signal indicators.
• The assay's performance was evaluated for precise bacterial detection and reliability in contrived samples.

Main Results:

• The CALA assay achieved an ultrasensitive limit of detection (LOD) of 10 aM for target genomic DNA.
• The developed platform demonstrated precise detection capabilities for bacteria.
• High reliability in pathogen diagnostics was validated using the rapid visual CALA platform on contrived samples.

Conclusions:

• The Cas14a1-advanced LAMP assay (CALA) offers a promising solution for ultrasensitive and visual pathogen detection.
• The integration of REBGD and CALA provides a rapid, stable, and reliable diagnostic tool.
• This technology shows significant potential for practical pathogen testing and diagnostics.