Quencher-free CRISPR-based molecular detection using an amphiphilic DNA fluorescence probe

A novel amphiphilic DNA fluorescence probe enables sensitive detection of nucleic acids with results comparable to traditional PCR methods.

• The probe consists of a hydrophobic Cy5 fluorophore head and a hydrophilic single-stranded DNA tail.
• In the presence of target nucleic acid, the CRISPR-Cas12a enzyme cleaves the ssDNA tail, releasing the Cy5 head.
• The detection system is integrated with a paper-based microfluidic device, allowing for quencher-free quantification of nucleic acids.
• Fluorescence diffusion distance of the Cy5 head is inversely related to the concentration of target nucleic acid.
• Testing with human papillomavirus-16 (HPV-16) DNA demonstrated the system's analytical performance and clinical feasibility.

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