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J Hazard Mater . A triplex-readout CRISPR-Cas12a multimodal biosensing platform for point-of-care detection of avian influenza H5N1

tetano

Editor, Senior Moderator
J Hazard Mater


. 2026 Jun 1:514:142562.
doi: 10.1016/j.jhazmat.2026.142562. Online ahead of print.
A triplex-readout CRISPR-Cas12a multimodal biosensing platform for point-of-care detection of avian influenza H5N1

Bin Guo[SUP] 1 [/SUP], Yang Li[SUP] 2 [/SUP], Mengxin Shi[SUP] 3 [/SUP], Jingyu Zhang[SUP] 4 [/SUP], Qingqing Wu[SUP] 3 [/SUP], Xiaokang Wu[SUP] 5 [/SUP], Ruifang An[SUP] 6 [/SUP], Fu Wang[SUP] 7 [/SUP]


Affiliations
Abstract

The highly pathogenic avian influenza A (H5N1) virus poses a significant zoonotic threat to public and animal health. Conventional detection methods often face limitations in complexity, time, and equipment requirements. In this study, we report a rapid and lightweight-equipment triplex diagnostic platform for H5N1 detection, integrating locked nucleic acid (LNA)-assisted target recognition, toehold-mediated strand displacement for signal amplification, and the high specificity of the CRISPR-Cas12a system. The developed assay achieved a detection limit of 3.7 × 10² copies/μL for H5N1 pseudovirus RNA within a 60-minute core detection process, and exhibited excellent specificity without cross-reactivity to other influenza subtypes or coronaviruses. Moreover, it successfully identified H5N1 in clinical swab samples, yielding consistent results across three independent readout formats: fluorescence, lateral flow strip, and a portable glucose meter. With advantages of low cost, rapid operation, and multimodal verification capability, this diagnostic system is well suited for point-of-care screening in resource-limited settings and represents a promising tool for frontline outbreak response.

Keywords: Avian influenza; CRISPR-Cas12a; Glucometer; Locked nucleic acid; Multimodal detection.

 
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