CRISPR-Programmable Nanoparticle Transport in Salt-Gated Porous Media for Visual DNA Detection

Paper-based diagnostics typically treat porous cellulose as a passive capillary medium, with nanoparticle transport governed by channel geometry and capture chemistry. Here we show that CRISPR-programmable nanoparticle surface chemistry can actively regulate transport in paper through a salt-activated adhesion mechanism. Gold nanoparticles functionalized with hydrophobic Cy5-labeled DNA exhibit strong adhesion to cellulose fibers at elevated ionic strength (~1.5 M local NaCl generated upon rehydration of preloaded salt), whereas particles lacking hydrophobic domains migrate freely. This phenomenon enables a distance-based visual readout, in which the length of the colored nanoparticle front provides a semi-quantitative measure of DNA target concentration. Optical and electron microscopy reveal that nanoparticle immobilization arises from extensive fiber adhesion rather than pore occlusion or particle aggregation. As a proof of concept, coupling Cas12a-mediated cleavage of Cy5-labeled DNA probes to this salt-gated transport system enabled naked-eye, distance-based detection of Herpes simplex virus 1 (HSV-1) DNA with a limit of detection of 100 aM in ~45-60 min. Evaluation of 40 clinical samples showed excellent agreement with qPCR, achieving 96.2% sensitivity and 100% specificity. More broadly, this work establishes a general strategy for programming nanoparticle transport in porous media by encoding functionality directly into nanoparticle surface chemistry.