Broad-Spectrum HIV-1 Detection and Neutralization via Multivalent Designer DNA Nanostructures

Early and accurate detection of HIV-1 remains a critical unmet need, particularly during the acute phase of infection when viral loads are low and transmission risk is highest. Here, we report a modular diagnostic and antiviral platform based on designer DNA nanostructures engineered for high-affinity recognition of the HIV-1 envelope glycoprotein (GP120). A custom DNA aptamer, termed HINT, was developed to bind GP120 across major HIV-1 subtypes (Groups M and P; subtypes A and B) with nanomolar affinity. To amplify binding strength, HINT aptamers were spatially patterned onto a net-shaped DNA nanostructure (DNA-Net _HINT ) that geometrically matches the trimeric GP120 spikes on the viral surface. Using multivalent interactions, the nanostructure enabled up to 10 ⁴ -fold improvement in binding affinity (sub-picomolar K _D ), confirmed by surface plasmon resonance. Integration of DNA-Net _HINT into a paper-based lateral flow assay produced a low-cost, saliva-compatible self-testing device capable of detecting intact HIV-1 virions at concentrations as low as 328 viral copies per test, outperforming commercial fourth-generation rapid diagnostic tests. In addition to its diagnostic capabilities, the DNA-Net _HINT construct exhibited potent antiviral activity, reducing pseudovirus infection with an EC ₅₀ of ∼1.8 nM, nearly 1,000-fold more effective than free aptamers. This work demonstrates a dual-function DNA nanotechnology platform that enables both ultrasensitive HIV-1 detection and entry inhibition. The approach is broadly applicable to other enveloped viruses and represents a promising step toward next-generation molecular theranostics for infectious disease management.