Rapid, visual detection of pathogen nucleic acids has broad applications in infection management. Here we present a modular detection platform, termed enzyme-assisted nanocomplexes for visual identification of nucleic acids (enVision). The system consists of an integrated circuit of enzyme–DNA nanostructures, which function as independent recognition and signaling elements, for direct and versatile detection of pathogen nucleic acids from infected cells. The built-in enzymatic cascades produce a rapid color readout for the naked eye; the assay is thus fast (<2 h), sensitive (<10 amol), and readily quantified with smartphones. When implemented on a configurable microfluidic platform, the technology demonstrates superior programmability to perform versatile computations, for detecting diverse pathogen targets and their virus–host genome integration loci. We further design the enVision platform for molecular-typing of infections in patient endocervical samples. The technology not only improves the clinical inter-subtype differentiation, but also expands the intra-subtype coverage to identify previously undetectable infections.

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Extracellular vesicles (EVs) are diverse, nanoscale membrane vesicles released by cells into the circulation. As an emerging class of circulating biomarkers, EVs contain a trove of molecular information and play important roles in mediating intercellular communication. These EV molecular cargoes are differentially organized in the vesicles; they could be inherited from the parent cells or bound to the EV membrane through surface interactions. While the inherited constituents could serve as cell surrogate biomarkers, extravesicular association could reflect structural states of the bound molecules, revealing distinct subpopulations with different biophysical and/or biochemical properties. Despite the clinical potential of EVs and their diverse contents, conventional sensing technologies have limited compatibility to reveal nanoscale EV features. Complementary analytical platforms are being developed to address these technical challenges and expand the biomedical applications of EVs, to establish novel correlations and empower new diagnostics. This article provides a perspective on recent developments in sensor technologies to profile the diverse contentsdifferent molecular types, quantities, and organizational statesin extracellular vesicles.

Accurate and accessible nucleic acid diagnostics is critical to reducing the spread of COVID-19 and resuming socioeconomic activities. Here, we present an integrated platform for the direct detection of SARS-CoV-2 RNA targets near patients. Termed e lectrochemical s ystem i ntegrating r econfigurable e nzyme-DNA n anostructures ( eSIREN ), the technology leverages responsive molecular nanostructures and automated microfluidics to seamlessly transduce target-induced molecular activation into an enhanced electrochemical signal. Through responsive enzyme-DNA nanostructures, the technology establishes a molecular circuitry that directly recognizes specific RNA targets and catalytically enhances signaling; only upon target hybridization, the molecular nanostructures activate to liberate strong enzymatic activity and initiate cascading reactions. Through automated microfluidics, the system coordinates and interfaces the molecular circuitry with embedded electronics; its pressure actuation and liquid-guiding structures improve not only analytical performance but also automated implementation. The developed platform establishes a detection limit of 7 copies of RNA target per μl, operates against the complex biological background of native patient samples, and is completed in <20 min at room temperature. When clinically evaluated, the technology demonstrates accurate detection in extracted RNA samples and direct swab lysates to diagnose COVID-19 patients.

Despite the importance of nucleic acid testing in managing the COVID-19 pandemic, current detection approaches remain limited due to their high complexity and extensive processing. Here, we describe a molecular nanotechnology that enables direct and sensitive detection of viral RNA targets in native clinical samples. The technology, termed catalytic amplification by transition-state molecular switch (CATCH), leverages DNA-enzyme hybrid complexes to form a molecular switch. By ratiometric tuning of its constituents, the multicomponent molecular switch is prepared in a hyperresponsive state—the transition state—that can be readily activated upon the binding of sparse RNA targets to turn on substantial enzymatic activity. CATCH thus achieves superior performance (~8 RNA copies/μl), direct fluorescence detection that bypasses all steps of PCR (<1 hour at room temperature), and versatile implementation (high-throughput 96-well format and portable microfluidic assay). When applied for clinical COVID-19 diagnostics, CATCH demonstrated direct and accurate detection in minimally processed patient swab samples.