Net-Shaped DNA Nanostructures Designed for Rapid/Sensitive Detection and Potential Inhibition of the SARS-CoV-2 Virus.

We present a net-shaped DNA nanostructure (called "DNA Net" herein) design strategy for selective recognition and high-affinity capture of intact SARS-CoV-2 virions through spatial pattern-matching and multivalent interactions between the aptamers (targeting wild-type spike-RBD) positioned on the DNA Net and the trimeric spike glycoproteins displayed on the viral outer surface. Carrying a designer nanoswitch, the DNA Net-aptamers release fluorescence signals upon virus binding that are easily read with a handheld fluorimeter for a rapid (in 10 min), simple (mix-and-read), sensitive (PCR equivalent), room temperature compatible, and inexpensive (∼$1.26/test) COVID-19 test assay. The DNA Net-aptamers also impede authentic wild-type SARS-CoV-2 infection in cell culture with a near 1 × 103-fold enhancement of the monomeric aptamer. Furthermore, our DNA Net design principle and strategy can be customized to tackle other life-threatening and economically influential viruses like influenza and HIV, whose surfaces carry class-I viral envelope glycoproteins like the SARS-CoV-2 spikes in trimeric forms.

DNA Origami-Based Nanoprinting for the Assembly of Plasmonic Nanostructures with Single-Molecule Surface-Enhanced Raman Scattering.

Metallic nanocube ensembles exhibit tunable localized surface plasmon resonance to induce the light manipulation at the subwavelength scale. Nevertheless, precisely control anisotropic metallic nanocube ensembles with relative spatial directionality remains a challenge. Here, we report a DNA origami based nanoprinting (DOBNP) strategy to transfer the essential DNA strands with predefined sequences and positions to the surface of the gold nanocubes (AuNCs). These DNA strands ensured the specific linkages between AuNCs and gold nanoparticles (AuNPs) that generating the stereo-controlled AuNC-AuNP nanostructures (AANs) with controlled geometry and composition. By anchoring the single dye molecule in hot spot regions, the dramatic enhanced electromagnetic field aroused stronger surface enhanced Raman scattering (SERS) signal amplification. Our approach opens the opportunity for the fabrication of stereo-controlled metal nanostructures for designing highly sensitive photonic devices.

Au nanoparticles on two-dimensional MoS2 nanosheets as a photoanode for efficient photoelectrochemical miRNA detection.

MiRNAs are small regulatory RNAs that play crucial roles in the oncogenic state in various cancers and have shown highly promising clinical applications as plasma-based markers for cancer classification and prognostication. Due to their electroanalytical advantages, photoelectrochemical biosensors are a very attractive alternative technology for miRNA sensing and detection. In this work, we demonstrated a novel photoelectrochemical (PEC) sensor using the in situ grown Au nanoparticles/two-dimensional molybdenum disulfide (MoS2) nanosheet heterojunction (MoS2-AuNPs) on ITO glass as the photoanode (MoS2-AuNPs/ITO). AuNPs were used as a photoelectronic transfer promoter and DNA probe immobilization carrier as well. The thiol modified biotin DNA with a hairpin structure was tethered to the MoS2-AuNPs/ITO surface to form a specific capturing layer for miRNA detection. The biotin specific protein streptavidin was used as the signal amplifying species. This PEC sensor is structurally simple and possesses sensitivity and specificity toward miRNA. The CV and EIS responses were evaluated to monitor the PEC anode fabrication. The stability and reproducibility of this PEC design strategy were both evaluated before it was used in analyzing the samples of miRNA in human serum. Finally, we found that this PEC sensor displayed a broad detection linear range and a low detection limit of 4.21 fM, and it can excellently discriminate the mismatched miRNA. These findings pave the way for developing PEC sensors targeting miRNA by using noble metals/MoS2 heterojunctions.

Designer DNA nanostructures for viral inhibition.

Emerging viral diseases can substantially threaten national and global public health. Central to our ability to successfully tackle these diseases is the need to quickly detect the causative virus and neutralize it efficiently. Here we present the rational design of DNA nanostructures to inhibit dengue virus infection. The designer DNA nanostructure (DDN) can bind to complementary epitopes on antigens dispersed across the surface of a viral particle. Since these antigens are arranged in a defined geometric pattern that is unique to each virus, the structure of the DDN is designed to mirror the spatial arrangement of antigens on the viral particle, providing very high viral binding avidity. We describe how available structural data can be used to identify unique spatial patterns of antigens on the surface of a viral particle. We then present a procedure for synthesizing DDNs using a combination of in silico design principles, self-assembly, and characterization using gel electrophoresis, atomic force microscopy and surface plasmon resonance spectroscopy. Finally, we evaluate the efficacy of a DDN in inhibiting dengue virus infection via plaque-forming assays. We expect this protocol to take 2-3 d to complete virus antigen pattern identification from existing cryogenic electron microscopy data, ~2 weeks for DDN design, synthesis, and virus binding characterization, and ~2 weeks for DDN cytotoxicity and antiviral efficacy assays.

Designer DNA architecture offers precise and multivalent spatial pattern-recognition for viral sensing and inhibition.

DNA, when folded into nanostructures with a specific shape, is capable of spacing and arranging binding sites into a complex geometric pattern with nanometre precision. Here we demonstrate a designer DNA nanostructure that can act as a template to display multiple binding motifs with precise spatial pattern-recognition properties, and that this approach can confer exceptional sensing and potent viral inhibitory capabilities. A star-shaped DNA architecture, carrying five molecular beacon-like motifs, was constructed to display ten dengue envelope protein domain III (ED3)-targeting aptamers into a two-dimensional pattern precisely matching the spatial arrangement of ED3 clusters on the dengue (DENV) viral surface. The resulting multivalent interactions provide high DENV-binding avidity. We show that this structure is a potent viral inhibitor and that it can act as a sensor by including a fluorescent output to report binding. Our molecular-platform design strategy could be adapted to detect and combat other disease-causing pathogens by generating the requisite ligand patterns on customized DNA nanoarchitectures.

Antibody-Functionalized MoS2 Nanosheets for Targeted Photothermal Therapy of Staphylococcus aureus Focal Infection.

Bacterial biofilm-related diseases cause serious hazard to public health and bring great challenge to the traditional antibiotic treatment. Photothermal therapy (PTT) has been recognized as a promising alternative solution. However, the therapeutic efficacy of PTT is often compromised by the collateral damage to normal tissues due to the lack of bacteria-targeting capability. Here, a Staphylococcus aureus (S. aureus)-targeted PTT nanoagent is prepared based on antibody (anti-protein A IgG), polydopamine (PDA), and PEG-SH (thiolated poly (ethylene glycol)) functionalized MoS2 nanosheets (MoS2@PDA-PEG/IgG NSs, MPPI NSs). The PDA was used as bio-nano interface to facilitate the covalent conjugation of antibody and PEG-SH onto the surface of MoS2 NSs via facile catechol chemistry. Targeted PTT of MPPI NSs shows excellent inactivation efficiency of larger than 4 log (>99.99%) to S. aureus both in biofilms (in vitro) and in infected tissues (in vivo) without causing damage to normal mammalian cells. By contrast, non-targeted PTT of MoS2@PDA-PEG NSs (MPP NSs) only kills S. aureus by <90% in vitro and <50% in vivo. As a result, S. aureus focal infection in mice healed much faster after PTT of MPPI NSs than that of MPP NSs. The superiority of targeted PTT may originate from the efficient accumulation and close binding of PTT agents to bacterial cells. Therefore, MPPI NSs with bacteria-targeting capability are promising photothermal agents for effective treatment of S. aureus focal infection.

Single-Step Organization of Plasmonic Gold Metamaterials with Self-Assembled DNA Nanostructures.

Self-assembled DNA nanostructures hold great promise as nanoscale templates for organizing nanoparticles (NPs) with near-atomistic resolution. However, large-scale organization of NPs with high yield is highly desirable for nanoelectronics and nanophotonic applications. Here, we design five-strand DNA tiles that can readily self-assemble into well-organized micrometer-scale DNA nanostructures. By organizing gold nanoparticles (AuNPs) on these self-assembled DNA nanostructures, we realize the fabrication of one- and two-dimensional Au nanostructures in single steps. We further demonstrate the one-pot synthesis of Au metamaterials for highly amplified surface-enhanced Raman Scattering (SERS). This single-step and high-yield strategy thus holds great potential for fabricating plasmonic metamaterials.

Cavity-Type DNA Origami-Based Plasmonic Nanostructures for Raman Enhancement.

DNA origami has been established as addressable templates for site-specific anchoring of gold nanoparticles (AuNPs). Given that AuNPs are assembled by charged DNA oligonucleotides, it is important to reduce the charge repulsion between AuNPs-DNA and the template to realize high yields. Herein, we developed a cavity-type DNA origami as templates to organize 30 nm AuNPs, which formed dimer and tetramer plasmonic nanostructures. Transmission electron microscopy images showed that high yields of dimer and tetramer plasmonic nanostructures were obtained by using the cavity-type DNA origami as the template. More importantly, we observed significant Raman signal enhancement from molecules covalently attached to the plasmonic nanostructures, which provides a new way to high-sensitivity Raman sensing.