Validation of Rapid and Economic Colorimetric Nanoparticle Assay for SARS-CoV-2 RNA Detection in Saliva and Nasopharyngeal Swabs.

Even with the widespread uptake of vaccines, the SARS-CoV-2-induced COVID-19 pandemic continues to overwhelm many healthcare systems worldwide. Consequently, massive scale molecular diagnostic testing remains a key strategy to control the ongoing pandemic, and the need for instrument-free, economic and easy-to-use molecular diagnostic alternatives to PCR remains a goal of many healthcare providers, including WHO. We developed a test (Repvit) based on gold nanoparticles that can detect SARS-CoV-2 RNA directly from nasopharyngeal swab or saliva samples with a limit of detection (LOD) of 2.1 × 105 copies mL-1 by the naked eye (or 8 × 104 copies mL-1 by spectrophotometer) in less than 20 min, without the need for any instrumentation, and with a manufacturing price of <$1. We tested this technology on 1143 clinical samples from RNA extracted from nasopharyngeal swabs (n = 188), directly from saliva samples (n = 635; assayed by spectrophotometer) and nasopharyngeal swabs (n = 320) from multiple centers and obtained sensitivity values of 92.86%, 93.75% and 94.57% and specificities of 93.22%, 97.96% and 94.76%, respectively. To our knowledge, this is the first description of a colloidal nanoparticle assay that allows for rapid nucleic acid detection at clinically relevant sensitivity without the need for external instrumentation that could be used in resource-limited settings or for self-testing.

Generalization of Self-Assembly Toward Differently Shaped Colloidal Nanoparticles for Plasmonic Superlattices.

Periodic superlattices of noble metal nanoparticles  have demonstrated superior plasmonic properties compared to randomly distributed plasmonic arrangements due to near-field coupling and constructive far-field interference. Here, a chemically driven, templated self-assembly process of colloidal gold nanoparticles is investigated and optimized, and the technology is extended toward a generalized assembly process for variously shaped particles, such as spheres, rods, and triangles. The process yields periodic superlattices of homogenous  nanoparticle clusters on a centimeter scale. Electromagnetically simulated absorption spectra and corresponding experimental extinction measurements demonstrate excellent agreement in the far-field for all particle types and different lattice periods. The electromagnetic simulations reveal the specific nano-cluster near-field behavior, predicting the experimental findings provided by surface-enhanced Raman scattering measurements. It turns out that periodic arrays of spherical nanoparticles produce higher surface-enhanced Raman scattering enhancement factors than particles with less symmetry as a result of very well-defined strong hotspots.

Mechanically Tunable Lattice-Plasmon Resonances by Templated Self-Assembled Superlattices for Multi-Wavelength Surface-Enhanced Raman Spectroscopy.

Lattice plasmons, i.e., diffractively coupled localized surface plasmon resonances, occur in long-range ordered plasmonic nanostructures such as 1D and 2D periodic lattices. Such far-field coupled resonances can be employed for ultrasensitive surface-enhanced Raman spectroscopy (SERS), provided they are spectrally matched to the excitation wavelength. The spectral positions of lattice plasmon modes critically depend on the lattice period and uniformity, owing to their pronounced sensitivity to structural disorder. We report the fabrication of superlattices by templated self-assembly of gold nanoparticles on a flexible support, with tunable lattice-plasmon resonances by means of macroscopic strain. We demonstrate that the highest SERS performance is achieved by matching the lattice plasmon mode to the excitation wavelength, by post-assembly fine-tuning of long-range structural parameters. Both asymmetric and symmetric lattice deformations can be used to adapt a single lattice structure to both red-shifted and blue-shifted excitation lines, as exemplified by lattice expansion and contraction, respectively. This proof-of-principle study represents a basis for alternative designs of adaptive functional nanostructures with mechanically tunable lattice resonances using strain as a macroscopic control parameter.

Preventing Memory Effects in Surface-Enhanced Raman Scattering Substrates by Polymer Coating and Laser-Activated Deprotection.

The development of continuous monitoring systems requires in situ sensors that are capable of screening multiple chemical species and providing real-time information. Such in situ measurements, in which the sample is analyzed at the point of interest, are hindered by underlying problems derived from the recording of successive measurements within complex environments. In this context, surface-enhanced Raman scattering (SERS) spectroscopy appears as a noninvasive technology with the ability of identifying low concentrations of chemical species as well as resolving dynamic processes under different conditions. To this aim, the technique requires the use of a plasmonic substrate, typically made of nanostructured metals such as gold or silver, to enhance the Raman signal of adsorbed molecules (the analyte). However, a common source of uncertainty in real-time SERS measurements originates from the irreversible adsorption of (analyte) molecules onto the plasmonic substrate, which may interfere in subsequent measurements. This so-called "SERS memory effect" leads to measurements that do not accurately reflect varying conditions of the sample over time. We introduce herein the design of plasmonic substrates involving a nonpermeable poly(lactic-co-glycolic acid) (PLGA) thin layer on top of the plasmonic nanostructure, toward controlling the adsorption of molecules at different times. The polymeric layer can be locally degraded by irradiation with the same laser used for SERS measurements (albeit at a higher fluence), thereby creating a micrometer-sized window on the plasmonic substrate available to molecules present in solution at a selected measurement time. Using SERS substrates coated with such thermolabile polymer layers, we demonstrate the possibility of performing over 10,000 consecutive measurements per substrate as well as accurate continuous monitoring of analytes in microfluidic channels and biological systems.