Synthesis and characterization of Au-decorated graphene oxide nanocomposite for magneto-electrochemical detection of SARS-CoV-2 nucleocapsid gene.

Fast and effective diagnosis is the first step in monitoring the current coronavirus 2 (CoV-2) pandemic. Herein, we establish a simple and sensitive electrochemical assay using magnetic nanocomposite and DNA sandwich probes to rapidly quantify the CoV-2 nucleocapsid (N) gene down to the 0.37 fM level. This assay uses a pair of specific DNA probes. The capture probe is covalently conjugated to Au-decorated magnetic reduced graphene oxide (AMrGO) nanocomposite for efficiently capturing target RNA. In contrast, the detection probe is linked to peroxidase for signal amplification. The probes target the COV-2 gene, allowing for specific magnetic separation, enzymatic signal amplification, and subsequent generation of voltammetric current with a total assay time of 45 min. The developed biosensor has high selectivity and can discriminate non-specific gene sequences. Synthetic COV-2 N-gene can be detected efficiently in serum and saliva, while 1-bp mismatch gene yielded a low response. The performance of the genosensor was good in an extensive linear range of 5 aM-50 pM. For synthetic N-gene, we achieved the detection limit of 0.37, 0.33, and 0.19 fM in human saliva, urine, and serum. This simple, selective, and sensitive genosensor could have various genetics-based biosensing and diagnostic applications.

Localized surface plasmon resonance biosensor chip surface modification and signal amplifications toward rapid and sensitive detection of COVID-19 infections.

We developed a multi-pronged approach to enhance the detection sensitivity of localized surface plasmon resonance (LSPR) sensor chips to detect SARS-CoV-2. To this end, poly(amidoamine) dendrimers were immobilized onto the surface of LSPR sensor chips to serve as templates to further conjugate aptamers specific for SARS-CoV-2. The immobilized dendrimers were shown to reduce surface nonspecific adsorptions and increase capturing ligand density on the sensor chips, thereby improving detection sensitivity. To characterize the detection sensitivity of the surface-modified sensor chips, SARS-CoV-2 spike protein receptor-binding domain was detected using LSPR sensor chips with different surface modifications. The results showed that the dendrimer-aptamer modified LSPR sensor chip exhibited a limit of detection (LOD) of 21.9 pM, a sensitivity that was 9 times and 152 times more sensitive than the traditional aptamer- or antibody-based LSPR sensor chips, respectively. In addition, detection sensitivity was further improved by combining rolling circle amplification product and gold nanoparticles to further amplify the detection signals by increasing both the target mass and plasmonic coupling effects. Using pseudo SARS-CoV-2 viral particles as detection targets, we demonstrated that this combined signal intensification approach further enhanced the detection sensitivity by 10 folds with a remarkable LOD of 148 vp/mL, making it one of the most sensitive SARS-CoV-2 detection assays reported to date. These results highlight the potential of a novel LSPR-based detection platform for sensitive and rapid detection of COVID-19 infections, as well as other viral infections and point-of-care applications.

PdPtRu trimetallic nanozymes and application to electrochemical immunosensor for sensitive SARS-COV-2 antigen detection.

Herein, a ternary PdPtRu nanodendrite as novel trimetallic nanozyme was reported, which possessed excellent peroxidase-like activity as well as electro-catalytic activity on account of the synergistic effect between the three metals. Based on the excellent electro-catalytic activity of trimetallic PdPtRu nanozyme toward the reduction of H2O2, the trimetallic nanozyme was applied to construct a brief electrochemical immunosensor for SARS-COV-2 antigen detection. Concretely, trimetallic PdPtRu nanodendrite was used to modify electrode surface, which not only generated high reduction current of H2O2 for signal amplification, but also provided massive active sites for capture antibody (Ab1) immobilization to construct immunosensor. In the presence of target SARS-COV-2 antigen, SiO2 nanosphere labeled detection antibody (Ab2) composites were introduced on the electrode surface according sandwich immuno-reaction. Due to the inhibitory effect of SiO2 nanosphere on the current signal, the current signal was decreased with the increasing target SARS-COV-2 antigen concentration. As a result, the proposed electrochemical immunosensor presented sensitive detection of SARS-COV-2 antigen with linear range from 1.0 pg/mL to 1.0 µg/mL and limit of detection down to 51.74 fg/mL. The proposed immunosensor provide a brief but sensitive antigen detection tool for rapid diagnosis of COVID-19.

Ultra-sensitive and specific detection of pathogenic nucleic acids using composite-excited hyperfine plasma spectroscopy combs sensitized by Au nanoarrays functionalized with 2D Ta2C-MXene.

Accurate and rapid screening techniques on a population scale are crucial for preventing and managing epidemics like COVID-19. The standard gold test for nucleic acids in pathogenic infections is primarily the reverse transcription polymerase chain reaction (RT-PCR). However, this method is not suitable for widespread screening due to its reliance on large-scale equipment and time-consuming extraction and amplification processes. Here, we developed a collaborative system that combines high-load hybridization probes targeting N and OFR1a with Au NPs@Ta2C-M modified gold-coated tilted fiber Bragg grating (TFBG) sensors to enable direct nucleic acid detection. Multiple activation sites of SARS-CoV-2 were saturable modified on the surface of a homogeneous arrayed AuNPs@Ta2C-M/Au structure based on a segmental modification approach. The combination of hybrid probe synergy and composite polarisation response in the excitation structure results in highly specific hybridization analysis and excellent signal transduction of trace target sequences. The system demonstrates excellent trace specificity, with a limit of detection of 0.2 pg/mL, and achieves a rapid response time of 1.5 min for clinical samples without amplification. The results showed high agreement with the RT-PCR test (Kappa index = 1). And the gradient-based detection of 10-in-1 mixed samples exhibits high-intensity interference immunity and excellent trace identification. Therefore, the proposed synergistic detection platform has a good tendency to curb the global spread of epidemics such as COVID-19.

Electrochemical sensing of dual biomolecules in live cells and whole blood samples: A flexible gold wire-modified copper-organic framework-based hybrid composite.

For clinical research, the precise measurement of hydrogen peroxide (H2O2) and glucose (Glu) is of paramount importance, due to their imbalanced concentrations in blood glucose, and reactive oxygen species (ROS) play a huge role in COVID-19 viral disease. It is critical to construct and develop a simple, rapid, flexible, long-term, and sensitive detection of H2O2 and glucose. In this paper, we have developed a unique morphological structure of MOF(Cu) on a single-walled carbon nanotube-modified gold wire (swnt@gw). Highly designed frameworks with nanotube composites enhance electron rate-transfer behavior while extending conductance and electroactive surface area.The composite sensing system delivers wide linear-range concentrations, low detection limit, and interference-free performance in co-existence with other biomolecules and metal ions. Endogenous quantitative tracking of H2O2 was performed in macrophage live-cells with the help of a strong stimulator lipopolysaccharide.The composite device was effectively utilized for the measurement of H2O2 and glucose in turbid samples of whole blood and milk samples without a pretreatment process. The practical results of biofluids showed favorable voltammetric results and acceptance recovery percentage levels between 97.49 and 98.88%. Finally, a flexible MOF-based hybrid system may provide a suitable detection platform in the construction of electro-biosensors and hold potential promise for clinical-sensory applications.

Empirical Optimization of Peptide Sequence and Nanoparticle Colloidal Stability: The Impact of Surface Ligands and Implications for Colorimetric Sensing.

Surface ligands play a critical role in controlling and defining the properties of colloidal nanocrystals. These aspects have been exploited to design nanoparticle aggregation-based colorimetric sensors. Here, we coated 13-nm gold nanoparticles (AuNPs) with a large library of ligands (e.g., from labile monodentate monomers to multicoordinating macromolecules) and evaluated their aggregation propensity in the presence of three peptides containing charged, thiolate, or aromatic amino acids. Our results show that AuNPs coated with the polyphenols and sulfonated phosphine ligands were good choices for electrostatic-based aggregation. AuNPs capped with citrate and labile-binding polymers worked well for dithiol-bridging and π-π stacking-induced aggregation. In the example of electrostatic-based assays, we stress that good sensing performance requires aggregating peptides of low charge valence paired with charged NPs with weak stability and vice versa. We then present a modular peptide containing versatile aggregating residues to agglomerate a variety of ligated AuNPs for colorimetric detection of the coronavirus main protease. Enzymatic cleavage liberates the peptide segment, which in turn triggers NP agglomeration and thus rapid color changes in <10 min. The protease detection limit is 2.5 nM.

Label-free integrated microfluidic plasmonic biosensor from vertical-cavity surface-emitting lasers for SARS-CoV-2 receptor binding domain protein detection.

The nanoplasmonic sensor of the nanograting array has a remarkable ability in label-free and rapid biological detection. The integration of the nanograting array with the standard vertical-cavity surface-emitting lasers (VCSEL) platform can achieve a compact and powerful solution to provide on-chip light sources for biosensing applications. Here, a high sensitivity and label-free integrated VCSELs sensor was developed as a suitable analysis technique for COVID-19 specific receptor binding domain (RBD) protein. The gold nanograting array is integrated on VCSELs to realize the integrated microfluidic plasmonic biosensor of on-chip biosensing. The 850 nm VCSELs are used as a light source to excite the localized surface plasmon resonance (LSPR) effect of the gold nanograting array to detect the concentration of attachments. The refractive index sensitivity of the sensor is 2.99 × 106 nW/RIU. The aptamer of RBD was modified on the surface of the gold nanograting to detect the RBD protein successfully. The biosensor has high sensitivity and a wide detection range of 0.50 ng/mL - 50 µg/mL. This VCSELs biosensor provides an integrated, portable, and miniaturized idea for biomarker detection.

Introduction of Multilayered Dual-Signal Nanotags into a Colorimetric-Fluorescent Coenhanced Immunochromatographic Assay for Ultrasensitive and Flexible Monitoring of SARS-CoV-2.

Timely, accurate, and rapid diagnosis of SARS-CoV-2 is a key factor in controlling the spread of the epidemic and guiding treatments. Herein, a flexible and ultrasensitive immunochromatographic assay (ICA) was proposed based on a colorimetric/fluorescent dual-signal enhancement strategy. We first fabricated a highly stable dual-signal nanocomposite (SADQD) by continuously coating one layer of 20 nm AuNPs and two layers of quantum dots onto a 200 nm SiO2 nanosphere to provide strong colorimetric signals and enhanced fluorescence signals. Two kinds of SADQD with red and green fluorescence were conjugated with spike (S) antibody and nucleocapsid (N) antibody, respectively, and used as dual-fluorescence/colorimetric tags for the simultaneous detection of S and N proteins on one test line of ICA strip, which can not only greatly reduce the background interference and improve the detection accuracy but also achieve a higher colorimetric sensitivity. The detection limits of the method for target antigens via colorimetric and fluorescence modes were as low as 50 and 2.2 pg/mL, respectively, which were 5 and 113 times more sensitive than those from the standard AuNP-ICA strips, respectively. This biosensor will provide a more accurate and convenient way to diagnose COVID-19 in different application scenarios.

Designing and fabrication of electrochemical nano-biosensor for the fast detection of SARS-CoV-2-RNA.

SARS-CoV-2 caused a global panic among populations. Rapid diagnostic procedures for the virus are crucial for disease control. Thus, the designed signature probe from a highly conserved region of the virus was chemically immobilized onto the nanostructured-AuNPs/WO3-screen printed electrodes. Different concentrations of the matched oligonucleotides were spiked to test the specificity of the hybridization affinity whereas the electrochemical impedance spectroscopy was used for tracking the electrochemical performance. After a full assay optimization, limits of detection and quantification were calculated based on linear regression and were valued at 298 and 994 fM, respectively. Further, the high performance of the fabricated RNA-sensor chips was confirmed after testing the interference status in the presence of the mismatched oligos in one nucleotide and completely one. Worthy to mention that the single-stranded matched oligos can be hybridized to the immobilized probe in 5 min at room temperature. The designed disposable sensor chips are capable of detecting the virus genome directly. Therefore, the chips are a rapid tool for SARS-CoV-2 detection.

Resorc[4]arene Modifiers for Supramolecular Site-Directed Immobilization of Antibodies on Multi-Walled Carbon Nanotubes.

One of the main problems in developing immunosensors featuring carbon nanotubes (CNTs) is immobilizing antibodies (Abs) onto the CNT surface to afford selective binding to target antigens (Ags). In this work, we developed a practical supramolecular Ab conjugation strategy based on resorc[4]arene modifiers. To improve the Ab orientation on the CNTs surface and optimizing the Ab/Ag interaction, we exploited the host-guest approach by synthesizing two newly resorc[4]arene linkers R1 and R2 via well-established procedures. The upper rim was decorated with eight methoxyl groups to promote selective recognition of the fragment crystallizable (Fc ) region of the Ab. Moreover, the lower rim was functionalized with 3-bromopropyloxy or 3-azidopropiloxy substituents to bind the macrocycles on the multi-walled carbon nanotubes (MWCNTs) surface. Accordingly, several chemical modifications of MWCNTs were evaluated. After the morphological and electrochemical characterization of nanomaterials, the resorc[4]arene-modified MWCNTs were deposited onto a glassy carbon electrode surface to evaluate their potential applicability for label-free immunosensor development. The most promising system showed an improved electrode active area (AEL ) of almost 20 % and a site-oriented immobilization of the SARS-CoV-2 spike protein S1 antibody (Ab-SPS1). The developed immunosensor revealed a good sensitivity (23.64 µA mL ng-1 cm-2 ) towards the SPS1 antigen and a limit of detection (LOD) of 1.01 ng mL-1 .

Detection of live SARS-CoV-2 virus and its variants by specially designed SERS-active substrates and spectroscopic analyses.

A method using label-free surface enhanced Raman spectroscopy (SERS) based on substrate design is provided for an early detection and differentiation of spike glycoprotein mutation sites in live SARS-CoV-2 variants. Two SERS-active substrates, Au nanocavities (Au NCs) and Au NPs on porous ZrO2 (Au NPs/pZrO2), were used to identify specific peaks of A.3, Alpha, and Delta variants at different concentrations and demonstrated the ability to provide their SERS spectra with detection limits of 0.1-1.0% (or 104-5 copies/mL). Variant identification can be achieved by cross-examining reference spectra and analyzing the substrate-analyte relationship between the suitability of the analyte upon the hotspot(s) formed at high concentrations and the effective detection distance at low concentrations. Mutation sites on the S1 chain of the spike glycoprotein for each variant may be related and distinguishable. This method does not require sample preprocessing and therefore allows for fast screening, which is of high value for more comprehensive and specific studies to distinguish upcoming variants.

Trends in Application of SERS Substrates beyond Ag and Au, and Their Role in Bioanalysis.

This article compares the applications of traditional gold and silver-based SERS substrates and less conventional (Pd/Pt, Cu, Al, Si-based) SERS substrates, focusing on sensing, biosensing, and clinical analysis. In recent decades plethora of new biosensing and clinical SERS applications have fueled the search for more cost-effective, scalable, and stable substrates since traditional gold and silver-based substrates are quite expensive, prone to corrosion, contamination and non-specific binding, particularly by S-containing compounds. Following that, we briefly described our experimental experience with Si and Al-based SERS substrates and systematically analyzed the literature on SERS on substrate materials such as Pd/Pt, Cu, Al, and Si. We tabulated and discussed figures of merit such as enhancement factor (EF) and limit of detection (LOD) from analytical applications of these substrates. The results of the comparison showed that Pd/Pt substrates are not practical due to their high cost; Cu-based substrates are less stable and produce lower signal enhancement. Si and Al-based substrates showed promising results, particularly in combination with gold and silver nanostructures since they could produce comparable EFs and LODs as conventional substrates. In addition, their stability and relatively low cost make them viable alternatives for gold and silver-based substrates. Finally, this review highlighted and compared the clinical performance of non-traditional SERS substrates and traditional gold and silver SERS substrates. We discovered that if we take the average sensitivity, specificity, and accuracy of clinical SERS assays reported in the literature, those parameters, particularly accuracy (93-94%), are similar for SERS bioassays on AgNP@Al, Si-based, Au-based, and Ag-based substrates. We hope that this review will encourage research into SERS biosensing on aluminum, silicon, and some other substrates. These Al and Si based substrates may respond efficiently to the major challenges to the SERS practical application. For instance, they may be not only less expensive, e.g., Al foil, but also in some cases more selective and sometimes more reproducible, when compared to gold-only or silver-only based SERS substrates. Overall, it may result in a greater diversity of applicable SERS substrates, allowing for better optimization and selection of the SERS substrate for a specific sensing/biosensing or clinical application.

Silent Antibodies Start Talking: Enhanced Lateral Flow Serodiagnosis with Two-Stage Incorporation of Labels into Immune Complexes.

The presence of pathogen-specific antibodies in the blood is widely controlled by a serodiagnostic technique based on the lateral flow immunoassay (LFIA). However, its common one-stage format with an antigen immobilized in the binding zone of a test strip and a nanodispersed label conjugated with immunoglobulin-binding proteins is associated with risks of very low analytical signals. In this study, the first stage of the immunochromatographic serodiagnosis was carried out in its traditional format using a conjugate of gold nanoparticles with staphylococcal immunoglobulin-binding protein A and an antigen immobilized on a working membrane. At the second stage, a labeled immunoglobulin-binding protein was added, which enhanced the coloration of the bound immune complexes. The use of two separated steps, binding of specific antibodies, and further coloration of the formed complexes, allowed for a significant reduction of the influence of non-specific immunoglobulins on the assay results. The proposed approach was applied for the serodiagnosis using a recombinant RBD protein of SARS-CoV-2. As a result, an increase in the intensity of test zone coloration by more than two orders of magnitude was demonstrated, which enabled the significant reduction of false-negative results. The diagnostic sensitivity of the LFIA was 62.5% for the common format and 100% for the enhanced format. Moreover, the diagnostic specificity of both variants was 100%.

Carbon Dots Conjugated Antibody as an Effective FRET-Based Biosensor for Progesterone Hormone Screening.

In this work, carbon dots (CDs) were synthesized by a one-step hydrothermal method using citric acid and ethylene diamine, and covalently functionalized with antibodies for the sensing of progesterone hormone. The structural and morphological analysis reveals that the synthesized CDs are of average size (diameter 8-10 nm) and the surface functionalities are confirmed by XPS, XRD and FT-IR. Further graphene oxide (GO) is used as a quencher due to the fluorescence resonance energy transfer (FRET) mechanism, whereas the presence of the analyte progesterone turns on the fluorescence because of displacement of GO from the surface of CDs effectively inhibiting FRET efficiency due to the increased distance between donor and acceptor moieties. The linear curve is obtained with different progesterone concentrations with 13.8 nM detection limits (R2 = 0.974). The proposed optical method demonstrated high selectivity performance in the presence of structurally resembling interfering compounds. The PL intensity increased linearly with the increased progesterone concentration range (10-900 nM) under the optimal experimental parameters. The developed level-free immunosensor has emerged as a potential platform for simplified progesterone analysis due to the high selectivity performance and good recovery in different samples of spiked water.

A photonic resonator interferometric scattering microscope for label-free detection of nanometer-scale objects with digital precision in point-of-use environments.

Label-free detection and digital counting of nanometer-scaled objects such as nanoparticles, viruses, extracellular vesicles, and protein molecules enable a wide range of applications in cancer diagnostics, pathogen detection, and life science research. Here, we report the design, implementation, and characterization of a compact Photonic Resonator Interferometric Scattering Microscope (PRISM) designed for point-of-use environments and applications. The contrast of interferometric scattering microscopy is amplified through a photonic crystal surface, upon which scattered light from an object combines with illumination from a monochromatic source. The use of a photonic crystal substrate for interferemetric scattering microscopy results in reduced requirements for high-intensity lasers or oil-immersion objectives, thus opening a pathway toward instruments that are more suitable for environments outside the optics laboratory. The instrument incorporates two innovative elements that facilitate operation on a desktop in ordinary laboratory environments by users that do not have optics expertise. First, because scattering microscopes are extremely sensitive to vibration, we incorporated an inexpensive but effective solution of suspending the instrument's main components from a rigid metal framework using elastic bands, resulting in an average of 28.7 dBV reduction in vibration amplitude compared to an office desk. Second, an automated focusing module based on the principle of total internal reflection maintains the stability of image contrast over time and spatial position. In this work, we characterize the system's performance by measuring the contrast from gold nanoparticles with diameters in the 10-40 nm range and by observing various biological analytes, including HIV virus, SARS-CoV-2 virus, exosome, and ferritin protein.

Electrochemical Biosensor for the Determination of Specific Antibodies against SARS-CoV-2 Spike Protein.

In this article, we report the development of an electrochemical biosensor for the determination of the SARS-CoV-2 spike protein (rS). A gold disc electrode was electrochemically modified to form the nanocrystalline gold structure on the surface. Then, it was further altered by a self-assembling monolayer based on a mixture of two alkane thiols: 11-mercaptoundecanoic acid (11-MUA) and 6-mercapto-1-hexanol (6-MCOH) (SAMmix). After activating carboxyl groups using a N-(3-dimethylaminopropyl)-N'-ethyl-carbodiimide hydrochloride and N-hydroxysuccinimide mixture, the rS protein was covalently immobilized on the top of the SAMmix. This electrode was used to design an electrochemical sensor suitable for determining antibodies against the SARS-CoV-2 rS protein (anti-rS). We assessed the association between the immobilized rS protein and the anti-rS antibody present in the blood serum of a SARS-CoV-2 infected person using three electrochemical methods: cyclic voltammetry, differential pulse voltammetry, and potential pulsed amperometry. The results demonstrated that differential pulse voltammetry and potential pulsed amperometry measurements displayed similar sensitivity. In contrast, the measurements performed by cyclic voltammetry suggest that this method is the most sensitive out of the three methods applied in this research.

A label-free electrochemical DNA biosensor used a printed circuit board gold electrode (PCBGE) to detect SARS-CoV-2 without amplification.

The emergence of coronavirus disease 2019 (COVID-19) motivates continuous efforts to develop robust and accurate diagnostic tests to detect severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). Detection of viral nucleic acids provides the highest sensitivity and selectivity for diagnosing early and asymptomatic infection because the human immune system may not be active at this stage. Therefore, this work aims to develop a label-free electrochemical DNA biosensor for SARS-CoV-2 detection using a printed circuit board-based gold substrate (PCBGE). The developed sensor used the nucleocapsid phosphoprotein (N) gene as a biomarker. The DNA sensor-based PCBGE was fabricated by self-assembling a thiolated single-stranded DNA (ssDNA) probe onto an Au surface, which performed as the working electrode (WE). The Au surface was then treated with 6-mercapto-1-hexanol (MCH) before detecting the target N gene to produce a well-oriented arrangement of the immobilized ssDNA chains. The successful fabrication of the biosensor was characterized using cyclic voltammetry (CV), electrochemical impedance spectroscopy (EIS), and atomic force microscopy (AFM). The DNA biosensor performances were evaluated using a synthetic SARS-CoV-2 genome and 20 clinical RNA samples from healthy and infected individuals through EIS. The developed DNA biosensor can detect as low as 1 copy per µL of the N gene within 5 minutes with a LOD of 0.50 µM. Interestingly, the proposed DNA sensor could distinguish the expression of SARS-CoV-2 RNA in a patient diagnosed with COVID-19 without any amplification technique. We believe that the proposed DNA sensor platform is a promising point-of-care (POC) device for COVID-19 viral infection since it offers a rapid detection time with a simple design and workflow detection system, as well as an affordable diagnostic assay.

Recent Advances in Colorimetric Sensors Based on Gold Nanoparticles for Pathogen Detection.

Infectious pathogens cause severe threats to public health due to their frightening infectivity and lethal capacity. Rapid and accurate detection of pathogens is of great significance for preventing their infection. Gold nanoparticles have drawn considerable attention in colorimetric biosensing during the past decades due to their unique physicochemical properties. Colorimetric diagnosis platforms based on functionalized AuNPs are emerging as a promising pathogen-analysis technique with the merits of high sensitivity, low-cost, and easy operation. This review summarizes the recent development in this field. We first introduce the significance of detecting pathogens and the characteristics of gold nanoparticles. Four types of colorimetric strategies, including the application of indirect target-mediated aggregation, chromogenic substrate-mediated catalytic activity, point-of-care testing (POCT) devices, and machine learning-assisted colorimetric sensor arrays, are systematically introduced. In particular, three biomolecule-functionalized AuNP-based colorimetric sensors are described in detail. Finally, we conclude by presenting our subjective views on the present challenges and some appropriate suggestions for future research directions of colorimetric sensors.

Ultrasensitive detection of SARS-CoV-2 S protein with aptamers biosensor based on surface-enhanced Raman scattering.

A rapid and accurate diagnostic modality is essential to prevent the spread of SARS-CoV-2. In this study, we proposed a SARS-CoV-2 detection sensor based on surface-enhanced Raman scattering (SERS) to achieve rapid and ultrasensitive detection. The sensor utilized spike protein deoxyribonucleic acid aptamers with strong affinity as the recognition entity to achieve high specificity. The spherical cocktail aptamers-gold nanoparticles (SCAP) SERS substrate was used as the base and Au nanoparticles modified with the Raman reporter molecule that resonates with the excitation light and spike protein aptamers were used as the SERS nanoprobe. The SCAP substrate and SERS nanoprobes were used to target and capture the SARS-CoV-2 S protein to form a sandwich structure on the Au film substrate, which can generate ultra-strong "hot spots" to achieve ultrasensitive detection. Analysis of SARS-CoV-2 S protein was performed by monitoring changes in SERS peak intensity on a SCAP SERS substrate-based detection platform. This assay detects S protein with a LOD of less than 0.7 fg mL-1 and pseudovirus as low as 0.8 TU mL-1 in about 12 min. The results of the simulated oropharyngeal swab system in this study indicated the possibility of it being used for clinical detection, providing a potential option for rapid and accurate diagnosis and more effective control of SARS-CoV-2 transmission.

SARS-CoV-2 proteins monitored by long-range surface plasmon field-enhanced Raman scattering with hybrid bowtie nanoaperture arrays and nanocavities.

The identification of biomacromolecules by using surface-enhanced Raman scattering (SERS) remains a challenge because of the near-field effect of traditional substrates. Long-range surface plasmon resonance (LRSPR) is a special type of surface optical phenomenon that provides higher electromagnetic field enhancement and longer penetration depth than conventional surface plasmon resonance. To break the limit of SERS detection distance and obtain a SERS substrate with increased enhancement ability, a bowtie nanoaperture array was sandwiched between two symmetric dielectric environments. Then, an Au mirror was inserted to form a metal-insulator-metal configuration. Finite-difference time-domain simulations revealed that numerous hybrid modes can be provided by this novel configuration (denoted as long-range SERS [LR-SERS] substrate). In particular, the LRSPR mode can be excited and reach the maximum value through the regulation of the polarizations of the incident light and the geometrical parameters of the LR-SERS substrate. The optimized LR-SERS substrate was then applied to detect SARS-CoV-2 spike (S) and nucleocapsid (N) proteins. This substrate displayed ultralow detection limits of ∼9.2 and ∼11.3 pg mL-1 for the S and N proteins, respectively. Moreover, with the help of principal component analysis and receiver operating characteristic methods, our fabricated sensors exhibited excellent selectivity and hold great potential for the diagnosis of SARS-CoV-2 proteins in real samples.