SARS-Cov-2 Spike-S1 Antigen Test Strip with High Sensitivity Endowed by High-Affinity Antibodies and Brightly Fluorescent QDs/Silica Nanospheres.

The extensive research into developing novel strategies for detecting respiratory syndrome coronavirus 2 (SARS-CoV-2) antigens in clinical specimens, especially the sensitive point-of-care testing method, is still urgently needed to reach rapid screening of viral infections. Herein, a new lateral flow immunoassay (LFIA) platform was reported for the detection of SARS-CoV-2 spike-S1 protein antigens, in which four sensitive and specific SARS-CoV-2 mouse monoclonal antibodies (MmAbs) were tailored by using quantum dot (QD)-loaded dendritic mesoporous silica nanoparticles modified further for achieving the -COOH group surface coating (named Q/S-COOH nanospheres). Importantly, compact QD adsorption was achieved in mesoporous channels of silica nanoparticles on account of highly accessible central-radial pores and electrostatic interactions, leading to significant signal amplification. As such, a limit of detection for SARS-CoV-2 spike-S1 testing was found to be 0.03 ng/mL, which is lower compared with those of AuNPs-LFIA (traditional colloidal gold nanoparticles, Au NPs) and enzyme-linked immunosorbent assay methods. These results show that optimizing the affinity of antibody and the intensity of fluorescent nanospheres simultaneously is of great significance to improve the sensitivity of LFIA.

A Novel SPR Immunosensor Based on Dual Signal Amplification Strategy for Detection of SARS-CoV-2 Nucleocapsid Protein.

Since the global outbreak of coronavirus disease 2019 (COVID-19), it has spread rapidly around the world. The nucleocapsid (N) protein is one of the most abundant SARS-CoV-2 proteins. Therefore, a sensitive and effective detection method for SARS-CoV-2 N protein is the focus of research. Here, we developed a surface plasmon resonance (SPR) biosensor based on the dual signal-amplification strategy of Au@Ag@Au nanoparticles (NPs) and graphene oxide (GO). Additionally, a sandwich immunoassay was utilized to sensitively and efficiently detect SARS-CoV-2 N protein. On the one hand, Au@Ag@Au NPs have a high refractive index and the capability to electromagnetically couple with the plasma waves propagating on the surface of gold film, which are harnessed for amplifying the SPR response signal. On the other hand, GO, which has the large specific surface area and the abundant oxygen-containing functional groups, could provide unique light absorption bands that can enhance plasmonic coupling to further amplify the SPR response signal. The proposed biosensor could efficiently detect SARS-CoV-2 N protein for 15 min and the detection limit for SARS-CoV-2 N protein was 0.083 ng/mL, with a linear range of 0.1 ng/mL~1000 ng/mL. This novel method can meet the analytical requirements of artificial saliva simulated samples, and the developed biosensor had a good anti-interference capability.

Recent Trends in Lateral Flow Immunoassays with Optical Nanoparticles.

Rapid, accurate, and convenient diagnosis is essential for effective disease management. Various detection methods, such as enzyme-linked immunosorbent assay, have been extensively used, with lateral flow immunoassay (LFIA) recently emerging as a major diagnostic tool. Nanoparticles (NPs) with characteristic optical properties are used as probes for LFIA, and researchers have presented various types of optical NPs with modified optical properties. Herein, we review the literature on LFIA with optical NPs for the detection of specific targets in the context of diagnostics.

Updates on the Biofunctionalization of Gold Nanoparticles for the Rapid and Sensitive Multiplatform Diagnosis of SARS-CoV-2 Virus and Its Proteins: From Computational Models to Validation in Human Samples.

Since the outbreak of the pandemic respiratory virus SARS-CoV-2 (COVID-19), academic communities and governments/private companies have used several detection techniques based on gold nanoparticles (AuNPs). In this emergency context, colloidal AuNPs are highly valuable easy-to-synthesize biocompatible materials that can be used for different functionalization strategies and rapid viral immunodiagnosis. In this review, the latest multidisciplinary developments in the bioconjugation of AuNPs for the detection of SARS-CoV-2 virus and its proteins in (spiked) real samples are discussed for the first time, with reference to the optimal parameters provided by three approaches: one theoretical, via computational prediction, and two experimental, using dry and wet chemistry based on single/multistep protocols. Overall, to achieve high specificity and low detection limits for the target viral biomolecules, optimal running buffers for bioreagent dilutions and nanostructure washes should be validated before conducting optical, electrochemical, and acoustic biosensing investigations. Indeed, there is plenty of room for improvement in using gold nanomaterials as stable platforms for ultrasensitive and simultaneous "in vitro" detection by the untrained public of the whole SARS-CoV-2 virus, its proteins, and specific developed IgA/IgM/IgG antibodies (Ab) in bodily fluids. Hence, the lateral flow assay (LFA) approach is a quick and judicious solution to combating the pandemic. In this context, the author classifies LFAs according to four generations to guide readers in the future development of multifunctional biosensing platforms. Undoubtedly, the LFA kit market will continue to improve, adapting researchers' multidetection platforms for smartphones with easy-to-analyze results, and establishing user-friendly tools for more effective preventive and medical treatments.

Alternative platform for COVID-19 diagnosis based on AuNP-modified lab-on-paper.

COVID-19 has caused global health problems, and so rapid diagnosis is crucial to slow spread of the disease. Herein, a novel lab-on-paper screening method for SARS-CoV-2 Omicron BA.2 variant was developed using a gold nanoparticle-based colorimetric biosensor along with sensitive detection of SARS-CoV-2 antigen using laser desorption ionization-mass spectrometry (LDI-MS). As a result of antigen-antibody interaction, in the presence of SARS-CoV-2 antigen the gold nanoparticles undergo aggregation and change color from red to light purple, allowing for rapid determination of SARS-CoV-2 antigen with the naked eye. Furthermore, the lab-on-paper method can be directly applied as a substrate for sensitive quantitation of SARS-CoV-2 antigen in saliva using LDI-MS without the use of a conventional organic matrix and sample preparation. LDI-MS offers early diagnosis with high sensitivity, rapidity without sample preparation and lower cost per test compared with reverse transcriptase-PCR, which is crucial for preventing mortality in patients with underlying conditions. This method showed linearity over 0.01-1 µg mL-1 covering the cut-off value of 0.048 µg mL-1 for COVID-19 detection in human saliva. Moreover, a colorimetric sensor for urea was also fabricated in-parallel, for prediction of COVID-19 severity in patients with chronic kidney disease. The color change upon increasing urea concentration directly reflected kidney damage, which is related to increasing risk of mortality among patients with COVID-19. Hence, this platform might be a potential device for non-invasive diagnosis of SARS-CoV-2 Omicron BA.2 variant, which is the variant of most concern because it is transmitted more rapidly than the original SARS-CoV-2 virus and the Delta variant.

A dual-labeled fluorescence quenching lateral flow assay based on one-pot enzyme-free isothermal cascade amplification for the rapid and sensitive detection of pathogens.

Rapid detection of nucleic acids is integral for clinical diagnostics, especially if a major public-health emergency occurs. However, such detection cannot be carried out efficiently in remote areas limited by medical resources. Herein, a dual-labeled fluorescence resonance energy transfer (FRET) lateral flow assay (LFA) based on one-pot enzyme-free cascade amplification was developed for rapid, convenient, and sensitive detection of open reading frame (ORF)1ab of severe acute respiratory syndrome-coronavirus-2. The catalyzed hairpin assembly (CHA) reaction of two well-designed hairpin probes was initiated by a target sequence and generated a hybridization chain reaction (HCR) initiator. Then, HCR probes modified with biotin were initiated to produce long DNA nanowires. After two-level amplification, the cascade-amplified product was detected by dual-labeled lateral flow strips. Gold nanoparticles (AuNPs)-streptavidin combined with the product and then ran along a nitrocellulose membrane under the action of capillary force. After binding with fluorescent microsphere-labeled-specific probes on the T line, a positive signal (red color) could be observed. Meanwhile, AuNPs could quench the fluorescence of the T line, and an inverse relationship between fluorescence intensity and the concentration of the CHA-HCR-amplified product was formed. The proposed strategy achieved a satisfactory limit of detection of 2.46 pM for colorimetric detection and 174 fM for fluorescent detection, respectively. Benefitting from the features of being one-pot, enzyme-free, low background, high sensitivity, and selectivity, this strategy shows great potential in bioanalysis and clinical diagnostics upon further development.

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.

Antibacterial, antiviral, and biodegradable collagen network mask for effective particulate removal and wireless breath monitoring.

Functional face masks that can effectively remove particulate matter and pathogens are critical to addressing the urgent health needs arising from industrial air pollution and the COVID-19 pandemic. However, most commercial masks are manufactured by tedious and complicated network-forming procedures (e.g., meltblowing and electrospinning). In addition, the materials used (e.g., polypropylene) have significant limitations such as a lack of pathogen inactivation and degradability, which can cause secondary infection and serious environmental concerns if discarded. Here, we present a facile and straightforward method for creating biodegradable and self-disinfecting masks based on collagen fiber networks. These masks not only provide superior protection against a wide range of hazardous substances in polluted air, but also address environmental concerns associated with waste disposal. Importantly, collagen fiber networks with naturally existing hierarchical microporous structures can be easily modified by tannic acid to improve its mechanical characteristics and enable the in situ production of silver nanoparticles. The resulting masks exhibit excellent antibacterial (>99.99%, 15 min) and antiviral (>99.999%, 15 min) capabilities, as well as high PM2.5 removal efficiency (>99.9%, 30 s). We further demonstrate the integration of the mask into a wireless platform for respiratory monitoring. Therefore, the smart mask has enormous promise for combating air pollution and contagious viruses, managing personal health, and alleviating waste issues caused by commercial masks.

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.

Synergistically functionalized molybdenum disulfide-reduced graphene oxide nanohybrid based ultrasensitive electrochemical immunosensor for real sample analysis of COVID-19.

Herein, we have proposed a straightforward and label-free electrochemical immunosensing strategy supported on a glassy carbon electrode (GCE) modified with a biocompatible and conducting biopolymer functionalized molybdenum disulfide-reduced graphene oxide (CS-MoS2/rGO) nanohybrid to investigate the SARS-CoV-2 virus. CS-MoS2/rGO nanohybrid-based immunosensor employs recombinant SARS-CoV-2 Spike RBD protein (rSP) that specifically identifies antibodies against the SARS-CoV-2 virus via differential pulse voltammetry (DPV). The antigen-antibody interaction diminishes the current responses of the immunosensor. The obtained results indicate that the fabricated immunosensor is extraordinarily capable of highly sensitive and specific detection of the corresponding SARS-CoV-2 antibodies with a LOD of 2.38 zg mL-1 in phosphate buffer saline (PBS) samples over a broad linear range between 10 zg mL-1-100 ng mL-1. In addition, the proposed immunosensor can detect attomolar concentrations in spiked human serum samples. The performance of this immunosensor is assessed using actual serum samples from COVID-19-infected patients. The proposed immunosensor can accurately and substantially differentiate between (+) positive and (-) negative samples. As a result, the nanohybrid can provide insight into the conception of Point-of-Care Testing (POCT) platforms for cutting-edge infectious disease diagnostic methods.

Bifunctional Au@Pt/Au nanoparticles as electrochemiluminescence signaling probes for SARS-CoV-2 detection.

A novel immunosensor based on electrochemiluminescence resonance energy transfer (ECL-RET) for the sensitive determination of N protein of the SARS-CoV-2 coronavirus is described. For this purpose, bifunctional core@shell nanoparticles composed of a Pt-coated Au core and finally decorated with small Au inlays (Au@Pt/Au NPs) have been synthesized to act as ECL acceptor, using [Ru (bpy)3]2+ as ECL donor. These nanoparticles are efficient signaling probes in the immunosensor developed. The proposed ECL-RET immunosensor has a wide linear response to the concentration of N protein of the SARS-CoV-2 coronavirus with a detection limit of 1.27 pg/mL. Moreover, it has a high stability and shows no response to other proteins related to different virus. The immunosensor has achieved the quantification of N protein of the SARS-CoV-2 coronavirus in saliva samples. Results are consistent with those provided by a commercial colorimetric ELISA kit. Therefore, the developed immunosensor provides a feasible and reliable tool for early and effective detection of the virus to protect the population.

Enhanced Single-Particle Collision Electrochemistry at Polysulfide-Functionalized Microelectrodes for SARS-CoV-2 Detection.

Single-particle collision electrochemistry (SPCE) has shown great promise in biosensing applications due to its high sensitivity, high flux, and fast response. However, a low effective collision frequency and a large number of interfering substances in complex matrices limit its broad application in clinical samples. Herein, a novel and universal SPCE biosensor was proposed to realize sensitive detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) based on the collision and oxidation of single silver nanoparticles (Ag NPs) on polysulfide-functionalized gold ultramicroelectrodes (Ps-Au UMEs). Taking advantage of the strong interaction of the Ag-S bond, collision and oxidation of Ag NPs on the Ps-Au UME surface could be greatly promoted to generate enhanced Faraday currents. Compared with bare Au UMEs, the collision frequency of Ps-Au UMEs was increased by 15-fold, which vastly improved the detection sensitivity and practicability of SPCE in biosensing. By combining magnetic separation, liposome encapsulation release, and DNAzyme-assisted signal amplification, the SPCE biosensor provided a dynamic range of 5 orders of magnitude for spike proteins with a detection limit of 6.78 fg/mL and a detection limit of 21 TCID50/mL for SARS-CoV-2. Furthermore, SARS-CoV-2 detection in nasopharyngeal swab samples of infected patients was successfully conducted, indicating the potential of the SPCE biosensor for use in clinically relevant diagnosis.

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.

Quercetin-Mediated Silver Nanoparticle Formation for the Colorimetric Detection of Infectious Pathogens Coupled with Loop-Mediated Isothermal Amplification.

Here, quercetin-mediated silver nanoparticle (AgNP) formation combined with loop-mediated isothermal amplification (LAMP) was introduced to colorimetrically detect two major infectious pathogens, SARS-CoV-2 and Enterococcus faecium, using a foldable PMMA microdevice. The nitrogenous bases of LAMP amplicons can readily form a complex with Ag+ ions, and the catechol moiety in quercetin, which acted as a reducing agent, could be chelated with Ag+ ions, resulting in the easy electron transfer from the oxidant to the reductant and producing brown-colored AgNPs within 5 min. The introduced method exhibited higher sensitivity than agarose gel electrophoresis due to more active redox centers in quercetin. The detection limit was attained at 101 copies µL-1 and 101 CFU mL-1 for SARS-CoV-2 RNA and E. faecium, respectively. A foldable microdevice made of two pieces of PMMA that fully integrates DNA extraction, amplification, and detection processes was fabricated to establish practical applicability. On one PMMA, DNA extraction was performed in a reaction chamber inserted with an FTA card, and then LAMP reagents were added for amplification. Silver nitrate was added to the reaction chamber after LAMP. On the other PMMA, quercetin-soaked paper discs loaded in the detection chamber were folded toward the reaction chamber for colorimetric detection. An intense brown color was produced within 5 min when heated at 65 °C. The introduced colorimetric assay, which is highly favorable for laboratory and on-site applications, could be a valuable alternative to conventional methods for detecting infectious diseases, given its unique principle, simplicity, and naked-eye detection.

CRISPR-Cas13a-powered electrochemical biosensor for the detection of the L452R mutation in clinical samples of SARS-CoV-2 variants.

Since the end of 2019, a highly contagious disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has deprived numerous lives worldwide, called COVID-19. Up to date, omicron is the latest variant of concern, and BA.5 is replacing the BA.2 variant to become the main subtype rampaging worldwide. These subtypes harbor an L452R mutation, which increases their transmissibility among vaccinated people. Current methods for identifying SARS-CoV-2 variants are mainly based on polymerase chain reaction (PCR) followed by gene sequencing, making time-consuming processes and expensive instrumentation indispensable. In this study, we developed a rapid and ultrasensitive electrochemical biosensor to achieve the goals of high sensitivity, the ability of distinguishing the variants, and the direct detection of RNAs from viruses simultaneously. We used electrodes made of MXene-AuNP (gold nanoparticle) composites for improved sensitivity and the CRISPR/Cas13a system for high specificity in detecting the single-base L452R mutation in RNAs and clinical samples. Our biosensor will be an excellent supplement to the RT-qPCR method enabling the early diagnosis and quick distinguishment of SARS-CoV-2 Omicron BA.5 and BA.2 variants and more potential variants that might arise in the future.

Lateral Flow Immunoassay Coupled with Copper Enhancement for Rapid and Sensitive SARS-CoV-2 Nucleocapsid Protein Detection.

The coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory coronavirus 2 (SARS-CoV-2) is still raging all over the world. Hence, the rapid and sensitive screening of the suspected population is in high demand. The nucleocapsid protein (NP) of SARS-CoV-2 has been selected as an ideal marker for viral antigen detection. This study describes a lateral flow immunoassay (LFIA) based on colloidal gold nanoparticles for rapid NP antigen detection, in which sensitivity was improved through copper deposition-induced signal amplification. The detection sensitivity of the developed LFIA for NP antigen detection (using certified reference materials) under the optimized parameters was 0.01 µg/mL and was promoted by three orders of magnitude to 10 pg/mL after copper deposition signal amplification. The LFIA coupled with the copper enhancement technique has many merits such as low cost, high efficiency, and high sensitivity. It provides an effective approach to the rapid screening, diagnosis, and monitoring of the suspected population in the COVID-19 outbreak.

Gold-silver alloy hollow nanoshells-based lateral flow immunoassay for colorimetric, photothermal, and SERS tri-mode detection of SARS-CoV-2 neutralizing antibody.

Although many approaches have been developed for the quick assessment of SARS-CoV-2 infection, few of them are devoted to the detection of the neutralizing antibody, which is essential for assessing the effectiveness of vaccines. Herein, we developed a tri-mode lateral flow immunoassay (LFIA) platform based on gold-silver alloy hollow nanoshells (Au-Ag HNSs) for the sensitive and accurate quantification of neutralizing antibodies. By tuning the shell-to-core ratio, the surface plasmon resonance (SPR) absorption band of the Au-Ag HNSs is located within the near infrared (NIR) region, endowing them with an excellent photothermal effect under the irradiation of optical maser at 808 nm. Further, the Raman reporter molecule 4-mercaptobenzoic acid (MBA) was immobilized on the gold-silver alloy nanoshell to obtain an enhanced SERS signal. Thus, these Au-Ag HNSs could provide colorimetric, photothermal and SERS signals, with which, tri-mode strips for SARS-CoV-2 neutralizing antibody detection were constructed by competitive immunoassay. Since these three kinds of signals could complement one another, a more accurate detection was achieved. The tri-mode LFIA achieved a quantitative detection with detection limit of 20 ng/mL. Moreover, it also successfully detected the serum samples from 98 vaccinated volunteers with 79 positive results, exhibiting great application value in neutralizing antibody 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.

The detection of IgG class antibodies against SARS-CoV-2 nucleocapsid protein by application of nanoparticles.

OBJECTIVE: Enzyme-linked immunosorbent assay (ELISA) is a widely used biochemical analytical method for the detection of a biomarker, through a specific antigen-antibody reaction. A common with ELISA is the amount of concrete biomarkers falling below the detection limit. Thus, the approach that will contribute to enhanced sensitivity of enzyme-linked immunosorbent assay is of great importance for medical practice. To address this issue, we used nanoparticles to improve the detection limit of traditional ELISA. MATERIALS AND METHODS: 80 samples were used, for which the presence of IgG antibodies against SARS-CoV-2 nucleocapsid protein were already determined qualitatively.  We tested the samples using an in vitro ELISA kit [SARS-CoV-2 IgG ELISA, COVG0949 (NovaTec, Leinfelden-Echterdingen, Germany)]. Additionally, we tested the same sample with the same ELISA kit but with the addition of 50 nm diameter citrate-capped silver nanoparticles. The reaction was performed, and data were calculated according to manufacturer guidelines. To measure ELISA results absorbance (optical density - OD) at 450 nm was read. RESULTS: Greater absorbance values have been revealed in case of silver nanoparticles application (66 cases, 82.5%, p<0.05). ELISA with application of nanoparticles classified 19 equivocal cases as positive and 3 equivocal ones as negative, 1 negative case as equivocal. CONCLUSIONS: Our findings suggest that nanoparticles can be used to improve the sensitivity of ELISA method and increase the detection limit. Thus, it is logical and desirable to enhance the sensitivity of ELISA method by application of nanoparticles; the approach is low cost and with a positive impact on accuracy.