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1.
Dokl Biochem Biophys ; 506(1): 220-222, 2022 Oct.
Article in English | MEDLINE | ID: covidwho-2088453

ABSTRACT

Based on previous studies, two antibody-like molecules, monobodies, capable of high-affinity interaction with the SARS-CoV-2 nucleocapsid protein (dissociation constant of tens of nM) were selected. For delivery to target cells, genetically engineered constructs containing monobody and TAT peptide, placed either at the N- or C-terminus of the resulting polypeptide, were produced and expressed in E. coli. The construct with the highest affinity to the SARS-CoV-2 nucleocapsid protein was revealed with the use of thermophoresis technique. Cellular thermal shift assay demonstrated the ability of this construct to interact with the nucleocapsid protein within HEK293T cells transfected with the SARS-CoV-2 nucleocapsid protein fused to the mRuby3 fluorescent protein. Replacement of TAT peptide to S10 shuttle peptide, containing endosomolytic peptide, significantly improved the penetration of the construct into the target cells.


Subject(s)
COVID-19 , SARS-CoV-2 , Humans , Escherichia coli/genetics , Escherichia coli/metabolism , HEK293 Cells , Nucleocapsid Proteins/chemistry , Nucleocapsid Proteins/metabolism , Antibodies, Viral
2.
Sci Bull (Beijing) ; 2022 Oct 27.
Article in English | MEDLINE | ID: covidwho-2086702

ABSTRACT

Nucleocapsid (N) protein plays crucial roles in the life cycle of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), including the formation of ribonucleoprotein (RNP) complex with the viral RNA. Here we reported the crystal structures of the N-terminal domain (NTD) and C-terminal domain (CTD) of the N protein and an NTD-RNA complex. Our structures reveal a unique tetramer organization of NTD and identify a distinct RNA binding mode in the NTD-RNA complex, which could contribute to the formation of the RNP complex. We also screened small molecule inhibitors of N-NTD and N-CTD and discovered that ceftriaxone sodium, an antibiotic, can block the binding of RNA to NTD and inhibit the formation of the RNP complex. These results together could facilitate the further research of antiviral drug design targeting N protein.

3.
Embase; 26.
Preprint in English | EMBASE | ID: ppcovidwho-346543

ABSTRACT

While significant attention has been paid to the immunologic determinants of disease states associated with COVID-191,2, their contributions to post-acute sequelae of COVID-19 (PASC) remain less clear3-5. Due to the wide array of PASC presentations6, it is critical to understand if specific features of the disease are associated with discrete immune processes, and whether those processes may be therapeutically targeted. To this end, we performed wide immunologic and serological characterization of patients in the early recovery phase of COVID-19 across a breadth of symptomatic presentations. Using high-parameter proteomics screening and applied machine learning (ML), we identify clear signatures of immunologic activity between PASC patients and uncomplicated recovery, dominated by inflammatory cytokine signaling, neutrophil activity, and markers of cell death. Consistent with disease complexity, heterogeneity in plasma profiling reveals distinct PASC subsets with striking divergence in these ongoing inflammatory processes, here termed plasma quiescent (plaq) and inflammatory (infl) PASC. In addition to elevated inflammatory blood proteomics, inflPASC patients display positive clinical tests of acute inflammation including C-reactive protein and fibrinogen, increased B cell activity with extrafollicular involvement coupled with elevated targeting of viral nucleocapsid protein and clinical autoreactivity. Further, the unique plasma signatures of PASC patients allowed for the creation of refined models with high sensitivity and specificity for the positive identification of inflPASC with a streamlined assessment of 12 blood markers. Additionally, refined ML modeling highlights the unexpected significance of several markers of potential diagnostic or therapeutic use for PASC in general, including the peptide hormone, epiregulin. In all, this work identifies clear biological signatures of PASC with potential diagnostic and therapeutic potential and establishes clear disease subtypes that are both easily identifiable and highly relevant to ongoing efforts in both therapeutic targeting and epidemiological investigation of this highly complex disease. Copyright The copyright holder for this preprint is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. All rights reserved. No reuse allowed without permission.

4.
J Med Virol ; 2022 Oct 13.
Article in English | MEDLINE | ID: covidwho-2075076

ABSTRACT

The ongoing coronavirus disease 2019 (COVID-19) pandemic has a significant global social and economic impact, and the emergence of new and more destructive mutant strains highlights the need for accurate virus detection. Here, 90 monoclonal antibodies (MAbs) that exclusively reacted with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nucleocapsid protein (NP) were generated. These MAbs did not cross-react with NPs of common human coronaviruses (HCoVs, i.e., 229E, OC43, HKU1, and NL63) and Middle East Respiratory Syndrome Coronavirus. Subsequently, overlapped peptides in individual fragments (N1-N4) of NP were synthesized. N1-3 (25-GSNQNGERSGARSKQ-39), N3-1 (217-AALALLLLDRLNQL-230), and N4-8 (393-TLLPAADLDDFSKQL-407) were identified as major epitopes using enzyme-linked immunoassay (ELISA) and recognized by 47, 1, and 18 MAbs, respectively. The 24 remaining MAbs exhibited no reactivity with all synthetic peptides. Among MAb-epitope pairs, only MAbs targeting epitope N1-3 displayed no cross-reaction with NPs of SARS-CoV-1 and other SARS-related CoVs. All Omicron variants contained a three-amino acid deletion (31ERS33) in the N1-3 region. Thus, MAbs targeting N1-3 failed to recognize these variants. Furthermore, a double-antibody sandwich ELISA for antigen detection was established using the optimal MAbs. Overall, a series of MAbs targeting SARS-CoV-2 NP was prepared, characterized with epitope mapping, and applied for the detection of SARS-CoV-2 antigens, and some novel B-cell epitopes of the viral NP were identified.

5.
Viruses ; 14(10)2022 10 16.
Article in English | MEDLINE | ID: covidwho-2071840

ABSTRACT

Host-virus protein interactions are critical for intracellular viral propagation. Understanding the interactions between cellular and viral proteins may help us develop new antiviral strategies. Porcine epidemic diarrhea virus (PEDV) is a highly contagious coronavirus that causes severe damage to the global swine industry. Here, we employed co-immunoprecipitation and liquid chromatography-mass spectrometry to characterize 426 unique PEDV nucleocapsid (N) protein-binding proteins in infected Vero cells. A protein-protein interaction network (PPI) was created, and gene ontology (GO) annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) database analyses revealed that the PEDV N-bound proteins belong to different cellular pathways, such as nucleic acid binding, ribonucleoprotein complex binding, RNA methyltransferase, and polymerase activities. Interactions of the PEDV N protein with 11 putative proteins: tripartite motif containing 21, DEAD-box RNA helicase 24, G3BP stress granule assembly factor 1, heat shock protein family A member 8, heat shock protein 90 alpha family class B member 1, YTH domain containing 1, nucleolin, Y-box binding protein 1, vimentin, heterogeneous nuclear ribonucleoprotein A2/B1, and karyopherin subunit alpha 1, were further confirmed by in vitro co-immunoprecipitation assay. In summary, studying an interaction network can facilitate the identification of antiviral therapeutic strategies and novel targets for PEDV infection.


Subject(s)
Coronavirus Infections , Nucleic Acids , Porcine epidemic diarrhea virus , Swine Diseases , Chlorocebus aethiops , Swine , Animals , Porcine epidemic diarrhea virus/genetics , Vimentin/metabolism , Vero Cells , Nucleocapsid/metabolism , Nucleocapsid Proteins/genetics , Viral Proteins/metabolism , Coronavirus Infections/metabolism , Antiviral Agents/metabolism , RNA/metabolism , Heat-Shock Proteins/metabolism , Methyltransferases/metabolism , Heterogeneous-Nuclear Ribonucleoproteins/metabolism , DEAD-box RNA Helicases/metabolism , Ribonucleoproteins/metabolism , Karyopherins/metabolism , Nucleic Acids/metabolism
6.
Dig Liver Dis ; 2022 Oct 17.
Article in English | MEDLINE | ID: covidwho-2068870

ABSTRACT

BACKGROUND AND AIMS: COVID-19 mRNA vaccines were approved to prevent severe forms of the disease, but their immunogenicity and safety in cirrhosis is poorly known. METHOD: In this prospective single-center study enrolling patients with cirrhosis undergoing COVID-19 vaccination (BNT162b2 and mRNA-1273), we assessed humoral and cellular responses vs healthy controls, the incidence of breakthrough infections and adverse events (AEs). Antibodies against spike- and nucleocapsid-protein (anti-S and anti-N) and Spike-specific T-cells responses were quantified at baseline, 21 days after the first and second doses and during follow-up. RESULTS: 182 cirrhotics (85% SARS-CoV-2-naïve) and 38 controls were enrolled. After 2 doses of vaccine, anti-S titres were significantly lower in cirrhotics vs controls [1,751 (0.4-25,000) U/mL vs 4,523 (259-25,000) U/mL, p=0.012] and in SARS-CoV-2-naïve vs previously infected cirrhotics [999 (0.4-17,329) U/mL vs 7,500 (12.5-25,000) U/mL, (p<0.001)]. T-cell responses in cirrhotics were similar to controls, although with different kinetics. In SARS-CoV-2-naïve cirrhotics, HCC, Child-Pugh B/C and BNT162b2 were independent predictors of low response. Neither unexpected nor severe AEs emerged. During follow-up, 2% turned SARS-CoV-2 positive, all asymptomatic. CONCLUSION: Humoral response to COVID-19 vaccines appeared suboptimal in patients with cirrhosis, particularly in SARS-CoV-2-naïve decompensated cirrhotics, although cellular response appeared preserved, and low breakthrough infections rate was registered.

7.
Anal Chim Acta ; 1234: 340522, 2022 Nov 22.
Article in English | MEDLINE | ID: covidwho-2068604

ABSTRACT

At the end of 2019, the novel coronavirus disease 2019 (COVID-19), a cluster of atypical pneumonia caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has been known as a highly contagious disease. Herein, we report the MXene/P-BiOCl/Ru(bpy)32+ heterojunction composite to construct an electrochemiluminescence (ECL) immunosensor for SARS-CoV-2 nucleocapsid protein (CoVNP) determination. Two-dimensional (2D) material ultrathin phosphorus-doped bismuth oxychloride (P-BiOCl) is exploited and first applied in ECL. 2D architectures MXene not only act as "soft substrate" to improve the properties of P-BiOCl, but also synergistically work with P-BiOCl. Owing to the inimitable set of bulk and interfacial properties, intrinsic high electrochemical conductivity, hydrophilicity and good biocompatible of 2D/2D MXene/P-BiOCl/Ru(bpy)32+, this as-exploited heterojunction composite is an efficient signal amplifier and co-reaction accelerator in the presence of tri-n-propylamine (TPA) as a coreactant. The proposed MXene/P-BiOCl/Ru(bpy)32+-TPA system exhibits a high and stable ECL signal and achieves ECL emission quenching for "signal on-off" recognition of CoVNP. Fascinatingly, the constructed ECL biosensor towards CoVNP allows a wide linear concentration range from 1 fg/mL to 10 ng/mL and a low limit of detection (LOD) of 0.49 fg/mL (S/N = 3). Furthermore, this presented strategy sheds light on designing a highly efficient ECL nanostructure through the combination of 2D MXene architectures with 2D semiconductor materials in the field of nanomedicine. This ECL biosensor can successfully detect CoVNP in human serum, which can promote the prosperity and development of diagnostic methods of SARS-CoV-2.


Subject(s)
Biosensing Techniques , COVID-19 , Humans , Biosensing Techniques/methods , Bismuth , COVID-19/diagnosis , Electrochemical Techniques/methods , Immunoassay/methods , Luminescent Measurements/methods , Nucleocapsid Proteins , SARS-CoV-2
8.
Clinical and Experimental Rheumatology ; 40(10):83, 2022.
Article in English | EMBASE | ID: covidwho-2067780

ABSTRACT

Sjogren's syndrome (SS) is a chronic and systemic autoimmune disease characterized by lymphocytic infiltration and the development of dry eyes and dry mouth due to the secretory dysfunction of the lacrimal and salivary glands. In recent years, infectious pathogens have been proved to be associated with SS, including Cytomegalovirus, Coxsackie, EBV, and lymphotropic virus-1 (HTLV-1). Studies suggest that infections caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) may trigger an autoimmune response, as evidenced by increased autoantibodies in patients diagnosed with Coronavirus disease 2019 (COVID-19). To investigate the relationship between SARS-CoV-2 and SS, the study was performed by infecting humanized ACE2 mice with SARS-CoV-2. Mice infected with the virus showed a decreased saliva flow rate, elevated antinuclear antibodies (ANAs) and anti-SSB/La, and lymphocyte infiltration in the lacrimal and salivary glands. We detected the viral nucleocapsid protein in mice exocrine glands with significant apoptotic bodies by the acinar cells. Confirmed with clinical data, we also observed the elevation of SS-specific autoantibodies (ANA, anti-SSB/Ro52, and anti-SSA/La) and specific ANA patterns in sera from COVID-19 patients. One unique aspect of SS is the high degree of sexual dimorphism, with women being affected 10-20 times more than men. To determine whether COVID-19 patients exhibited an element of sexual dimorphism in the autoantibody response, we grouped the sera by sex. We found the male patients showed elevated anti-SSA/Ro52 compared to female patients (p=0.0029), and female patients had more diverse ANA patterns. Lastly, monoclonal antibodies isolated from recovered patients using singlecell antibody nanowells technology were shown to recognize the nuclear antigens. Overall, by observing SS-like phenotypes in mouse models and patients, our study confirms a direct pathogenic role of SARS-CoV-2 in SS.

9.
Vaccines (Basel) ; 10(10)2022 Oct 01.
Article in English | MEDLINE | ID: covidwho-2066613

ABSTRACT

The current report presents the case of a 76-year-old man with Parkinson's disease (PD) who died three weeks after receiving his third COVID-19 vaccination. The patient was first vaccinated in May 2021 with the ChAdOx1 nCov-19 vector vaccine, followed by two doses of the BNT162b2 mRNA vaccine in July and December 2021. The family of the deceased requested an autopsy due to ambiguous clinical signs before death. PD was confirmed by post-mortem examinations. Furthermore, signs of aspiration pneumonia and systemic arteriosclerosis were evident. However, histopathological analyses of the brain uncovered previously unsuspected findings, including acute vasculitis (predominantly lymphocytic) as well as multifocal necrotizing encephalitis of unknown etiology with pronounced inflammation including glial and lymphocytic reaction. In the heart, signs of chronic cardiomyopathy as well as mild acute lympho-histiocytic myocarditis and vasculitis were present. Although there was no history of COVID-19 for this patient, immunohistochemistry for SARS-CoV-2 antigens (spike and nucleocapsid proteins) was performed. Surprisingly, only spike protein but no nucleocapsid protein could be detected within the foci of inflammation in both the brain and the heart, particularly in the endothelial cells of small blood vessels. Since no nucleocapsid protein could be detected, the presence of spike protein must be ascribed to vaccination rather than to viral infection. The findings corroborate previous reports of encephalitis and myocarditis caused by gene-based COVID-19 vaccines.

10.
Toxins (Basel) ; 14(10)2022 10 04.
Article in English | MEDLINE | ID: covidwho-2066493

ABSTRACT

With the outbreak and spread of COVID-19, a deep investigation of SARS-CoV-2 is urgent. Direct usage of this virus for scientific research could provide reliable results and authenticity. However, it is strictly constrained and unrealistic due to its high pathogenicity and infectiousness. Considering its biosafety, different systems and technologies have been employed in immunology and biomedical studies. In this study, phage display technology was used to construct a nonpathogenic model for COVID-19 research. The nucleocapsid protein of SARS-CoV-2 was fused with the M13 phage capsid p3 protein and expressed on the M13 phages. After validation of its successful expression, its potential as the standard for qPCR quantification and affinity with antibodies were confirmed, which may show the possibility of using this nonpathogenic bacteriophage to replace the pathogenic virus in scientific research concerning SARS-CoV-2. In addition, the model was used to develop a system for the classification and identification of different samples using ATR-FTIR, which may provide an idea for the development and evaluation of virus monitoring equipment in the future.


Subject(s)
COVID-19 , Viruses , Humans , SARS-CoV-2/genetics , Cell Surface Display Techniques , Bacteriophage M13/genetics , Bacteriophage M13/metabolism , Capsid Proteins/genetics , Capsid Proteins/metabolism , Nucleocapsid Proteins/genetics , Nucleocapsid Proteins/metabolism
11.
Life (Basel) ; 12(10)2022 Sep 30.
Article in English | MEDLINE | ID: covidwho-2066223

ABSTRACT

Background: Case reports are available showing that patients develop symptoms of acute arthritis during or after recovery from SARS-CoV-2 infection. Since the interrelation is still unknown, our aim was to study the impact of the SARS-CoV-2 nucleocapsid protein (NP) on human fibroblast-like synoviocytes and human endothelial cells (hEC) in terms of complement and cytokine regulation. Methods: Non-arthritic (K4IM) synoviocyte, arthritic (HSE) synoviocyte cell lines and primary hEC were stimulated with recombinant NP and/or TNFα. Analyses of cell viability, proliferation, gene and protein expression of cytokines and complement factors were performed. Results: NP suppressed significantly the vitality of hEC and proliferation of HSE. NP alone did not induce any significant changes in the examined gene expressions. However, NP combined with TNFα induced significantly higher TNFα in HSE and K4IM as well as higher IL-6 and CD55 gene expression in HSE and suppressed C3aR1 gene expression in hEC. HSE proliferated twice as fast as K4IM, but showed significantly lesser gene expressions of CD46, CD55, CD59 and TNFα with significantly higher IL-6 gene expression. CD35 gene expression was undetectable in K4IM, HSE and hEC. Conclusions: NP might contribute in combination with other inflammatory factors to complement regulation in arthritis.

12.
Chest ; 162(4):A507, 2022.
Article in English | EMBASE | ID: covidwho-2060615

ABSTRACT

SESSION TITLE: COVID-19 Case Report Posters 2 SESSION TYPE: Case Report Posters PRESENTED ON: 10/19/2022 12:45 pm - 01:45 pm INTRODUCTION: The SARS-CoV-2 pandemic spawned the use and study of novel therapeutics, re-purposed drugs, and interventions– all with limited success. Seraph® 100 Microbind Affinity Blood Filter® [Seraph®] is an investigational device that was given Emergency Use Authorization (EUA) by the Food and Drug Administration in 2019 to treat severe coronavirus disease. Higher viremia is correlated with higher mortality. Seraph® uses extracorporeal filtration to aid the innate immune system by reducing viral load and mitigating downstream effects of inflammation. CASE PRESENTATION: A forty-eight year old gentleman presented to the emergency room with coughs and fever of 101° F. He tested positive via polymerase chain reaction [PCR] testing for SARS-CoV-19 pneumonia. A computed-tomography angiogram [CTA] of the chest was negative for pulmonary embolism, but demonstrated significant bilateral ground-glass opacities consistent with viral pneumonia. Vitals were notable for an oxygen saturation of 69% on room air that improved to 96% on 6 liters/minute [L/min] of supplemental oxygen via nasal cannula. He was initiated on both dexamethasone and remdesivir. Within hours, the patient's oxygen requirements escalated to 15 L/min via non-rebreather to high flow humidified nasal cannula with a flow rate of 40 L/min and 60% FiO2. On day three, he was transferred to the ICU for treatment with Seraph®. After one treatment, the patient was weaned from high flow humidified nasal cannula to room air. On day five, after a second treatment, he transferred to the floor. He was discharged on day six, on room air, having completed his course of remdesivir, with an additional 5 days of oral steroids. DISCUSSION: Preliminary data regarding Seraph® remain limited with only select eligible patients undergoing therapy. Cases like our patient demonstrate dramatic improvements with even one or two treatments which correlate well with data that show up to 99% reduction in the bloodstream of targeted pathogens per pass. While database collection data have revealed trends towards improved outcomes, further investigation into populations most likely to benefit from treatment is needed. Timing, immunocompromised status and other comorbidities that raise or lower the chances of successful hemofiltration need to be considered. CONCLUSIONS: Studies in the SARS-CoV-2 pandemic augment ongoing research as filtration devices are used to target bacterial infections, cytokines and inflammatory markers. Since the invention of antibiotics, multi-drug resistant organisms have increased in prevalence. Novel interventions such as Seraph® warrant investigation to prevent infectious diseases from becoming unmanageable threats. Reference #1: Kielstein JT, Borchina DN, Fühner T, Hwang S, Mattoon D, Ball AJ. Hemofiltration with the Seraph® 100 Microbind® Affinity filter decreases SARS-CoV-2 nucleocapsid protein in critically ill COVID-19 patients. Crit Care. 2021;25(1):190. Published 2021 Jun 1. doi:10.1186/s13054-021-03597-3 Reference #2: Pape A, Kielstein JT, Krüger T, Fühner T, Brunkhorst R. Treatment of a Critically Ill COVID-19 Patient with the Seraph 100 Microbind Affinity Filter. TH Open. 2021;5(2):e134-e138. Published 2021 Apr 14. doi:10.1055/s-0041-1727121 Reference #3: Schmidt JJ, Borchina DN, van T Klooster M, et al. Interim-analysis of the COSA (COVID-19 patients treated with the Seraph® 100 Microbind® Affinity filter) registry [published online ahead of print, 2021 Dec 7]. Nephrol Dial Transplant. 2021;gfab347. doi:10.1093/ndt/gfab347 DISCLOSURES: No relevant relationships by Aneesa Afroze No relevant relationships by Lydia Meece No relevant relationships by Angela Park

13.
Embase; 2020.
Preprint in English | EMBASE | ID: ppcovidwho-344370

ABSTRACT

Effective public-health measures and vaccination campaigns against SARS-CoV-2 require granular knowledge of population-level immune responses. We developed a Tripartite Automated Blood Immunoassay (TRABI) to assess the IgG response against the ectodomain and the receptor-binding domain of the spike protein as well as the nucleocapsid protein of SARS-CoV-2. We used TRABI for continuous seromonitoring of hospital patients and healthy blood donors (n=72'222) in the canton of Zurich from December 2019 to December 2020 (pre-vaccine period). Seroprevalence peaked in May 2020 and rose again in November 2020 in both cohorts. Validations of results included antibody diffusional sizing and Western Blotting. Using an extended Susceptible-Exposed-Infectious-Removed model, we found that antibodies waned with a half-life of 75 days, whereas the cumulative incidence rose from 2.3% in June 2020 to 12.2% in mid-December 2020 in the population of the canton of Zurich. A follow-up health survey indicated that about 10% of patients infected with wildtype SARS-CoV-2 sustained some symptoms at least twelve months post COVID-19 and up to the timepoint of survey participation. Crucially, we found no evidence for a difference in long-term complications between those whose infection was symptomatic and those with asymptomatic acute infection. The cohort of asymptomatic SARS-CoV-2-infected subjects represents a resource for the study of chronic and possibly unexpected sequelae. Copyright The copyright holder for this preprint is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-NC 4.0 International license.

14.
Anal Chim Acta ; 1233: 340486, 2022 Nov 15.
Article in English | MEDLINE | ID: covidwho-2060277

ABSTRACT

The SARS-CoV-2 pandemic has posed a huge challenge to rapid and accurate diagnosis of SARS-CoV-2 in the early stage of infection. In this work, we developed a novel magnetic/fluorescent dual-modal lateral flow immunoassay (LFIA) based on multifunctional nanobeads for rapid and accurate determination of SARS-CoV-2 nucleocapsid protein (NP). The multifunctional nanobeads were fabricated by using polyethyleneimine (PEI) as a mediate shell to combine superparamagnetic Fe3O4 core with dual quantum dot shells (MagDQD). The core-shell structure of MagDQD label with high loading density of quantum dots (QDs) and superior magnetic content realized LFIA with dual quantitative analysis modal from the assemblies of individual single nanoparticles. The LFIA integrated the advantages of magnetic signal and fluorescent signal, resulting excellent accuracy for quantitative analysis and high elasticity of the overall detection. In addition, magnetic signal and fluorescent signal both had high sensitivity with the limit of detection (LOD) as 0.235 ng mL-1 and 0.012 ng mL-1, respectively. The recovery rates of the methods in simulated saliva samples were 91.36%-103.60% (magnetic signal) and 94.39%-104.38% (fluorescent signal). The results indicate the method has a considerable potential to be an effective tool for diagnose SARS-CoV-2 in the early stage of infection.


Subject(s)
COVID-19 , SARS-CoV-2 , Humans , Polyethyleneimine , COVID-19/diagnosis , Immunoassay/methods , Magnetic Phenomena
15.
mSphere ; 7(5): e0025722, 2022 10 26.
Article in English | MEDLINE | ID: covidwho-2053133

ABSTRACT

Accurate, highly specific immunoassays for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are needed to evaluate seroprevalence. This study investigated the concordance of results across four immunoassays targeting different antigens for sera collected at the beginning of the SARS-CoV-2 pandemic in the United States. Specimens from All of Us participants contributed between January and March 2020 were tested using the Abbott Architect SARS-CoV-2 IgG (immunoglobulin G) assay (Abbott) and the EuroImmun SARS-CoV-2 enzyme-linked immunosorbent assay (ELISA) (EI). Participants with discordant results, participants with concordant positive results, and a subset of concordant negative results by Abbott and EI were also tested using the Roche Elecsys anti-SARS-CoV-2 (IgG) test (Roche) and the Ortho-Clinical Diagnostics Vitros anti-SARS-CoV-2 IgG test (Ortho). The agreement and 95% confidence intervals were estimated for paired assay combinations. SARS-CoV-2 antibody concentrations were quantified for specimens with at least two positive results across four immunoassays. Among the 24,079 participants, the percent agreement for the Abbott and EI assays was 98.8% (95% confidence interval, 98.7%, 99%). Of the 490 participants who were also tested by Ortho and Roche, the probability-weighted percentage of agreement (95% confidence interval) between Ortho and Roche was 98.4% (97.9%, 98.9%), that between EI and Ortho was 98.5% (92.9%, 99.9%), that between Abbott and Roche was 98.9% (90.3%, 100.0%), that between EI and Roche was 98.9% (98.6%, 100.0%), and that between Abbott and Ortho was 98.4% (91.2%, 100.0%). Among the 32 participants who were positive by at least 2 immunoassays, 21 had quantifiable anti-SARS-CoV-2 antibody concentrations by research assays. The results across immunoassays revealed concordance during a period of low prevalence. However, the frequency of false positivity during a period of low prevalence supports the use of two sequentially performed tests for unvaccinated individuals who are seropositive by the first test. IMPORTANCE What is the agreement of commercial SARS-CoV-2 immunoglobulin G (IgG) assays during a time of low coronavirus disease 2019 (COVID-19) prevalence and no vaccine availability? Serological tests produced concordant results in a time of low SARS-CoV-2 prevalence and no vaccine availability, driven largely by the proportion of samples that were negative by two immunoassays. The CDC recommends two sequential tests for positivity for future pandemic preparedness. In a subset analysis, quantified antinucleocapsid and antispike SARS-CoV-2 IgG antibodies do not suggest the need to specify the antigen targets of the sequential assays in the CDC's recommendation because false positivity varied as much between assays targeting the same antigen as it did between assays targeting different antigens.


Subject(s)
COVID-19 , Population Health , Humans , SARS-CoV-2 , COVID-19/diagnosis , COVID-19/epidemiology , Prevalence , Seroepidemiologic Studies , Sensitivity and Specificity , Antibodies, Viral , Immunoglobulin G
16.
ACS Appl Mater Interfaces ; 14(40): 45189-45200, 2022 Oct 12.
Article in English | MEDLINE | ID: covidwho-2050254

ABSTRACT

The worldwide spread of coronavirus disease 2019 (COVID-19) highlights the need for rapid, simple, and accurate tests to detect various variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The antigen test, based on the lateral flow immunoassay (LFI), is a suitable "first line of defense" test that enables early identification and timely isolation of patients to minimize viral transmission among communities. However, it is generally less accurate than nucleic acid testing, and its sensitivity needs improvement. Here, a novel rapid detection method is designed to sensitively detect SARS-CoV-2 using isolated gold nanoparticle (AuNP)-assembled SiO2 core-satellite nanoparticles (SiO2@Au CSNPs). Well-grown AuNP satellites in the synthesis of SiO2@Au CSNPs significantly enhanced their light absorption, increased the detection sensitivity, and lowered the detection limit by 2 orders of magnitude relative to conventional gold colloids. The proposed system enabled highly sensitive detection of the SARS-CoV-2 nucleocapsid protein with a detection limit of 0.24 pg mL-1 within 20 min. This is the first study to develop a highly sensitive antigen test using the absorption-modulated SiO2@Au CSNPs. Our findings demonstrate the capacity of this platform to serve as an effective sensing strategy for managing pandemic conditions and preventing the spread of viral infections.


Subject(s)
Biosensing Techniques , COVID-19 , Metal Nanoparticles , Nucleic Acids , COVID-19/diagnosis , Colloids , Gold , Humans , Immunoassay/methods , SARS-CoV-2 , Sensitivity and Specificity , Silicon Dioxide
17.
Biosensors (Basel) ; 12(10)2022 Sep 23.
Article in English | MEDLINE | ID: covidwho-2043580

ABSTRACT

The global pandemic of COVID-19 has created an unrivalled need for sensitive and rapid point-of-care testing (POCT) methods for the detection of infectious viruses. For the novel coronavirus SARS-CoV-2, the nucleocapsid protein (N-protein) is one of the most abundant structural proteins of the virus and it serves as a useful diagnostic marker for detection. Herein, we report a fiber optic particle plasmon resonance (FOPPR) biosensor which employed a single-stranded DNA (ssDNA) aptamer as the recognition element to detect the SARS-CoV-2 N-protein in 15 min with a limit of detection (LOD) of 2.8 nM, meeting the acceptable LOD of 106 copies/mL set by the WHO target product profile. The sensor chip is a microfluidic chip based on the balance between the gravitational potential and the capillary force to control fluid loading, thus enabling the power-free auto-flowing function. It also has a risk-free self-contained design to avoid the risk of the virus leaking into the environment. These findings demonstrate the potential for designing a low-cost and robust POCT device towards rapid antigen detection for early screening of SARS-CoV-2 and its related mutants.


Subject(s)
Biosensing Techniques , COVID-19 , Humans , SARS-CoV-2 , DNA, Single-Stranded , Microfluidics , COVID-19/diagnosis , Nucleocapsid Proteins/genetics
18.
Vaccines (Basel) ; 10(9)2022 Sep 15.
Article in English | MEDLINE | ID: covidwho-2044009

ABSTRACT

BACKGROUND: We conducted a longitudinal study to estimate immunity produced in response to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection among university students over seven months. METHODS: All participants were attending a public university and resided in Pitt County, North Carolina. University students enrolled weekly for 10 weeks between 26 August 2020 and 28 October 2020, resulting in 136 young adults completing at least one study visit by 17 November 2020. Enrolled students completed an online survey and nasal swab collection at two-week intervals and monthly blood collection between 26 August 2020 and 31 March 2021. RESULTS: Amongst 695 serum samples tested during follow-up, the prevalence of a positive result for anti-nucleocapsid antibodies (N-IgG) was 9.78%. In 22 students with more than one positive N-IgG serum sample, 68.1% of the group lost persistence of N-IgG below the positive threshold over 140 days. Anti-spike IgG antibodies were significantly higher among 11 vaccinated compared to 10 unvaccinated. CONCLUSIONS: In healthy young adults, N-IgG wanes below the detectable threshold within five months. S-IgG titer remained consistently elevated months after infection, and significantly increased after vaccination.

19.
Biosens Bioelectron ; 218: 114737, 2022 Dec 15.
Article in English | MEDLINE | ID: covidwho-2041594

ABSTRACT

Lateral flow immunoassays (LFIA) have a plethora of applications in health, environmental and food sectors for low-cost, simple, and rapid point-of-need testing. Typically, the user only needs to add the sample without any other intervention from sample application to results. A compelling challenge, and a constant pursuit in LFIA is to improve the assay sensitivity without compromising the simplicity and practicality. We report that the addition of water-soluble macromolecular crowding agents leads to an enhancement of the sensitivity, which is attributed to the fact that the exposure of antibodies and micro/nanoparticle conjugates to macromolecularly crowded environment, while migrating through the confining pores of the strip-pads by capillary forces, promotes the interactions that are responsible for analyte recognition and signal generation. The effect was shown by using two of the most widely established LFIA tests worldwide, that is, detection of nucleocapsid protein from SARS-CoV-2 associated with COVID-19 and detection of Strep-A antigen from Streptococcus pyogenes associated with pharyngitis. For immediate demonstration of the sensitivity enhancement, we worked directly on commercially available devices already optimized in terms of reagents and conditions. Of the crowders used, ficoll, Mr 400000, and ficoll, Mr 70000, gave a 5-10-fold improvement of the signal without affecting the background. Because the addition of macromolecular crowding agents is complementary to other strategies of sensitivity enhancement, such as the design of novel labels and the introduction of signal amplification, we anticipate that the proposed modulation will be extended to numerous analytes with a variety of reporters and LFIA configurations.


Subject(s)
Biosensing Techniques , COVID-19 , Humans , SARS-CoV-2 , COVID-19/diagnosis , Ficoll , Immunoassay/methods , Nucleocapsid Proteins , Water , Sensitivity and Specificity
20.
Swiss Medical Weekly ; 152:31S, 2022.
Article in English | EMBASE | ID: covidwho-2040896

ABSTRACT

Background: Rapid antigen tests (RATs) are widely used for pointof- care or self-testing to identify SARS-CoV-2 (SCoV2)-positive cases. Aim: To assess the performance of RATs with currently circulating Omicron variants that may impair detection. Methods: We prospectively evaluated the Roche-SARS-CoV-2- Antigen and Acon-FlowFlex-SARS-CoV-2-Antigen in 150 consecutively collected nasopharyngeal patient swabs (50 SCoV2 RNA undetectable;100 SCoV2 Omicron BA.1). Omicron BA.1 results were compared to 92 Ct-matched early pandemic SCoV2 variants (B.1.160 and B.1.177), to 100 Omicron BA.2 positive and to 100 Omicron BA.5 positive samples. Results: For Omicron BA.1, Roche-SARS-CoV-2-Antigen detected 87% of samples having Ct-values <29 reflecting 3.6% lower rates compared to B.1.160 and B.1.177. Acon-FlowFlex-SARS-CoV-2- Antigen was less affected and detected 90% of Omicron BA.1 with Ct-values <29. Omicron BA.2 and BA.5 detection rates were significantly reduced by 20% and 10%, respectively, for the Roche-SARSCoV- 2-Antigen in samples with Ct-values <29, but remained similar for Acon-FlowFlex-SARS-CoV-2-Antigen. RATs need to be continuously evaluated as new SCoV2-variants emerge. Conclusion: This study provides evidence that variation within the nucleocapsid protein as seen in recently emerged and now globally spreading Omicron BA.2 and Omicron BA.5 variants significantly impairs detection rates of widely used antigen tests. Consequently, antigen tests need to be re-evaluated when new pandemic SCoV2 variants emerge and start to predominate globally.

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