scholarly journals Development of a Unique Rapid Test to Detect Anti-bodies Directed Against an Extended RBD of SARS-CoV-2 Spike Protein

2021 ◽  
Vol 75 (5) ◽  
pp. 446-452
Author(s):  
Larissa Brosi ◽  
Eric Kübler ◽  
Anna Weston ◽  
Patrick Romann ◽  
Sherin Panikulam ◽  
...  
Keyword(s):  
Immune Response ◽  
Receptor Binding ◽  
Recombinant Expression ◽  
Rapid Test ◽  
High Sensitivity ◽  
Spike Protein ◽  
Receptor Binding Domain ◽  
Serological Testing ◽  
Humoral Immune ◽  
Previous Infection

Serological testing for antibodies directed against SARS-CoV-2 in patients may serve as a diagnostic tool to verify a previous infection and as surrogate for an elicited humoral immune response, ideally conferring immunity after infection or vaccination. Here, we present the recombinant expression of an extended receptor binding domain (RBD) of the SARS-CoV-2 Spike protein used as capture antigen in a unique rapid immunoassay to detect the presence of RBD binding antibodies with high sensitivity and specificity. As currently available vaccines focus on the Spike RBD as target, the developed test can also be used to monitor a successful immune response after vaccination with an RBD based vaccine.

scholarly journals SARS-CoV-2 Infection Is Asymptomatic in Nearly Half of Adults with Robust Anti-Spike Protein Receptor-Binding Domain Antibody Response

Vaccines ◽  
2021 ◽  
Vol 9 (3) ◽  
pp. 207
Author(s):  
Ourania E. Tsitsilonis ◽  
Dimitrios Paraskevis ◽  
Evi Lianidou ◽  
Evangelos Terpos ◽  
Athanasios Akalestos ◽  
...  
Keyword(s):  
Receptor Binding ◽  
Clinical Symptoms ◽  
Binding Domain ◽  
Spike Protein ◽  
Receptor Binding Domain ◽  
Humoral Immune ◽  
Protein Receptor ◽  
Previous Infection ◽  
And Gender ◽  
Asymptomatic Individuals

Between June and November 2020, we assessed plasma antibodies against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nucleocapsid protein in 4996 participants (aged 18–82 years, 34.5% men) from the National and Kapodistrian University of Athens. The weighted overall prevalence was 1.6% and monthly prevalence correlated with viral RNA-confirmed SARS-CoV-2 infections in Greece, in the same period. Notably, 49% of seropositive cases reported no history of SARS-CoV-2 infection-related clinical symptoms and 33% were unsuspected of their previous infection. Additionally, levels of anti-SARS-CoV-2 antibodies against the spike-protein receptor-binding domain were similar between symptomatic and asymptomatic individuals, irrespective of age and gender. Using Food and Drug Administration Emergency Use Authorization-approved assays, these results support the need for such studies on pandemic evaluation and highlight the development of robust humoral immune responses even among asymptomatic individuals. The high percentage of unsuspected/asymptomatic active cases, which may contribute to community transmission for more days than that of cases who are aware and self-isolate, underscores the necessity of measures across the population for the efficient control of the pandemic.

scholarly journals Engineered ACE2 counteracts vaccine-evading SARS-CoV-2 Omicron variant

2021 ◽  
Author(s):  
Nariko Ikemura ◽  
Shunta Taminishi ◽  
Tohru Inaba ◽  
Takao Arimori ◽  
Daisuke Motooka ◽  
...  
Keyword(s):  
Receptor Binding ◽  
Mutational Analysis ◽  
High Sensitivity ◽  
Binding Domain ◽  
Spike Protein ◽  
The Novel ◽  
Receptor Binding Domain ◽  
Therapeutic Drugs ◽  
Viral Mutation ◽  
Therapeutic Value

The novel SARS-CoV-2 variant, Omicron (B.1.1.529) contains an unusually high number of mutations (>30) in the spike protein, raising concerns of escape from vaccines, convalescent sera and therapeutic drugs. Here we analyze the alteration of neutralizing titer with Omicron pseudovirus. Sera of 3 months after double BNT162b2 vaccination exhibit approximately 18-fold lower neutralization titers against Omicron. Convalescent sera from Alpha and Delta patients allow similar levels of breakthrough by Omicron. However, some Delta patients have relatively preserved neutralization efficacy, comparable to 3-month double BNT162b2 vaccination. Domain-wise analysis using chimeric spike revealed that this efficient evasion was, at least in part, caused by multiple mutations in the N-terminal domain. Omicron escapes the therapeutic cocktail of imdevimab and casirivimab, whereas sotrovimab, which targets a conserved region to avoid viral mutation, remains effective against Omicron. The ACE2 decoy is another virus-neutralizing drug modality that is free, at least in theory, from mutational escape. Deep mutational analysis demonstrated that, indeed, the engineered ACE2 overcomes every single-residue mutation in the receptor-binding domain, similar to immunized sera. Like previous SARS-CoV-2 variants, Omicron and some other sarbecoviruses showed high sensitivity against engineered ACE2, confirming the therapeutic value against diverse variants, including those that are yet to emerge.

scholarly journals Recombinant Mycobacterium paragordonae Expressing SARS-CoV-2 Receptor-Binding Domain as a Vaccine Candidate Against SARS-CoV-2 Infections

2021 ◽  
Vol 12 ◽  
Author(s):  
Byoung-Jun Kim ◽  
Hyein Jeong ◽  
Hyejun Seo ◽  
Mi-Hyun Lee ◽  
Hyun Mu Shin ◽  
...  
Keyword(s):  
Immune Response ◽  
Single Dose ◽  
Immune Responses ◽  
Receptor Binding ◽  
Humoral Immune Response ◽  
Vaccine Candidate ◽  
Binding Domain ◽  
Receptor Binding Domain ◽  
Live Virus ◽  
Humoral Immune

At present, concerns that the recent global emergence of SARS-CoV-2 variants could compromise the current vaccines have been raised, highlighting the urgent demand for new vaccines capable of eliciting T cell-mediated immune responses, as well as B cell-mediated neutralizing antibody production. In this study, we developed a novel recombinant Mycobacterium paragordonae expressing the SARS-CoV-2 receptor-binding domain (RBD) (rMpg-RBD-7) that is capable of eliciting RBD-specific immune responses in vaccinated mice. The potential use of rMpg-RBD-7 as a vaccine for SARS-CoV-2 infections was evaluated in in vivo using mouse models of two different modules, one for single-dose vaccination and the other for two-dose vaccination. In a single-dose vaccination model, we found that rMpg-RBD-7 versus a heat-killed strain could exert an enhanced cell-mediated immune (CMI) response, as well as a humoral immune response capable of neutralizing the RBD and ACE2 interaction. In a two-dose vaccination model, rMpg-RBD-7 in a two-dose vaccination could also exert a stronger CMI and humoral immune response to neutralize SARS-CoV-2 infections in pseudoviral or live virus infection systems, compared to single dose vaccinations of rMpg-RBD or two-dose RBD protein immunization. In conclusion, our data showed that rMpg-RBD-7 can lead to an enhanced CMI response and humoral immune responses in mice vaccinated with both single- or two-dose vaccination, highlighting its feasibility as a novel vaccine candidate for SARS-CoV-2. To the best of our knowledge, this study is the first in which mycobacteria is used as a delivery system for a SARS-CoV-2 vaccine.

scholarly journals High seroprevalence for SARS-CoV-2 among household members of essential workers detected using a dried blood spot assay

2020 ◽  
Author(s):  
Thomas W. McDade ◽  
Elizabeth M. McNally ◽  
Aaron S. Zelikovich ◽  
Richard D’Aquila ◽  
Brian Mustanski ◽  
...  
Keyword(s):  
Receptor Binding ◽  
Venous Blood ◽  
Binding Domain ◽  
Spike Protein ◽  
Dried Blood Spot ◽  
Receptor Binding Domain ◽  
Serological Testing ◽  
Igg Antibodies ◽  
Household Members ◽  
Blood Spot

AbstractObjectiveSerological testing is needed to investigate the extent of transmission of SARS-CoV-2 from front-line essential workers to their household members. However, the requirement for serum/plasma limits serological testing to clinical settings where it is feasible to collect and process venous blood. To address this problem we developed a serological test for SARS-CoV-2 IgG antibodies that requires only a single drop of finger stick capillary whole blood, collected in the home and dried on filter paper (dried blood spot, DBS).MethodsAn ELISA to the receptor binding domain of the SARS-CoV-2 spike protein was optimized to quantify IgG antibodies in DBS. Samples were self-collected from a community sample of 232 participants enriched with health care workers, including 30 known COVID-19 cases and their household members.ResultsAmong 30 individuals sharing a household with a virus-confirmed case of COVID-19, 80% were seropositive. Of 202 community individuals without prior confirmed acute COVID-19 diagnoses, 36% were seropositive. Of documented convalescent COVID-19 cases from the community, 29 of 30 (97%) were seropositive for IgG antibodies to the receptor binding domain.ConclusionDBS ELISA provides a minimally-invasive alternative to venous blood collection. Early analysis suggests a high rate of transmission among household members. High rates of seroconversion were also noted following recovery from infection. Serological testing for SARSCoV-2 IgG antibodies in DBS samples can facilitate seroprevalence assessment in community settings to address epidemiological questions, monitor duration of antibody responses, and assess if antibodies against the spike protein correlate with protection from reinfection.

scholarly journals Comparative Immunomodulatory Evaluation of the Receptor Binding Domain of the SARS-CoV-2 Spike Protein; a Potential Vaccine Candidate Which Imparts Potent Humoral and Th1 Type Immune Response in a Mouse Model

2021 ◽  
Vol 12 ◽  
Author(s):  
Tripti Shrivastava ◽  
Balwant Singh ◽  
Zaigham Abbas Rizvi ◽  
Rohit Verma ◽  
Sandeep Goswami ◽  
...  
Keyword(s):  
Immune Response ◽  
Receptor Binding ◽  
Neutralizing Antibodies ◽  
Vaccine Development ◽  
Vaccine Candidate ◽  
Unmet Need ◽  
Binding Domain ◽  
Spike Protein ◽  
Cell Immune Response ◽  
Receptor Binding Domain

The newly emerged novel coronavirus, SARS-CoV-2, the causative agent of COVID-19 has proven to be a threat to the human race globally, thus, vaccine development against SARS-CoV-2 is an unmet need driving mass vaccination efforts. The receptor binding domain of the spike protein of this coronavirus has multiple neutralizing epitopes and is associated with viral entry. Here we have designed and characterized the SARS-CoV-2 spike protein fragment 330-526 as receptor binding domain 330-526 (RBD330-526) with two native glycosylation sites (N331 and N343); as a potential subunit vaccine candidate. We initially characterized RBD330-526 biochemically and investigated its thermal stability, humoral and T cell immune response of various RBD protein formulations (with or without adjuvant) to evaluate the inherent immunogenicity and immunomodulatory effect. Our result showed that the purified RBD immunogen is stable up to 72 h, without any apparent loss in affinity or specificity of interaction with the ACE2 receptor. Upon immunization in mice, RBD generates a high titer humoral response, elevated IFN-γ producing CD4+ cells, cytotoxic T cells, and robust neutralizing antibodies against live SARS-CoV-2 virus. Our results collectively support the potential of RBD330-526 as a promising vaccine candidate against SARS-CoV-2.

scholarly journals A compromised specific humoral immune response against the SARS-CoV-2 receptor-binding domain is related to viral persistence and periodic shedding in the gastrointestinal tract

2020 ◽  
Vol 17 (11) ◽  
pp. 1119-1125 ◽  
Author(s):  
Fengyu Hu ◽  
Fengjuan Chen ◽  
Zhihua Ou ◽  
Qinghong Fan ◽  
Xinghua Tan ◽  
...  
Keyword(s):  
Immune Response ◽  
Gastrointestinal Tract ◽  
Receptor Binding ◽  
Humoral Immune Response ◽  
Clinical Manifestations ◽  
Viral Persistence ◽  
Binding Domain ◽  
Health Concern ◽  
Receptor Binding Domain ◽  
Humoral Immune

Abstract Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been redetected after discharge in some coronavirus disease 2019 (COVID-19) patients. The reason for the recurrent positivity of the test and the potential public health concern due to this occurrence are still unknown. Here, we analyzed the viral data and clinical manifestations of 289 domestic Chinese COVID-19 patients and found that 21 individuals (7.3%) were readmitted for hospitalization after detection of SARS-CoV-2 after discharge. First, we experimentally confirmed that the virus was involved in the initial infection and was not a secondary infection. In positive retests, the virus was usually found in anal samples (15 of 21, 71.4%). Through analysis of the intracellular viral subgenomic messenger RNA (sgmRNA), we verified that positive retest patients had active viral replication in their gastrointestinal tracts (3 of 16 patients, 18.7%) but not in their respiratory tracts. Then, we found that viral persistence was not associated with high viral titers, delayed viral clearance, old age, or more severe clinical symptoms during the first hospitalization. In contrast, viral rebound was associated with significantly lower levels of and slower generation of viral receptor-binding domain (RBD)-specific IgA and IgG antibodies. Our study demonstrated that the positive retest patients failed to create a robust protective humoral immune response, which might result in SARS-CoV-2 persistence in the gastrointestinal tract and possibly in active viral shedding. Further exploration of the mechanism underlying the rebound in SARS-CoV-2 in this population will be crucial for preventing virus spread and developing effective vaccines.

scholarly journals Humoral and cell-mediated response in colostrum after exposure to severe acute respiratory syndrome coronavirus 2

2021 ◽  
Author(s):  
Vignesh Narayanaswamy ◽  
Brian Pentecost ◽  
Dominique Alfandari ◽  
Emily Chin ◽  
Kathleen Minor ◽  
...  
Keyword(s):  
Immune Response ◽  
Severe Acute Respiratory Syndrome ◽  
Inflammatory Markers ◽  
Large Scale ◽  
Spike Protein ◽  
Receptor Binding Domain ◽  
New Mothers ◽  
Sandwich Assay ◽  
Humoral Immune ◽  
Liquid Samples

AbstractBackgroundColostrum provides an immune sharing between a mother and her infant. The transfer in colostrum of antibodies against SARS-CoV-2 and the elicited cytokines may provide crucial protection to the infant. There is limited literature on the immune response to SARS-CoV-2 present in colostrum.ObjectiveTo evaluate the presence of antibodies specific to SARS-CoV-2 and the associated cytokines in colostrum from women who tested positive for the virus.Study DesignBetween March and September 2020 we obtained bilateral colostrum samples collected on spot cards within 48 hours of delivery from 15 new mothers who had previously tested positive for SARS-CoV-2. Five of these 15 COVID-19 positive women also provided bilateral liquid colostrum within 1-2 days of providing the spot card samples. Archived bilateral colostrum samples collected from 8 women during 2011-2013 were used as pre-COVID-19 controls. All samples were tested for reactivity to the Receptor Binding Domain (RBD) of the SARS-CoV-2 spike protein using an ELISA that measures SARS-CoV-2 RBD-specific IgA, IgG, and IgM, and for concentrations of 10 inflammatory cytokines (IFNγ, TNFα, IL-1β, IL-2, IL-4, IL-6, IL-8, IL-10, IL-12, IL-13) using a multiplex electrochemiluminescent sandwich assay.ResultsBilateral colostrum samples from 73%, 73% and 33% of the 15 COVID-19 mothers exhibited IgA, IgG, and IgM reactivity to RBD respectively. Colostrum samples from two of the 8 pre-pandemic controls showed IgA and IgG reactivity to RBD. Additionally, COVID-19 mothers had significantly higher levels of 9 of the 10 inflammatory markers (all except IFNγ) as compared to the pre-COVID-19 controls. Comparable results were obtained with both the spot card-eluates and liquid samples.ConclusionsA strong humoral immune response is present in the colostrum of women who were infected with SARS-CoV-2 before delivering. High levels of 9 inflammatory markers were also present in the colostrum. The evolution and duration of the antibody response, as well as dynamics of the cytokine response, remain to be determined. Our results also indicate that future large-scale studies can be conducted with milk easily collected on paper spot cards.

scholarly journals Covalent coupling of Spike’s receptor binding domain to a multimeric carrier produces a high immune response against SARS-CoV-2

2022 ◽  
Vol 12 (1) ◽  
Author(s):  
◽  
Paula M. Berguer ◽  
Matías Blaustein ◽  
Luis M. Bredeston ◽  
Patricio O. Craig ◽  
...  
Keyword(s):  
Immune Response ◽  
Receptor Binding ◽  
Neutralizing Antibodies ◽  
High Titer ◽  
Binding Domain ◽  
Efficient Production ◽  
Receptor Binding Domain ◽  
Promising Candidate ◽  
Humoral Immune ◽  
Lumazine Synthase

AbstractThe receptor binding domain (RBD) of the Spike protein from SARS-CoV-2 is a promising candidate to develop effective COVID-19 vaccines since it can induce potent neutralizing antibodies. We have previously reported the highly efficient production of RBD in Pichia pastoris, which is structurally similar to the same protein produced in mammalian HEK-293T cells. In this work we designed an RBD multimer with the purpose of increasing its immunogenicity. We produced multimeric particles by a transpeptidation reaction between RBD expressed in P. pastoris and Lumazine Synthase from Brucella abortus (BLS), which is a highly immunogenic and very stable decameric 170 kDa protein. Such particles were used to vaccinate mice with two doses 30 days apart. When the particles ratio of RBD to BLS units was high (6–7 RBD molecules per BLS decamer in average), the humoral immune response was significantly higher than that elicited by RBD alone or by RBD-BLS particles with a lower RBD to BLS ratio (1–2 RBD molecules per BLS decamer). Remarkably, multimeric particles with a high number of RBD copies elicited a high titer of neutralizing IgGs. These results indicate that multimeric particles composed of RBD covalent coupled to BLS possess an advantageous architecture for antigen presentation to the immune system, and therefore enhancing RBD immunogenicity. Thus, multimeric RBD-BLS particles are promising candidates for a protein-based vaccine.

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