scholarly journals IMMUNO-COV v2.0: Development and Validation of a High-Throughput Clinical Assay for Measuring SARS-CoV-2-Neutralizing Antibody Titers

mSphere ◽  
2021 ◽  
Author(s):  
Rianna Vandergaast ◽  
Timothy Carey ◽  
Samantha Reiter ◽  
Chase Lathrum ◽  
Patrycja Lech ◽  
...  
Keyword(s):  
Immune Responses ◽  
High Throughput ◽  
Causative Agent ◽  
Neutralizing Antibody ◽  
Clinical Assay ◽  
Antibody Titers ◽  
Development And Validation

Since its emergence at the end of 2019, SARS-CoV-2, the causative agent of COVID-19, has caused over 100 million infections and 2.4 million deaths worldwide. Recently, countries have begun administering approved COVID-19 vaccines, which elicit strong immune responses and prevent disease in most vaccinated individuals.

scholarly journals Statistical Approach To Estimate Vaccinia-Specific Neutralizing Antibody Titers Using a High-Throughput Assay

2009 ◽  
Vol 16 (8) ◽  
pp. 1105-1112 ◽  
Author(s):  
Richard Kennedy ◽  
V. Shane Pankratz ◽  
Eric Swanson ◽  
David Watson ◽  
Hana Golding ◽  
...  
Keyword(s):  
Immune Responses ◽  
Vaccinia Virus ◽  
High Throughput ◽  
Large Scale ◽  
Statistical Approach ◽  
Neutralizing Antibody ◽  
Neutralization Assay ◽  
Virus Neutralization Assay ◽  
Antibody Titers ◽  
High Throughput Assay

ABSTRACT Because of the bioterrorism threat posed by agents such as variola virus, considerable time, resources, and effort have been devoted to biodefense preparation. One avenue of this research has been the development of rapid, sensitive, high-throughput assays to validate immune responses to poxviruses. Here we describe the adaptation of a β-galactosidase reporter-based vaccinia virus neutralization assay to large-scale use in a study that included over 1,000 subjects. We also describe the statistical methods involved in analyzing the large quantity of data generated. The assay and its associated methods should prove useful tools in monitoring immune responses to next-generation smallpox vaccines, studying poxvirus immunity, and evaluating therapeutic agents such as vaccinia virus immune globulin.

scholarly journals IMMUNO-COV™ v2.0: Development and Validation of a High-Throughput Clinical Assay for Measuring SARS-CoV-2-Neutralizing Antibody Titers

2021 ◽  
Author(s):  
Rianna Vandergaast ◽  
Timothy Carey ◽  
Samantha Reiter ◽  
Chase Lathrum ◽  
Patrycja Lech ◽  
...  
Keyword(s):  
High Throughput ◽  
Neutralizing Antibodies ◽  
Dynamic Range ◽  
Limit Of Detection ◽  
Neutralizing Antibody ◽  
Clinical Samples ◽  
Range Limit ◽  
Specificity And Sensitivity ◽  
Antibody Titers ◽  
Development And Validation

ABSTRACTNeutralizing antibodies are key determinants of protection from future infection, yet well-validated high-throughput assays for measuring titers of SARS-CoV-2-neutralizing antibodies are not generally available. Here we describe the development and validation of IMMUNO-COV™ v2.0 a scalable surrogate virus assay, which titrates antibodies that block infection of Vero-ACE2 cells by a luciferase-encoding vesicular stomatitis virus displaying SARS-CoV-2 spike glycoproteins (VSV-SARS2-Fluc). Antibody titers, calculated using a standard curve consisting of stepped concentrations of SARS-CoV-2 spike monoclonal antibody, correlated closely (p < 0.0001) with titers obtained from a gold-standard PRNT50% assay performed using a clinical isolate of SARS-CoV-2. IMMUNO-COV™ v2.0 was comprehensively validated using data acquired from 242 assay runs performed over seven days by five analysts, utilizing two separate virus lots, and 176 blood samples. Assay performance was acceptable for clinical use in human serum and plasma based on parameters including linearity, dynamic range, limit of blank and limit of detection, dilutional linearity and parallelism, precision, clinical agreement, matrix equivalence, clinical specificity and sensitivity, and robustness. Sufficient VSV-SARS2-Fluc virus reagent has been banked to test 5 million clinical samples. Notably, a significant drop in IMMUNO-COV™ v2.0 neutralizing antibody titers was observed over a six-month period in people recovered from SARS-CoV-2 infection. Together, our results demonstrate the feasibility and utility of IMMUNO-COV™ v2.0 for measuring SARS-CoV-2-neutralizing antibodies in vaccinated individuals and those recovering from natural infections. Such monitoring can be used to better understand what levels of neutralizing antibodies are required for protection from SARS-CoV-2, and what booster dosing schedules are needed to sustain vaccine-induced immunity.

scholarly journals Immune responses to SARS-CoV-2 infection in hospitalized pediatric and adult patients

2020 ◽  
Vol 12 (564) ◽  
pp. eabd5487 ◽  
Author(s):  
Carl A. Pierce ◽  
Paula Preston-Hurlburt ◽  
Yile Dai ◽  
Clare Burn Aschner ◽  
Natalia Cheshenko ◽  
...  
Keyword(s):  
T Cells ◽  
Immune Responses ◽  
Pediatric Patients ◽  
Neutralizing Antibody ◽  
Children And Youth ◽  
Spike Protein ◽  
Serum Concentrations ◽  
Antiviral Immune Response ◽  
Antibody Titers ◽  
Ifn Γ

Children and youth infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have milder disease than do adults, and even among those with the recently described multisystem inflammatory syndrome, mortality is rare. The reasons for the differences in clinical manifestations are unknown but suggest that age-dependent factors may modulate the antiviral immune response. We compared cytokine, humoral, and cellular immune responses in pediatric (children and youth, age <24 years) (n = 65) and adult (n = 60) patients with coronavirus disease 2019 (COVID-19) at a metropolitan hospital system in New York City. The pediatric patients had a shorter length of stay, decreased requirement for mechanical ventilation, and lower mortality compared to adults. The serum concentrations of interleukin-17A (IL-17A) and interferon-γ (IFN-γ), but not tumor necrosis factor–α (TNF-α) or IL-6, were inversely related to age. Adults mounted a more robust T cell response to the viral spike protein compared to pediatric patients as evidenced by increased expression of CD25+ on CD4+ T cells and the frequency of IFN-γ+ CD4+ T cells. Moreover, serum neutralizing antibody titers and antibody-dependent cellular phagocytosis were higher in adults compared to pediatric patients with COVID-19. The neutralizing antibody titer correlated positively with age and negatively with IL-17A and IFN-γ serum concentrations. There were no differences in anti-spike protein antibody titers to other human coronaviruses. Together, these findings demonstrate that the poor outcome in hospitalized adults with COVID-19 compared to children may not be attributable to a failure to generate adaptive immune responses.

scholarly journals A Combination Adjuvant for the Induction of Potent Antiviral Immune Responses for a Recombinant SARS-CoV-2 Protein Vaccine

2021 ◽  
Vol 12 ◽  
Author(s):  
Sonia Jangra ◽  
Jeffrey J. Landers ◽  
Raveen Rathnasinghe ◽  
Jessica J. O’Konek ◽  
Katarzyna W. Janczak ◽  
...  
Keyword(s):  
Immune Responses ◽  
Neutralizing Antibody ◽  
Cellular Responses ◽  
Cross Protection ◽  
Protein Vaccine ◽  
Mucosal Vaccination ◽  
Adaptive Immune ◽  
Antibody Titers ◽  
High Virus

Several SARS-CoV-2 vaccines have received EUAs, but many issues remain unresolved, including duration of conferred immunity and breadth of cross-protection. Adjuvants that enhance and shape adaptive immune responses that confer broad protection against SARS-CoV-2 variants will be pivotal for long-term protection as drift variants continue to emerge. We developed an intranasal, rationally designed adjuvant integrating a nanoemulsion (NE) that activates TLRs and NLRP3 with an RNA agonist of RIG-I (IVT DI). The combination adjuvant with spike protein antigen elicited robust responses to SARS-CoV-2 in mice, with markedly enhanced TH1-biased cellular responses and high virus-neutralizing antibody titers towards both homologous SARS-CoV-2 and a variant harboring the N501Y mutation shared by B1.1.7, B.1.351 and P.1 variants. Furthermore, passive transfer of vaccination-induced antibodies protected naive mice against heterologous viral challenge. NE/IVT DI enables mucosal vaccination, and has the potential to improve the immune profile of a variety of SARS-CoV-2 vaccine candidates to provide effective cross-protection against future drift variants.

scholarly journals Immunogenicity of a Live-Attenuated Dengue Vaccine Using a Heterologous Prime-Boost Strategy in a Phase 1 Randomized Clinical Trial

2020 ◽  
Author(s):  
Leyi Lin ◽  
Michael A Koren ◽  
Kristopher M Paolino ◽  
Kenneth H Eckels ◽  
Rafael De La Barrera ◽  
...  
Keyword(s):  
Immune Responses ◽  
Dengue Virus ◽  
Phase 1 ◽  
Vaccination Strategy ◽  
Neutralizing Antibody ◽  
Dengue Vaccine ◽  
Serious Adverse Events ◽  
Live Attenuated Vaccine ◽  
Antibody Titers ◽  
Global Health Problem

Abstract Background Dengue is a global health problem and the development of a tetravalent dengue vaccine with durable protection is a high priority. A heterologous prime-boost strategy has the advantage of eliciting immune responses through different mechanisms and therefore may be superior to homologous prime-boost strategies for generating durable tetravalent immunity. Methods In this phase 1 first-in-human heterologous prime-boost study, 80 volunteers were assigned to 4 groups and received a tetravalent dengue virus (DENV-1–4) purified inactivated vaccine (TDENV-PIV) with alum adjuvant and a tetravalent dengue virus (DENV-1–4) live attenuated vaccine (TDENV-LAV) in different orders and dosing schedules (28 or 180 days apart). Results All vaccination regimens had acceptable safety profiles and there were no vaccine-related serious adverse events. TDEN-PIV followed by TDEN-LAV induced higher neutralizing antibody titers and a higher rate of tetravalent seroconversions compared to TDEN-LAV followed by TDEN-PIV. Both TDEN-PIV followed by TDEN-LAV groups demonstrated 100% tetravalent seroconversion 28 days following the booster dose, which was maintained for most of these subjects through the day 180 measurement. Conclusions A heterologous prime-boost vaccination strategy for dengue merits additional evaluation for safety, immunogenicity, and potential for clinical benefit. Clinical Trials Registration NCT02239614.

scholarly journals Potential SARS-CoV-2 Immune Correlates of Protection in Infection and Vaccine Immunization

Pathogens ◽  
2021 ◽  
Vol 10 (2) ◽  
pp. 138
Author(s):  
Yongjun Sui ◽  
Yonas Bekele ◽  
Jay A. Berzofsky
Keyword(s):  
Immune Responses ◽  
Neutralizing Antibody ◽  
Current Data ◽  
Innate Immune ◽  
Type I Interferons ◽  
Type I ◽  
Risk Of Infection ◽  
Correlates Of Protection ◽  
Immune Correlates ◽  
Antibody Titers

Both SARS-CoV-2 infections and vaccines induce robust immune responses. Current data suggested that high neutralizing antibody titers with sustained Th1 responses might correlate with protection against viral transmission and disease development and severity. In addition, genetic and innate immune factors, including higher levels of type I interferons, as well as the induction of trained immunity and local mucosal immunity also contribute to lower risk of infection and amelioration of disease severity. The identification of immune correlates of protection will facilitate the development of effective vaccines and therapeutics strategies.

scholarly journals Neutralizing antibody against SARS-CoV-2 spike in COVID-19 patients, health care workers and convalescent plasma donors: a cohort study using a rapid and sensitive high-throughput neutralization assay

2020 ◽  
Author(s):  
Cong Zeng ◽  
John P Evans ◽  
Rebecca Pearson ◽  
Panke Qu ◽  
Yi-Min Zheng ◽  
...  
Keyword(s):  
Health Care ◽  
High Throughput ◽  
Health Care Workers ◽  
Neutralizing Antibody ◽  
Neutralization Assay ◽  
Luciferase Reporter ◽  
Human Antibody ◽  
Icu Patients ◽  
Care Workers ◽  
Antibody Titers

Rapid and specific antibody testing is crucial for improved understanding, control, and treatment of COVID-19 pathogenesis. Herein, we describe and apply a rapid, sensitive, and accurate virus neutralization assay for SARS-CoV-2 antibodies. The new assay is based on an HIV-1 lentiviral vector that contains a secreted intron Gaussia luciferase or secreted Nano-luciferase reporter cassette, pseudotyped with the SARS-CoV-2 spike (S) glycoprotein, and is validated with a plaque reduction assay using an authentic, infectious SARS-CoV-2 strain. The new assay was used to evaluate SARS-CoV-2 antibodies in serum from individuals with a broad range of COVID-19 symptoms, including intensive care unit (ICU) patients, health care workers (HCWs), and convalescent plasma donors. The highest neutralizing antibody titers were observed among ICU patients, followed by general hospitalized patients, HCWs and convalescent plasma donors. Our study highlights a wide phenotypic variation in human antibody responses against SARS-CoV-2, and demonstrates the efficacy of a novel lentivirus pseudotype assay for high-throughput serological surveys of neutralizing antibody titers in large cohorts.

scholarly journals Competent immune responses to SARS-CoV-2 variants in older adults following mRNA vaccination

2021 ◽  
Author(s):  
Mladen Jergović ◽  
Jennifer L. Uhrlaub ◽  
Makiko Watanabe ◽  
Christine M. Bradshaw ◽  
Lisa M. White ◽  
...  
Keyword(s):  
Older Adults ◽  
Immune Responses ◽  
Cellular Immunity ◽  
Neutralizing Antibody ◽  
Humoral And Cellular Immunity ◽  
Age Cohorts ◽  
Receptor Repertoire ◽  
Depth Analysis ◽  
Repertoire Diversity ◽  
Antibody Titers

Aging is associated with a reduced magnitude of primary immune responses to vaccination and constriction of immune receptor repertoire diversity. Clinical trials demonstrate high efficacy of mRNA based SARS-CoV-2 vaccines in older adults but concerns about virus variant escape have not been well addressed. We have conducted an in-depth analysis of humoral and cellular immunity against an early-pandemic viral isolate and compared that to the P.1. (Gamma) and B.1.617.2 (Delta) variants, as well to a B.1.595 SARS-CoV-2 isolate bearing Spike mutation E484Q, in <55 and >65 age cohorts of mRNA vaccine recipients. As reported, robust immunity required the second dose of vaccine. Older vaccine recipients exhibited an expected 3-5x reduction (but not a complete loss) in neutralizing antibody titers against both P.1. (Gamma) and the B.1.595 virus at the peak of the boosted response. However, older vaccinees manifest robust cellular immunity against early-pandemic SARS-CoV-2 and more recent variants, which remained statistically comparable to the adult group. While the duration of these immune responses remains to be determined over longer periods of time, these results provide reasons for optimism regarding vaccine protection of older adults against SARS-CoV-2 variants and inform thinking about boost vaccination with variant vaccines.

scholarly journals Rapid Immune Responses to a Botulinum Neurotoxin Hc Subunit Vaccine throughIn VivoTargeting to Antigen-Presenting Cells

2011 ◽  
Vol 79 (8) ◽  
pp. 3388-3396 ◽  
Author(s):  
David M. White ◽  
Sabine Pellett ◽  
Mark A. Jensen ◽  
William H. Tepp ◽  
Eric A. Johnson ◽  
...  
Keyword(s):  
Immune Responses ◽  
Neutralizing Antibody ◽  
Antigen Presenting Cells ◽  
Binding Assays ◽  
Content Type ◽  
Botulinum Neurotoxins ◽  
Protein Toxins ◽  
Antibody Titers ◽  
Antigen Presenting

ABSTRACTThe clostridial botulinum neurotoxins (BoNTs) are the most potent protein toxins known. The carboxyl-terminal fragment of the toxin heavy chain (Hc) has been intensively investigated as a BoNT vaccine immunogen. We sought to determine whether targeting Hc to antigen-presenting cells (APCs) could accelerate the immune responses to vaccination with BoNT serotype A (BoNT/A) Hc. To test this hypothesis, we targeted Hc to the Fc receptors for IgG (FcγRs) expressed by dendritic cells (DCs) and other APCs. Hc was expressed as a fusion protein with a recombinant ligand for human FcγRs (R4) to produce HcR4 or a similar ligand for murine FcγRs to produce HcmR4. HcR4, HcmR4, and Hc were produced as secreted proteins using baculovirus-mediated expression in SF9 insect cells.In vitroreceptor binding assays showed that HcR4 effectively targets Hc to all classes of FcγRs. APCs loaded with HcR4 or HcmR4 are substantially more effective at stimulating Hc-reactive T cells than APCs loaded with nontargeted Hc. Mice immunized with a single dose of HcmR4 or HcR4 had earlier and markedly higher Hc-reactive antibody titers than mice immunized with nontargeted Hc. These results extend to BoNT neutralizing antibody titers, which are substantially higher in mice immunized with HcmR4 than in mice immunized with Hc. Our results demonstrate that targeting Hc to FcγRs augments the pace and magnitude of immune responses to Hc.

scholarly journals A Solution with Ginseng Saponins and Selenium as Vaccine Diluent to Increase Th1/Th2 Immune Responses in Mice

2020 ◽  
Vol 2020 ◽  
pp. 1-13
Author(s):  
Yong Wang ◽  
Xuemei Cui ◽  
Lijia Yuan ◽  
Babar Maqbool ◽  
Wei Xu ◽  
...  
Keyword(s):  
Immune Responses ◽  
Pseudorabies Virus ◽  
Saline Solution ◽  
Neutralizing Antibody ◽  
Cytolytic Activity ◽  
Routine Practice ◽  
Antibody Titers ◽  
Specific Igg ◽  
Swine Herds ◽  
Stem Leaf

Pseudorabies is an important infectious disease of swine, and immunization using attenuated pseudorabies virus (aPrV) vaccine is a routine practice to control this disease in swine herds. This study was to evaluate a saline solution containing ginseng stem-leaf saponins (GSLS) and sodium selenite (Se) as a vaccine adjuvant for its enhancement of immune response to aPrV vaccine. The results showed that aPrV vaccine diluted with saline containing GSLS-Se (aP-GSe) induced significantly higher immune responses than that of the vaccine diluted with saline alone (aP-S). The aP-GSe promoted higher production of gB-specific IgG, IgG1, and IgG2a, neutralizing antibody titers, secretion of Th1-type (IFN-γ, IL-2, IL-12), and Th2-type (IL-4, IL-6, IL-10) cytokines, and upregulated the T-bet/GATA-3 mRNA expression when compared to aP-S. In addition, cytolytic activity of NK cells, lymphocyte proliferation, and CD4+/CD8+ ratio was also significantly increased by aP-GSe. More importantly, aP-GSe conferred a much higher resistance of mice to a field virulent pseudorabies virus (fPrV) challenge. As the present study was conducted in mice, further study is required to evaluate the aP-GSe to improve the vaccination against PrV in swine.

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