scholarly journals Development of an in-house COVID-19 serology ELISA Test

2021 ◽  
Vol 2021 (2) ◽  
Author(s):  
Shazeda Chowdhury ◽  
Sara Taleb ◽  
Mahmoud Elgamal ◽  
Nesyia Hassan ◽  
Michail Nomikos ◽  
...  
Keyword(s):  
Neutralizing Antibodies ◽  
Elisa Test ◽  
Negative Control ◽  
Surface Proteins ◽  
Diagnostic Tools ◽  
High Antibody ◽  
Serum Samples ◽  
Specific Antibodies ◽  
Antibody Titers ◽  
Serology Test

Background: COVID-19 pandemic created an unprecedented demand for reagents and diagnostic tools to confirm COVID-19 cases. Thus, the development of a robust in-house diagnostic test is considered of high importance. Within a few days after exposure, the human body produces specific antibodies that recognize the surface proteins of the invading SARS-CoV-2 virus. Therefore, virus specific immunoglobulins are neutralizing antibodies and their appearance in the blood is a good sign of immunity. The aim of this study was to develop an in-house COVID-19 serology ELISA test to quantify induced antibody responses. This test can help identify convalescent plasma donors with high antibody titers that can be used to treat other patients. Methods: Spike protein antigen is highly expressed in SARS-CoV-2. Recombinant protein corresponding to the spike receptor-binding domain (RBD), which binds to specific antibodies circulating in COVID-19 patients’ blood was used as the antigen in this colorimetric ELISA test. Briefly, a 96-microtiter well plate was coated with RBD protein, where serum dilutions were added. Antibody titers were detected using an anti-human IgG- peroxidase labelled antibody and the substrate o-phenylenediamine dihydrochloride; measured at optical density (OD) of 450 nm. Results: The in-house quantitative serology test was validated using serum samples collected from severe COVID-19 patients (n = 282) admitted to the intensive care unit at Hamad General Hospital. Serum samples from non-COVID-19 (n = 10) were used as a negative control. We detected high antibody titers in ~90% of COVID-19 sera. In contrast, no SARS-CoV-2 specific antibodies were detected in the serum of non-infected subjects (n = 6), pooled human serum collected before 2019, or Middle East Respiratory Syndrome (MERS) infected subjects (n = 3) confirming the specificity and the sensitivity of this in-house serology test. Conclusion: This in-house quantitative serology test is sensitive, specific, and inexpensive. The test can address the rising issue of COVID-19 supply chain globally and foster the capacity-building efforts envisioned by Qatar University.

scholarly journals Serological response to rabies virus induced by commercial vaccines in cattle

2017 ◽  
Vol 47 (10) ◽  
Author(s):  
Mathias Martins ◽  
João Motta de Quadros ◽  
Eduardo Furtado Flores ◽  
Rudi Weiblen
Keyword(s):  
Rabies Virus ◽  
Neutralizing Antibodies ◽  
Vaccine Dose ◽  
Booster Vaccination ◽  
Serological Response ◽  
Serum Samples ◽  
Anamnestic Response ◽  
Post Vaccination ◽  
Antibody Titers ◽  
One Year

ABSTRACT: The antibody response to rabies virus (RABV) induced by commercial vaccines in heifers was investigated. For this, 84 heifers were vaccinated twice (30 days interval) with each of four vaccines (G1 = 14 animals; G2 = 24; G3 = 22 and G4 = 24) and received a booster vaccination 360 days later. Serum samples collected at different intervals after vaccination and 30 days after booster were submitted to a virus neutralizing (VN) assay for RABV antibodies. Thirty days after the second vaccine dose, 92% of the immunized animals presented VN titers ≥0.5UI/mL (geometric medium titers [GMT] 1.7 to 3.8UI/mL). At the day of the booster (360 days post-vaccination); however, the percentage of animals harboring antibody titers ≥0.5UI/mL had dropped to 31% (0-80% of the animals, depending on the vaccine), resulting in lower GMT (0.1 to 0.6UI/mL). Booster vaccination at day 360 resulted in a detectable anamnestic response in all groups, resulting in 83% of animals (65 to 100%) harboring VN titers ≥0.5UI/mL thirty days later (GMT 0.6 to 4.3UI/mL). These results indicated that these vaccines were able to induce an adequate anti-RABV response in all animals after prime vaccination (and after booster as well). However, the titers decreased, reaching titers <0.5UI/mL in approximately 70% of animals within the interval before the recommended booster. Thus, booster vaccination for rabies in cattle using the current vaccines should be performed before the recommended one-year interval, as to maintain neutralizing antibodies levels in most vaccinated animals.

scholarly journals Antibody titers measured by commercial assays are correlated with neutralizing antibody titers calibrated by international standards

2021 ◽  
Author(s):  
Yu-An Kung ◽  
Chung-Guei Huang ◽  
Sheng-Yu Huang ◽  
Kuan-Ting Liu ◽  
Peng-Nien Huang ◽  
...  
Keyword(s):  
Neutralizing Antibodies ◽  
Vaccine Development ◽  
Geometric Mean ◽  
Neutralizing Antibody ◽  
Diagnostic Techniques ◽  
International Standards ◽  
World Health ◽  
Serum Samples ◽  
Antibody Titers ◽  
Health Organization

The World Health Organization (WHO) has highlighted the importance of an international standard (IS) for SARS-CoV-2 neutralizing antibody titer detection, with the aim of calibrating different diagnostic techniques. In this study, IS was applied to calibrate neutralizing antibody titers (IU/mL) and binding antibody titers (BAU/mL) in response to SARS-CoV-2 vaccines. Serum samples were collected from participants receiving the Moderna (n = 20) and Pfizer (n = 20) vaccines at three time points: pre-vaccination, after one dose, and after two doses. We obtained geometric mean titers of 1404.16 and 928.75 IU/mL for neutralizing antibodies after two doses of the Moderna and Pfizer vaccines, respectively. These values provide an important baseline for vaccine development and the implementation of non-inferiority trials. We also compared three commercially available kits from Roche, Abbott, and MeDiPro for the detection of COVID-19 antibodies based on binding affinity to S1 and/or RBD. Our results demonstrated that antibody titers measured by commercial assays are highly correlated with neutralizing antibody titers calibrated by IS.

scholarly journals Can we predict antibody responses in SARS-CoV-2? A cohort analysis

2021 ◽  
Author(s):  
Mary Gaeddert ◽  
Philip Kitchen ◽  
Tobias Broger ◽  
Stefan Weber ◽  
Ralf Bartenschlager ◽  
...  
Keyword(s):  
Neutralizing Antibodies ◽  
Severe Disease ◽  
Cohort Analysis ◽  
Antibody Responses ◽  
High Antibody ◽  
Igg Antibodies ◽  
Rt Pcr ◽  
Median Increase ◽  
Antibody Titers

AbstractBackgroundAfter infection with severe acute respiratory syndrome coronavirus (SARS-CoV-2), Immunoglobulin G (IgG) antibodies and virus-specific neutralizing antibodies (nAbs) develop. This study describes antibody responses in a cohort of recovered COVID-19 patients to identify predictors.MethodsWe recruited patients with confirmed SARS-CoV-2 infection from Heidelberg, Germany. Blood samples were collected three weeks after COVID-19 symptoms ended. Participants with high antibody titers were invited for follow-up visits. IgG titers were measured by the Euroimmun Assay, and nAbs titers in a SARS-CoV-2 infection-based assay.Results281 participants were enrolled between April and August 2020 with IgG testing, 145 (51.6%) had nAbs, and 35 (12.5%) had follow-up. The median IgG optical density (OD) ratio was 3.1 (Interquartile range (IQR) 1.6-5.1), and 24.1% (35/145) had a nAb titer>1:80. Higher IgG titers were associated with increased age and more severe disease, and higher nAbs were associated with male gender and CT-value of 25-30 on RT-PCR at diagnosis. The median IgG OD ratio on follow-up was 3.7 (IQR 2.9-5.9), a median increase of 0.5 (IQR −0.3-1.7). Six participants with follow-up nAbs all had titers ≤ 1:80.ConclusionsWhile age and disease severity were correlated with IgG responses, predictive factors for nAbs in convalescent patients remain unclear.

Prevalence and Persistence of Maternal Dengue Neutralizing Antibodies in Infants From Central and Southern Thailand: A Retrospective Cohort Study

2019 ◽  
Vol 31 (4) ◽  
pp. 288-295 ◽  
Author(s):  
Adrienne Guignard ◽  
François Haguinet ◽  
Stéphanie Wéry ◽  
Phirangkul Kerdpanich
Keyword(s):  
Neutralizing Antibodies ◽  
Vaccine Development ◽  
Geometric Mean ◽  
Neutralizing Antibody ◽  
Maternal Antibodies ◽  
Childhood Immunization ◽  
Serum Samples ◽  
Denv Serotypes ◽  
Antibody Titers ◽  
Antibody Kinetics

Understanding maternal dengue virus (DENV) neutralizing antibody kinetics in infants remains timely to develop a safe and effective childhood immunization. This retrospective study evaluated the prevalence and persistence of maternal antibody titers against DENV serotypes 1 to 4 in 139 Thai infants at 2, 6, and 7 months of age, using serum samples collected in a vaccination trial ( http://clinicaltrials.gov ; NCT00197275). Neutralizing antibodies against all 4 DENV serotypes were detected in 87.8% and 22.9% of infants at 2 and 7 months, respectively. At 2 months, DENV-4 neutralizing antibody geometric mean titers were notably lower (80) compared with DENV-1 to DENV-3 (277-471). Our results corroborate previous findings that DENV-1 to DENV-4 maternal antibodies persist at 7 months despite titers decrease from 2 months onwards. As persisting maternal antibodies may inhibit immune responses in DENV-vaccinated infants, a comprehensive understanding of DENV antibody kinetics is required in the perspective of vaccine development for infants.

Lav/HTLV-III Neutralizing Antibodies in the Sera of Patients with Aids, Lymphadenopathy Syndrome and Asymptomatic Seropositive Individuals

1986 ◽  
Vol 72 (3) ◽  
pp. 219-224 ◽  
Author(s):  
Georgine P. Faulkner-Valle ◽  
Anita De Rossi ◽  
Oriella Dalla Gassa ◽  
Luigi Chieco-Bianchi
Keyword(s):  
Western Blot ◽  
Blot Analysis ◽  
Western Blot Analysis ◽  
Neutralizing Antibodies ◽  
Elisa Test ◽  
Antibody Specificity ◽  
Serum Samples ◽  
Neutralizing Activity ◽  
The Mean ◽  
Asymptomatic Individuals

Serum samples which had previously been found positive for LAV/HTLV-III antibodies by the ELISA test and then confirmed by radioimmunoassay (Western blot) were tested for the presence of neutralizing antibodies. No neutralizing activity was found in the sera of a group of patients with the clinical diagnosis of AIDS. However in patients with LAS and other related pathologic conditions the percentage of sera positive for neutralizing antibodies was 27 % and 55 % respectively. At least 50 % of the sera from seropositive asymptomatic individuals had neutralizing activity but with the exception of the haemophiliac group the mean titre was much lower than that of LAS patients. No relationship was found between the neutralizing titre and the antibody specificity detected by Western blot analysis for p41 and p120.

scholarly journals Age-Specific Seroprevalences of Merkel Cell Polyomavirus, Human Polyomaviruses 6, 7, and 9, and Trichodysplasia Spinulosa-Associated Polyomavirus

2013 ◽  
Vol 20 (3) ◽  
pp. 363-368 ◽  
Author(s):  
Jérôme T. J. Nicol ◽  
Rémy Robinot ◽  
Audrey Carpentier ◽  
Giovanni Carandina ◽  
Elisa Mazzoni ◽  
...  
Keyword(s):  
Merkel Cell Polyomavirus ◽  
Human Polyomavirus ◽  
Merkel Cell ◽  
High Antibody ◽  
Specific Antibodies ◽  
Virus Like Particle ◽  
Antibody Titers ◽  
Immunosorbent Assays ◽  
Human Polyomaviruses

ABSTRACTSix new human polyomaviruses have been identified since 2008 (Merkel cell polyomavirus [MCPyV], human polyomavirus 6 [HPyV6], HPyV7, HPyV9, trichodysplasia spinulosa polyomavirus [TSPyV], and Malawi polyomavirus [MWPyV]). The presence of specific antibodies against MCPyV, HPyV6, HPyV7, HPyV9, and TSPyV in 828 Italian subjects aged 1 to 100 years was investigated by virus-like particle-based enzyme-linked immunosorbent assays (ELISAs). The findings indicate that all of these new polyomaviruses circulate widely in humans, with seroprevalences in adulthood ranging from 39.4% for HPyV9 to 87.1% for MCPyV, and that primary exposure is most intense in childhood, with the exception of HPyV7 and HPyV9, for which the seroprevalence increased throughout life. The proportion of subjects with high antibody titers was found to increase with age for MCPyV and to decrease with age for TSPyV.

scholarly journals Detection of neutralizing antibodies against SARS-CoV-2 by using a commercial surrogate virus neutralization ELISA: can it substitute the classical neutralization test?

2021 ◽  
Author(s):  
Natalie E Hofmann ◽  
Marica Grossegesse ◽  
Markus Neumann ◽  
Lars Schaade ◽  
Andreas Nitsche
Keyword(s):  
Neutralizing Antibodies ◽  
Diagnostic Performance ◽  
Neutralization Test ◽  
Virus Neutralization ◽  
Serum Samples ◽  
Population Immunity ◽  
Vaccine Trials ◽  
Binding Inhibition ◽  
Antibody Titers ◽  
High Throughput Detection

Background: High-throughput detection of neutralizing antibodies against SARS-CoV-2 presents a valuable tool for vaccine trials or investigations of population immunity. We evaluate the performance of the first commercial surrogate virus neutralization test (sVNT, GenScript Biotech) against SARS-CoV-2 plaque reduction neutralization test (PRNT) in convalescent and vaccinated individuals. We compare it to five other ELISAs, two of which are designed to detect neutralizing antibodies. Results: In 491 pre-vaccination serum samples, sVNT missed 23.6% of PRNT-positive samples when using the manufacturer-recommended cutoff of 30% binding inhibition. Introducing a equivocal area between 15 and 35% maximized sensitivity and specificity against PRNT to 72.8-93.1 % and 73.5-97.6%, respectively. The overall diagnostic performance of the other ELISAs for neutralizing antibodies was below that of sVNT. Vaccinated individuals exhibited higher antibody titers by PRNT (median 119.8, IQR 56.7-160) and binding inhibition by sVNT (median 95.7, IQR 88.1-96.8) than convalescent patients (median 49.1, IQR 20-62; median 52.9, IQR 31.2-76.2). Conclusion: GenScript sVNT is suitable to screen for SARS-CoV-2-neutralizing antibodies; however, to obtain accurate results, confirmatory testing by PRNT in a equivocal area is required. This equivocal area may require adaptation for use in vaccinated individuals, due to higher antibody titers.

scholarly journals A high throughput reporter virus particle microneutralization assay for quantitation of Zika virus neutralizing antibodies in multiple species

PLoS ONE ◽  
2021 ◽  
Vol 16 (4) ◽  
pp. e0250516
Author(s):  
Kelly Bohning ◽  
Stephanie Sonnberg ◽  
Hui-Ling Chen ◽  
Melissa Zahralban-Steele ◽  
Timothy Powell ◽  
...  
Keyword(s):  
Virus Particle ◽  
High Throughput ◽  
Zika Virus ◽  
Neutralizing Antibodies ◽  
Surface Proteins ◽  
Mouse Serum ◽  
Reporter Virus ◽  
Serum Samples ◽  
Economic Sectors ◽  
Virus Surface

Zika virus is a Flavivirus, transmitted via Aedes mosquitos, that causes a range of symptoms including Zika congenital syndrome. Zika has posed a challenging situation for health, public and economic sectors of affected countries. To quantitate Zika virus neutralizing antibody titers in serum samples, we developed a high throughput plate based Zika virus reporter virus particle (RVP) assay that uses an infective, non-replicating particle encoding Zika virus surface proteins and capsid (CprME) and a reporter gene (Renilla luciferase). This is the first characterization of a Zika virus RVP assay in 384-well format using a Dengue replicon Renilla reporter construct. Serially diluted test sera were incubated with RVPs, followed by incubation with Vero cells. RVPs that have not been neutralized by antibodies in the test sera entered the cells and expressed Renilla luciferase. Quantitative measurements of neutralizing activity were determined using a plate-based assay and commercially available substrate. The principle of limiting the infection to a single round increases the precision of the assay measurements. RVP log10EC50 titers correlated closely with titers determined using a plaque reduction neutralization test (PRNT) (R2>95%). The plate-based Zika virus RVP assay also demonstrated high levels of precision, reproducibility and throughput. The assay employs identical reagents for human, rhesus macaque and mouse serum matrices. Spiking studies indicated that the assay performs equally well in different species, producing comparable titers irrespective of the serum species. The assay is conducted in 384-well plates and can be automated to simultaneously achieve high throughput and high reproducibility.

scholarly journals Commercial Serology Assays Predict Neutralization Activity against SARS-CoV-2

2020 ◽  
Author(s):  
Raymond T Suhandynata ◽  
Melissa A Hoffman ◽  
Deli Huang ◽  
Jenny T Tran ◽  
Michael J Kelner ◽  
...  
Keyword(s):  
Neutralizing Antibodies ◽  
Neutralizing Antibody ◽  
Neutralization Assay ◽  
Negative Control ◽  
Percentage Agreement ◽  
Positive Serology ◽  
Neutralization Activity ◽  
Antibody Titers ◽  
The Relationship ◽  
Positive Results

Abstract Background It is unknown whether a positive serology result correlates with protective immunity against SARS-CoV-2. There are also concerns regarding the low positive predictive value of SARS-CoV-2 serology tests, especially when testing populations with low disease prevalence. Methods A neutralization assay was validated in a set of PCR-confirmed positive specimens and in a negative cohort. In addition, 9530 specimens were screened using the Diazyme SARS-CoV-2 IgG serology assay and all positive results (N = 164 individuals) were reanalyzed using the neutralization assay, the Roche total immunoglobin assay, and the Abbott IgG assay. The relationship between the magnitude of a positive SARS-CoV-2 serology result and neutralizing activity was determined. Neutralizing antibody titers (50% inhibitory dilution, ID50) were also longitudinally monitored in patients confirmed to have SARS-CoV-2 by PCR. Results The SARS-CoV-2 neutralization assay had a positive percentage agreement (PPA) of 96.6% with a SARS-CoV-2 PCR test and a negative percentage agreement (NPA) of 98.0% across 100 negative control individuals. ID50 neutralization titers positively correlated with all 3 clinical serology platforms. Longitudinal monitoring of hospitalized PCR-confirmed patients with COVID-19 demonstrated they made high neutralization titers against SARS-CoV-2. PPA between the Diazyme IgG assay alone and the neutralization assay was 50.6%, while combining the Diazyme IgG assay with either the Roche or Abbott platforms increased the PPA to 79.2 and 78.4%, respectively. Conclusions These 3 clinical serology assays positively correlate with SARS-CoV-2 neutralization activity observed in patients with COVID-19. All patients confirmed SARS-CoV-2 positive by PCR develop neutralizing antibodies.

scholarly journals Antigenicity of the surface proteins of non o1 non o139 vibrio cholerae in rabbit model

1970 ◽  
Vol 19 (2) ◽  
pp. 129-135
Author(s):  
Tania S Bonny ◽  
Fazle Rabbi ◽  
Mahmuda Yasmin ◽  
Jamalun Nessa ◽  
Chowdhury R Ahsan
Keyword(s):  
Rabbit Model ◽  
Surface Antigens ◽  
Immunoblot Analysis ◽  
Surface Proteins ◽  
Diagnostic Tools ◽  
Coomassie Brilliant Blue ◽  
Antigenic Proteins ◽  
Sds Page ◽  
Antibody Titers ◽  
Coomassie Brilliant

An investigation was carried out to demonstrate the antigenicity of the surface proteins of the non O1 non O139 V. cholerae in rabbit model. Three rabbits were immunized with the surface proteins and sera were collected at one week intervals for six weeks. Sera were examined for antibodies specific for surface antigens using ELISA and the results showed a significant increase in antibody titers with time. The SDS?PAGE analysis showed a number of protein bands ranging from 21 to 131 kDa after Coomassie brilliant blue R250 staining. However, immunoblot analysis showed four major antigenic bands of 38, 43, 74 and 85 kDa to be prominent. These identified proteins might have higher immunogenic effects among humans infected with non O1 non O139 V. cholerae and some of these antigenic proteins could be included as diagnostic tools based on serology and also in vaccine preparations. Key words: Antigenicity; Surface proteins; Non O1 non O139 V. cholerae; Rabbit model DOI: http://dx.doi.org/10.3329/dujbs.v19i2.8954 DUJBS 2010; 19(2): 129-135

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