scholarly journals Evaluation of a Commercial Culture-free Neutralization Antibody Detection Kit for Severe Acute Respiratory Syndrome-Related Coronavirus-2 and Comparison with an Anti-RBD ELISA Assay

2021 ◽  
Author(s):  
Jesse Papenburg ◽  
Matthew P. Cheng ◽  
Rachel Corsini ◽  
Chelsea Caya ◽  
Emelissa Mendoza ◽  
...  
Keyword(s):  
Severe Acute Respiratory Syndrome ◽  
Neutralizing Antibodies ◽  
Neutralization Test ◽  
Neutralization Assay ◽  
Added Value ◽  
Antibody Detection ◽  
Elisa Assay ◽  
Commercial Culture ◽  
Viral Culture ◽  
Iga Antibodies

ABSTRACTBackgroundSARS-CoV-2 surrogate neutralization assays that obviate the need for viral culture offer substantial advantages regarding throughput and cost. The cPass SARS-CoV-2 Neutralization Antibody Detection Kit (Genscript) is the first such commercially available assay, detecting antibodies that block RBD/ACE-2 interaction. We aimed to evaluate cPass to inform its use and assess its added value compared to anti-RBD ELISA assays.MethodsSerum reference panels comprising 205 specimens were used to compare cPass to plaque-reduction neutralization test (PRNT) and a pseudotyped lentiviral neutralization (PLV) assay for detection of neutralizing antibodies. We assessed the correlation of cPass with an ELISA detecting anti-RBD IgG, IgM, and IgA antibodies at a single timepoint and across intervals from onset of symptoms of SARS-CoV-2 infection.ResultsCompared to PRNT-50, cPass sensitivity ranged from 77% - 100% and specificity was 95% - 100%. Sensitivity was also high compared to the pseudotyped lentiviral neutralization assay (93% [95%CI 85-97]), but specificity was lower (58% [95%CI 48-67]). Highest agreement between cPass and ELISA was for anti-RBD IgG (r=0.823). Against the pseudotyped lentiviral neutralization assay, anti-RBD IgG sensitivity (99% [95%CI 94-100]) was very similar to that of cPass, but overall specificity was lower (37% [95%CI 28-47]). Against PRNT-50, results of cPass and anti-RBD IgG were nearly identical.ConclusionsThe added value of cPass compared to an IgG anti-RBD ELISA was modest.

scholarly journals Evaluation of a Commercial Culture-free Neutralization Antibody Detection Kit for Severe Acute Respiratory Syndrome-Related Coronavirus-2 and Comparison with an Anti-RBD ELISA Assay

2021 ◽  
Author(s):  
Jesse Papenburg ◽  
Matthew P Cheng ◽  
Rachel Corsini ◽  
Chelsea Caya ◽  
Emelissa Mendoza ◽  
...  
Keyword(s):  
Severe Acute Respiratory Syndrome ◽  
Neutralizing Antibodies ◽  
Neutralization Test ◽  
Neutralization Assay ◽  
Added Value ◽  
Antibody Detection ◽  
Elisa Assay ◽  
Commercial Culture ◽  
Viral Culture ◽  
Iga Antibodies

Abstract Background SARS-CoV-2 surrogate neutralization assays that obviate the need for viral culture offer substantial advantages regarding throughput and cost. The cPass SARS-CoV-2 Neutralization Antibody Detection Kit (Genscript) is the first such commercially available assay, detecting antibodies that block RBD/ACE-2 interaction. We aimed to evaluate cPass to inform its use and assess its added value compared to anti-RBD ELISA assays. Methods Serum reference panels comprising 205 specimens were used to compare cPass to plaque-reduction neutralization test (PRNT) and a pseudotyped lentiviral neutralization (PLV) assay for detection of neutralizing antibodies. We assessed the correlation of cPass with an ELISA detecting anti-RBD IgG, IgM, and IgA antibodies at a single timepoint and across intervals from onset of symptoms of SARS-CoV-2 infection. Results Compared to PRNT-50, cPass sensitivity ranged from 77% - 100% and specificity was 95% - 100%. Sensitivity was also high compared to the pseudotyped lentiviral neutralization assay (93% [95%CI 85-97]), but specificity was lower (58% [95%CI 48-67]). Highest agreement between cPass and ELISA was for anti-RBD IgG (r=0.823). Against the pseudotyped lentiviral neutralization assay, anti-RBD IgG sensitivity (99% [95%CI 94-100]) was very similar to that of cPass, but overall specificity was lower (37% [95%CI 28-47]). Against PRNT-50, results of cPass and anti-RBD IgG were nearly identical. Conclusions The added value of cPass compared to an IgG anti-RBD ELISA was modest.

scholarly journals Low serum neutralizing anti-SARS-CoV-2 S antibody levels in mildly affected COVID-19 convalescent patients revealed by two different detection methods

2020 ◽  
Author(s):  
Berislav Bošnjak ◽  
Saskia Catherina Stein ◽  
Stefanie Willenzon ◽  
Anne Katrin Cordes ◽  
Wolfram Puppe ◽  
...  
Keyword(s):  
Neutralizing Antibodies ◽  
Clinical Symptoms ◽  
Neutralization Test ◽  
Antibody Therapy ◽  
Neutralization Assay ◽  
Detection Methods ◽  
Angiotensin Converting Enzyme 2 ◽  
Iga Antibodies ◽  
Vesicular Stomatitis ◽  
Antibody Levels

Abstract Neutralizing antibodies targeting the receptor-binding domain (RBD) of the SARS-CoV-2 spike (S) block severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) entry into cells via surface-expressed angiotensin-converting enzyme 2 (ACE2). We used a surrogate virus neutralization test (sVNT) and SARS-CoV-2 S protein-pseudotyped vesicular stomatitis virus (VSV) vector-based neutralization assay (pVNT) to assess the degree to which serum antibodies from coronavirus disease 2019 (COVID-19) convalescent patients interfere with the binding of SARS-CoV-2 S to ACE2. Both tests revealed neutralizing anti-SARS-CoV-2 S antibodies in the sera of ~90% of mildly and 100% of severely affected COVID-19 convalescent patients. Importantly, sVNT and pVNT results correlated strongly with each other and to the levels of anti-SARS-CoV-2 S1 IgG and IgA antibodies. Moreover, levels of neutralizing antibodies correlated with the duration and severity of clinical symptoms but not with patient age. Compared to pVNT, sVNT is less sophisticated and does not require any biosafety labs. Since this assay is also much faster and cheaper, sVNT will not only be important for evaluating the prevalence of neutralizing antibodies in a population but also for identifying promising plasma donors for successful passive antibody therapy.

scholarly journals Novel surrogate virus neutralization test reveals low serum neutralizing anti-SARS-CoV-2-S antibodies levels in mildly affected COVID-19 convalescents

2020 ◽  
Author(s):  
Berislav Bošnjak ◽  
Saskia Catherina Stein ◽  
Stefanie Willenzon ◽  
Anne Katrin Cordes ◽  
Wolfram Puppe ◽  
...  
Keyword(s):  
Vesicular Stomatitis Virus ◽  
Neutralizing Antibodies ◽  
Clinical Symptoms ◽  
Neutralization Test ◽  
Antibody Therapy ◽  
Neutralization Assay ◽  
Receptor Binding Domain ◽  
Angiotensin Converting Enzyme 2 ◽  
Iga Antibodies ◽  
Vesicular Stomatitis

Neutralizing antibodies targeting the receptor-binding domain (RBD) of the SARS-CoV-2 spike (S) block severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) entry into cells using surface-expressed angiotensin-converting enzyme 2 (ACE2). We developed a surrogate neutralization test (sVNT) to assess at what degree serum antibodies interfere with the binding of SARS-CoV-2-S-RBD to ACE2. The sVNT revealed neutralizing anti-SARS-CoV-2-S antibodies in the sera of 90% of mildly and 100% of severely affected coronavirus-disease-2019 (COVID-19) convalescent patients. Importantly, sVNT results correlated strongly to the results from pseudotyped-vesicular stomatitis virus-vector-based neutralization assay and to levels of anti-SARS-CoV-2-S1 IgG and IgA antibodies. Moreover, levels of neutralizing antibodies also correlated to duration and severity of clinical symptoms, but not patient age or gender. These findings together with the sVNT will not only be important for evaluating the prevalence of neutralizing antibodies in a population but also for identifying promising plasma donors for successful passive antibody therapy.

scholarly journals Protective levels of canine distemper virus antibody in an urban dog population using plaque reduction neutralization test

2004 ◽  
Vol 71 (3) ◽  
Author(s):  
O.I. Oyedele ◽  
D.O. Oluwayelu ◽  
S.I.B. Cadmus ◽  
F.D. Adu
Keyword(s):  
Neutralizing Antibodies ◽  
Canine Distemper Virus ◽  
Neutralization Test ◽  
Neutralization Assay ◽  
Canine Distemper ◽  
Virus Antibody ◽  
Post Vaccination ◽  
Blood Samples ◽  
Highly Sensitive ◽  
Positive Serum

Blood samples from 50 dogs were collected at three veterinary clinics in Ibadan and Abuja, Nigeria and the serum from each sample was evaluated serologically for neutralizing antibodies against canine distemper virus (CDV) by the highly sensitive plaque reduction (PRN) neutralization assay. Thirteen dogs had plaque reduction neutralization titres of 0-100, seven had titres of 100-1 000 while 30 had titres ranging from 1 000-6 000. The PRN titres of vaccinated dogs were found to be significantly higher than unvaccinated dogs. The widespread use of the highly reproducible PRN test for the evaluation of antibody response to CDV may be very important in the generation of international CDV positive serum standards that should help to improve pre-and post-vaccination testing of dogs worldwide.

scholarly journals Evaluation of a multi-species SARS-CoV-2 surrogate virus neutralization test

2021 ◽  
Author(s):  
Carmen W.E. Embregts ◽  
Babs Verstrepen ◽  
Jan A.M. Langermans ◽  
Kinga P Boszormenyi ◽  
Reina S. Sikkema ◽  
...  
Keyword(s):  
Neutralizing Antibodies ◽  
Respiratory Infections ◽  
Neutralization Test ◽  
Rhesus Macaques ◽  
High Sensitivity ◽  
High Specificity ◽  
Virus Neutralization Test ◽  
Virus Neutralization ◽  
Antibody Detection ◽  
Intermediate Hosts

Assays to measure SARS-CoV-2-specific neutralizing antibodies are important to monitor seroprevalence, to study asymptomatic infections and to reveal (intermediate) hosts. A recently developed assay, the surrogate virus-neutralization test (sVNT) is a quick and commercially available alternative to the 'gold standard' virus neutralization assay using authentic virus, and does not require processing at BSL-3 level. The assay relies on the inhibition of binding of the receptor binding domain (RBD) on the spike (S) protein to human angiotensin-converting enzyme 2 (hACE2) by antibodies present in sera. As the sVNT does not require species- or isotype-specific conjugates, it can be similarly used for antibody detection in human and animal sera. In this study, we used 298 sera from PCR-confirmed COVID-19 patients and 151 sera from patients confirmed with other coronavirus or other (respiratory) infections, to evaluate the performance of the sVNT. To analyze the use of the assay in a One Health setting, we studied the presence of RBD-binding antibodies in 154 sera from nine animal species (cynomolgus and rhesus macaques, ferrets, rabbits, hamsters, cats, cattle, mink and dromedary camels). The sVNT showed a moderate to high sensitivity and a high specificity using sera from confirmed COVID-19 patients (91.3% and 100%, respectively) and animal sera (93.9% and 100%), however it lacked sensitivity to detect low titers. Significant correlations were found between the sVNT outcomes and PRNT50 and the Wantai total Ig and IgM ELISAs. While species-specific validation will be essential, our results show that the sVNT holds promise in detecting RBD-binding antibodies in multiple species.

scholarly journals The Dynamics of Neutralizing Antibody Level in One Year After SARS-CoV-2 Infection

2021 ◽  
Author(s):  
Dan Liang ◽  
Guanting Zhang ◽  
Mingxing Huang ◽  
Wenshan Hong ◽  
An'an Li ◽  
...  
Keyword(s):  
Severe Acute Respiratory Syndrome ◽  
Antibody Level ◽  
Diagnostic Tests ◽  
Neutralizing Antibodies ◽  
Neutralizing Antibody ◽  
Neutralization Assay ◽  
Blood Samples ◽  
Immune Ability ◽  
One Year

Abstract Background: A lot of recent researches have focused on the duration of the nature immunity elicited by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. An improved understanding of the immunity offered by the antibodies developed against SARS-CoV-2 in recovered patients is critical for the development of diagnostic tests and vaccines. Methods: We enrolled 114 donors, which providing blood samples after discharge for half a year and one year. Neutralizing antibodies (NAbs) were tested using a micro-neutralization assay. Results: In two tests, 82 of 114 recovered patients completed the first test half a year after discharge and NAbs remained detectable in the vast majority of patients (75/82, 91.46%). In the comparison of the two intervals, 50% (27/54) of individuals had increased NAbs titers. when 31.48% (17/54) of patients remained unchanged. Conclusion: Our results suggest that immune ability is acquired in most individuals infected with SARS-CoV-2 and is sustained in a majority of patients for up to a year after recovery.

scholarly journals Evaluation of Commercial Anti-SARS-CoV-2 Neutralizing Antibody Assays in seropositive subjects

2022 ◽  
Author(s):  
Kahina Saker ◽  
Bruno Pozzetto ◽  
Vanessa ESCURET ◽  
Virginie Pitiot ◽  
Amélie Massardier-Pilonchéry ◽  
...  
Keyword(s):  
Functional Ability ◽  
Neutralizing Antibodies ◽  
Neutralization Test ◽  
Neutralizing Antibody ◽  
Added Value ◽  
P Value ◽  
Antibody Assays ◽  
Qualitative Test ◽  
User Friendly ◽  
Commercial User

The virus neutralization test (VNT) is the reference for the assessment of the functional ability of neutralizing antibodies (NAb) to block SARS-CoV-2 entry into cells. New competitive immunoassays measuring antibodies preventing interaction between the spike protein and its cellular receptor are proposed as surrogate VNT (sVNT). We tested three commercial sVNT (a qualitative immunochromatographic test and two quantitative immunoassays named YHLO and TECO) together with a conventional anti-spike IgG assay (bioMerieux) in comparison with an in-house plaque reduction neutralization test (PRNT50) using the original 19A strain and different variants of concern (VOC), on a panel of 306 sera from naturally-infected or vaccinated patients. The qualitative test was rapidly discarded because of poor sensitivity and specificity. Areas under the curve of YHLO and TECO assays were, respectively, 85.83 and 84.07 (p-value >0.05) using a positivity threshold of 20 for PRNT50, and 95.63 and 90.35 (p-value =0.02) using a threshold of 80. However, the performances of YHLO and bioMerieux were very close for both thresholds, demonstrating the absence of added value of sVNT compared to a conventional assay for the evaluation of the presence of NAb in seropositive subjects. In addition, the PRNT50 assay showed a reduction of NAb titers towards different VOC in comparison to the 19A strain that could not be appreciated by the commercial tests. Despite the good correlation between the anti-spike antibody titer and the titer of NAb by PRNT50, our results highlight the difficulty to distinguish true NAb among the anti-RBD antibodies with commercial user-friendly immunoassays.

scholarly journals Viral Shedding among Re-Positive Severe Acute Respiratory Syndrome Coronavirus-2 Positive Individuals in Republic of Korea

Viruses ◽  
2021 ◽  
Vol 13 (10) ◽  
pp. 2089
Author(s):  
Jeong-Min Kim ◽  
Boyeong Ryu ◽  
Young June Choe ◽  
Hye-Jun Jo ◽  
Hyeokjin Lee ◽  
...  
Keyword(s):  
Severe Acute Respiratory Syndrome ◽  
Neutralizing Antibodies ◽  
Index Patient ◽  
Test Results ◽  
Serological Tests ◽  
Past Infection ◽  
Viral Shedding ◽  
Viral Culture ◽  
Polymerase Chain ◽  
Pcr Test

This study investigated the infectivity of severe acute respiratory syndrome (SARS-CoV-2) in individuals who re-tested positive for SARS-CoV-2 RNA after recovering from their primary illness. We investigated 295 individuals with re-positive SARS-CoV-2 polymerase chain reaction (PCR) test results and 836 of their close contacts. We attempted virus isolation in individuals with re-positive SARS-CoV-2 PCR test results using cell culture and confirmed the presence of neutralizing antibodies using serological tests. Viral culture was negative in all 108 individuals with re-positive SARS-CoV-2 PCR test results in whom viral culture was performed. Three new cases of SARS-CoV-2 infection were identified among household contacts using PCR. Two of the three new cases had had contact with the index patient during their primary illness, and all three had antibody evidence of past infection. Thus, there was no laboratory evidence of viral shedding and no epidemiological evidence of transmission among individuals with re-positive SARS-CoV-2 PCR test results.

Egg Yolk Antibodies for Detection and Neutralization of Clostridium botulinum Type A Neurotoxin

2009 ◽  
Vol 72 (5) ◽  
pp. 1005-1011 ◽  
Author(s):  
D. L. TROTT ◽  
M. YANG ◽  
J. GONZALEZ ◽  
A. E. LARSON ◽  
W. H. TEPP ◽  
...  
Keyword(s):  
Specific Antibody ◽  
Neutralizing Antibodies ◽  
Egg Production ◽  
Egg Yolk ◽  
Type A ◽  
Neutralization Assay ◽  
Antibody Detection ◽  
Enzyme Linked Immunosorbent Assay ◽  
Botulinum Type ◽  
Lethal Doses

The objective of this research project was to determine the usefulness of an egg antibody platform for producing materials for the detection and neutralization of botulinum type A neurotoxin. Yield estimates for detection and neutralizing antibodies produced using methods described were calculated. Antibody specific to botulinum toxoid A (aToxoid) and toxin A (aBoNT/A) was produced by immunizing hens with botulinum toxoid A (toxoid) followed by increasing amounts of botulinum neurotoxin A (BoNT/A) in Freund incomplete adjuvant. Egg yolks were extracted with polyethylene glycol (PEG) for antibody detection and neutralization experiments. A model aToxoid/toxoid immunoassay using only egg yolk antibody was developed and had a detection limit of 1 pg/ml of toxoid. In an indirect enzyme-linked immunosorbent assay of BoNT/A-specific antibody, the aBoNT/A contained more BoNT/A-specific antibody than did the aToxoid, and aBoNT/A was as effective as commercial rabbit antibody. The aToxoid provided no protection against BoNT/A in a standard mouse neutralization assay; however, 1 mg of PEG-extracted aBoNT/A neutralized 4,000 lethal doses of BoNT/A injected intraperitoneally. Based on these results, we calculated that in 1 month one hen could produce more than 100 liters of antibody detection reagents or enough antibody to neutralize approximately 11.6 million mouse lethal doses of botulinum toxin. Utilization of an egg antibody platform is potentially rapid (28 to 70 days) and scalable to kilogram quantities using current egg production facilities with as few as 1,000 hens.

scholarly journals Neutralizing Antibody Responses to Severe Acute Respiratory Syndrome Coronavirus 2 in Coronavirus Disease 2019 Inpatients and Convalescent Patients

2020 ◽  
Vol 71 (10) ◽  
pp. 2688-2694 ◽  
Author(s):  
Xiaoli Wang ◽  
Xianghua Guo ◽  
Qianqian Xin ◽  
Yang Pan ◽  
Yaling Hu ◽  
...  
Keyword(s):  
Severe Acute Respiratory Syndrome ◽  
Antibody Level ◽  
Neutralizing Antibodies ◽  
Clinical Characteristics ◽  
Early Stage ◽  
Geometric Mean ◽  
Neutralizing Antibody ◽  
Estimating Equation ◽  
Neutralization Assay ◽  
Enzyme Linked Immunosorbent Assay

Abstract Background Coronavirus disease 2019 (COVID-19) is a pandemic with no specific antiviral treatments or vaccines. There is an urgent need for exploring the neutralizing antibodies from patients with different clinical characteristics. Methods A total of 117 blood samples were collected from 70 COVID-19 inpatients and convalescent patients. Antibodies were determined with a modified cytopathogenic neutralization assay (NA) based on live severe acute respiratory syndrome coronavirus 2 and enzyme-linked immunosorbent assay (ELISA). The dynamics of neutralizing antibody levels at different time points with different clinical characteristics were analyzed. Results The seropositivity rate reached up to 100.0% within 20 days since onset, and remained 100.0% till days 41–53. The total geometric mean titer was 1:163.7 (95% confidence interval [CI], 128.5–208.6) by NA and 1:12 441.7 (95% CI, 9754.5–15 869.2) by ELISA. The antibody level by NA and ELISA peaked on days 31–40 since onset, and then decreased slightly. In multivariate generalized estimating equation analysis, patients aged 31–45, 46–60, and 61–84 years had a higher neutralizing antibody level than those aged 16–30 years (β = 1.0470, P = .0125; β = 1.0613, P = .0307; β = 1.3713, P = .0020). Patients with a worse clinical classification had a higher neutralizing antibody titer (β = 0.4639, P = .0227). Conclusions The neutralizing antibodies were detected even at the early stage of disease, and a significant response was shown in convalescent patients.

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