scholarly journals Correlation of SARS-CoV-2 Neutralizing Antibodies to an Automated Chemiluminescent Serological Immunoassay

2020 ◽  
Author(s):  
David G Grenache ◽  
Chunyan Ye ◽  
Steven B Bradfute
Keyword(s):  
Neutralizing Antibodies ◽  
Large Scale ◽  
Antibody Test ◽  
Negative Control ◽  
Standard Test ◽  
Serum Samples ◽  
Serological Tests ◽  
Signal Value ◽  
Positive Antibody ◽  
Chemiluminescent Immunoassay

Abstract Introduction Neutralizing antibodies (NAbs) are capable of binding to a virus to render it incapable of infection. The ability of commercially available SARS-CoV-2 serological tests to detect NAbs has not been widely reported. We sought to correlate the antibodies detected by an automated chemiluminescent immunoassay with NAbs. Methods Residual serum samples from 35 patients that had a positive antibody test using the LIAISON® SARS-CoV-2 S1/S2 IgG chemiluminescent immunoassay and 2 antibody-negative control sera were tested for NAbs using a plaque reduction neutralization test (PRNT). Results NAbs were detected in 66% (23/35) of the antibody-positive samples. The immunoassay signal value ranged from 21.7 to 131.3 AU/mL (median, 90.5) with significant correlation between it and the PRNT (r = 0.61, P = 0.002). In the samples without NAbs, the immunoassay signal ranged from 16.3 to 66.2 AU/mL (median, 27.2). An immunoassay signal cutoff of >41 AU/mL was 91% sensitive and 92% specific for the detection of NAbs. Discussion It is important that correlates of immunity to SARS-CoV-2 be identified and NAbs are considered to be central indicators of such. PRNT is the gold-standard test for identifying NAbs but it cannot be used for large-scale testing of populations. It is necessary to establish relationships between it and widely used commercial serological assays for SARS-CoV-2.

scholarly journals Comparative analysis of three point-of-care lateral flow immunoassays for detection of anti-SARS-CoV-2 antibodies in COVID-19 confirmed healthcare workers

2020 ◽  
Author(s):  
Danielle Dias Conte ◽  
Joseane Mayara Almeida Carvalho ◽  
Luciano Kleber de Souza Luna ◽  
Klinger Soares Faíco-Filho ◽  
Ana Helena Perosa ◽  
...  
Keyword(s):  
Healthcare Workers ◽  
Large Scale ◽  
Point Of Care ◽  
Antibody Test ◽  
Cost Effective ◽  
Lateral Flow ◽  
Rt Pcr ◽  
Serum Samples ◽  
Igg Antibody ◽  
Serological Tests

AbstractSince the Coronavirus Disease 2019 (COVID-19) pandemic, Brazil has the third-highest number of confirmed cases and the second-highest number of recovered patients. SARS-CoV-2 detection by real-time RT-PCR is the gold standard in certified infrastructured laboratories. However, for large-scale testing, diagnostics should be fast, cost-effective, widely available, and deployed for the community, such as serological tests based on lateral flow immunoassay (LFIA) for IgM/IgG detection. We evaluated three different commercial point-of-care (POC) LFIAs for anti-SARS-CoV-2 IgM and IgG detection in capillary whole blood of 100 healthcare workers (HCW) previously tested by RT-PCR: 1) COVID-19 IgG/IgM BIO (Bioclin, Brazil), 2) Diagnostic kit for IgM/IgG Antibody to Coronavirus (SARS-CoV-2) (Livzon, China); and 3) SARS-CoV-2 Antibody Test (Wondfo, China). A total of 84 positives and 16 negatives HCW were tested. The data was also analyzed by the number of days post symptoms (DPS) in three groups: <30 (n=26), 30-59 (n=42), and >59 (n=16). Overall detection was 85.71%, 47.62%, and 44.05% for Bioclin, Livzon, and Wondfo, respectively, with a specificity of 100%, and 98.75% for Livzon on storage serum samples. Bioclin was more sensitive (p<0.01), regardless of the DPS. Thus, the Bioclin can be used as a POC test to monitor SARS-CoV-2 seroconversion in HCW.

scholarly journals Monoclonal Antibody-Based Competitive Enzyme-Linked Immunosorbent Assay for Detecting and Quantifying West Nile Virus-Neutralizing Antibodies in Horse Sera

2006 ◽  
Vol 14 (2) ◽  
pp. 134-138 ◽  
Author(s):  
Kang-Seuk Choi ◽  
Young-Joon Ko ◽  
Jin-Ju Nah ◽  
Yong-Joo Kim ◽  
Shien-Young Kang ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
West Nile Virus ◽  
Immunosorbent Assay ◽  
Neutralizing Antibodies ◽  
Large Scale ◽  
Good Alternative ◽  
Standard Test ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serum Samples ◽  
West Nile

ABSTRACT A rapid immunoassay for detecting and quantifying West Nile virus (WNV)-neutralizing antibodies in sera was developed as an alternative to the plaque reduction neutralization test (PRNT), the gold standard test for WNV. The assay is a competitive, enzyme-linked immunosorbent assay using neutralizing monoclonal antibody 5E8 (NT-ELISA). A cutoff percent inhibition (PI) value of 35% (mean PI plus 3 standard deviations), with a specificity of 99%, was established based on analysis of 246 serum samples from horses free of WNV. The NT-ELISA detected neutralizing antibodies in all sera collected 7 or 14 days postinoculation from mice (n = 11) infected with lineage I (strain NY385-99) or II (strain B956) WNV. When sera from WNV-vaccinated horses (n = 212) were tested by NT-ELISA and PRNT, the NT-ELISA gave a positive result for 96.1% (173/180) of the PRNT-positive sera and 3.1% (1/32) of the PRNT-negative sera. Discrepancies between the two tests were observed mainly with sera with low PRNT90 titers (expressed as the reciprocal of the highest dilution yielding ≥90% reduction in the number of plaques) for WNV or low PIs by NT-ELISA. The overall agreement (k value) between the two tests was 0.86. A good correlation (r 2 = 0.77) was also observed between the tests for endpoint titration of sera (n = 116). In conclusion, the newly developed NT-ELISA may be a good alternative serologic assay for detecting WNV that can be used for large-scale testing of WNV-neutralizing antibodies in multiple species.

scholarly journals Correlation of the commercial anti-SARS-CoV-2 receptor binding domain antibody test with the chemiluminescent reduction neutralizing test and possible detection of antibodies to emerging variants

2021 ◽  
Author(s):  
Yoshitomo Morinaga ◽  
Hideki Tani ◽  
Yasushi Terasaki ◽  
Satoshi Nomura ◽  
Hitoshi Kawasuji ◽  
...  
Keyword(s):  
Receptor Binding ◽  
Disease Onset ◽  
Antibody Test ◽  
Binding Domain ◽  
Receptor Binding Domain ◽  
Healthy Individuals ◽  
Wild Type ◽  
Serum Samples ◽  
Serological Tests ◽  
Convalescent Phase

Background Serological tests are beneficial for recognizing the immune response against SARS-CoV-2. To identify protective immunity, optimization of the chemiluminescent reduction neutralizing test (CRNT), using pseudotyped SARS-CoV-2, is critical. Whether commercial antibody tests are comparably accurate is unknown. Methods Serum samples collected before variants were locally found were obtained from confirmed COVID-19 patients (n = 74), confirmed non-COVID-19 individuals (n = 179), and unscreened individuals (suspected healthy individuals, n = 229). The convalescent phase was defined as the period after day 10 from disease onset. The CRNT against pseudotyped viruses displaying the wild-type spike protein and a commercially available anti-receptor binding domain (RBD) antibody test were assayed. The CRNT was also assayed, using South African (SA) and United Kingdom (UK)-derived variants. Results The CRNT (cut off value, 50% inhibition) and the anti-RBD antibody test (cut off value, 0.8 U/mL) concurred regarding symptomatic COVID-19 patients in the convalescent phase and clearly differentiated between patients and suspected healthy individuals (sensitivity; 95.8% and 100%, specificity; 99.1% and 100%, respectively). Anti-RBD antibody test results correlated with neutralizing titer (r = 0.47, 95% CI 0.20-0.68). Compared with the wild-type, CRNT reduction was observed for the SA and UK-derived variants. Of the samples with ≥100 U/mL by the anti-RBD antibody test, 77.8% and 88.9% showed ≥50% neutralization against the UK and the SA variants, respectively. Conclusion The CRNT and commercial anti-RBD antibody test effectively classified convalescent COVID-19 patients. The strong positive results using the commercial antibody test can reflect neutralizing activity against emerging variants.

scholarly journals Occurrence of Antibodies against SARS-CoV-2 in the Domestic Cat Population of Germany

Vaccines ◽  
2020 ◽  
Vol 8 (4) ◽  
pp. 772
Author(s):  
Anna Michelitsch ◽  
Donata Hoffmann ◽  
Kerstin Wernike ◽  
Martin Beer
Keyword(s):  
Neutralizing Antibodies ◽  
Large Scale ◽  
Close Contact ◽  
Companion Animals ◽  
Cross Reactivity ◽  
Human Infection ◽  
Domestic Cat ◽  
Domestic Cats ◽  
Serum Samples ◽  
Transmission Cycle

Domestic cats (Felis catus) are popular companion animals that live in close contact with their human owners. Therefore, the risk of a trans-species spreading event between domestic cats and humans is ever-present. Shortly after the emergence of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and its rapid spread around the world, the role of domestic cats in the transmission cycle was questioned. In the present study, the first large-scale survey of antibody occurrence in the domestic cat population in Germany was conducted, in order to assess the incidence of naturally occurring human to cat transmission of SARS-CoV-2. A total of 920 serum samples, which were collected from April to September of 2020, were screened by an indirect multispecies ELISA. Positive samples were verified using an indirect immunofluorescence test (iIFT) and additionally tested for neutralizing antibodies. Furthermore, serum samples were screened for antibodies against feline coronavirus (FCoV), in order to rule out cross-reactivity in the described test systems. Overall, 0.69% (6/920) of serum samples were found to be positive for antibodies against SARS-CoV-2 by ELISA and iIFT. Two of these reactive sera also displayed neutralizing antibodies. No cross-reactivity with FCoV-specific antibodies was observed. The finding of SARS-CoV-2 antibody-positive serum samples in the domestic cat population of Germany, during a period when the incidence of human infection in the country was still rather low, indicates that human-to-cat transmission of SARS-CoV-2 happens, but there is no indication of SARS-CoV-2 circulation in cats.

scholarly journals Evaluation of the performance of three serological tests for diagnosis of Leishmania infantum infection in dogs using latent class analysis

2020 ◽  
Vol 29 (4) ◽  
Author(s):  
Asier Basurco ◽  
Alda Natale ◽  
Katia Capello ◽  
Antonio Fernández ◽  
María Teresa Verde ◽  
...  
Keyword(s):  
Sensitivity And Specificity ◽  
Leishmania Infantum ◽  
Latent Class ◽  
Rapid Test ◽  
Antibody Test ◽  
Latent Class Models ◽  
Canine Leishmaniasis ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serum Samples ◽  
Serological Tests

Abstract Canine leishmaniasis (CanL) is a disease caused by Leishmania infantum. Serological methods are the most common diagnostic techniques used for the diagnosis of the CanL. The objective of our study was to estimate the sensitivity and specificity of one in-house ELISA kit (ELISA UNIZAR) and three commercially available serological tests (MEGACOR Diagnostik GmbH) including an immunochromatographic rapid test (FASTest LEISH®), an immunofluorescent antibody test (MegaFLUO LEISH®) and an enzyme-linked immunosorbent assay (MegaELISA LEISH®), using latent class models in a Bayesian analysis. Two hundred fifteen serum samples were included. The highest sensitivity was achieved for FASTest LEISH® (99.38%), ELISA UNIZAR (99.37%), MegaFLUO LEISH® (99.36%) followed by MegaELISA LEISH® (98.49%). The best specificity was obtained by FASTest LEISH® (98.43%), followed by ELISA UNIZAR (97.50%), whilst MegaFLUO LEISH® and MegaELISA LEISH® obtained the lower specificity (91.94% and 91.93%, respectively). The results of present study indicate that the immunochromatographic rapid test evaluated FASTest LEISH® show similar levels of sensitivity and specificity to the quantitative commercial tests. Among quantitative serological tests, sensitivity and specificity were similar considering ELISA or IFAT techniques.

scholarly journals Calculating rabies virus neutralizing antibodies titres by flow cytometry

2002 ◽  
Vol 44 (3) ◽  
pp. 151-154 ◽  
Author(s):  
Juliano BORDIGNON ◽  
Fabiano COMIN ◽  
Sílvia Córdoba P. FERREIRA ◽  
Graciane M. M. CAPORALE ◽  
José Hermênio Cavalcante LIMA FILHO ◽  
...  
Keyword(s):  
Cell Culture ◽  
Flow Cytometry ◽  
Rabies Virus ◽  
Neutralizing Antibodies ◽  
Neutralizing Antibody ◽  
Standard Test ◽  
Serum Samples ◽  
Reference Serum ◽  
Infection Inhibition

The determination of the rabies neutralizing antibody (VNA) response after immunization against rabies is an acceptable index of the efficacy of a vaccine and a successful treatment. Several tests have been developed in attempt to improve the assessment of VNA, from mice inoculation to cell-culture fluorescence inhibition tests. All of them, however, present special difficulties in terms of reading or accuracy. The present study describes a neutralization test performed in cell-culture appraised by flow cytometry (FC). Serial dilutions of the serum samples were mixed in vitro with rabies virus before the addition of BHK-21 cells. After 24h-incubation, cells were released by trypsin treatment, fixed and permeabilized with a p-formaldehyde solution and stained with a rabies virus nucleocapsid protein-specific antibody conjugate. The percentage of virus infection inhibition caused by specific antibodies present in the serum were evaluated in a Beckton & Dickinson FACSCalibur® flow cytometer. A correlation curve between the IU/ml content and the percentage of infective inhibition was built with a reference serum and the VNA titers of serum samples were obtained by extrapolation. Titers obtained by FC and standard test showed an effective pairing results (p < 0.01), with a correlation coefficient (r) = 0.7. These results permit to envisage the FC as a suitable technique to evaluate VNA in sera from immunized animals and likely in human serum samples. Nevertheless, new studies comparing FC to gold-standard techniques are required for determining the FC values of Sensibility and Specificity .

scholarly journals A Spike protein-based subunit SARS-CoV-2 vaccine for pets: safety, immunogenicity, and protective efficacy in juvenile cats

2021 ◽  
Author(s):  
Kairat Tabynov ◽  
Madiana Orynbassar ◽  
Leila Yelchibayeva ◽  
Nurkeldi Turebekov ◽  
Toktassyn Yerubayev ◽  
...  
Keyword(s):  
Respiratory Tract ◽  
Neutralizing Antibodies ◽  
Large Scale ◽  
Protective Efficacy ◽  
Spike Protein ◽  
Wild Type Virus ◽  
Upper Respiratory Tract ◽  
Wild Type ◽  
Serum Samples ◽  
Virus Shedding

Abstract Whereas multiple vaccine types have been developed to curb the spread of Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) among humans, there are very few vaccines being developed for animals including pets. To combat the threat of human-to-animal, animal-to-animal and animal-to-human transmission and the generation of new virus variants, we developed a subunit SARS-CoV-2 vaccine which is based on recombinant spike protein extracellular domain expressed in insect cells then formulated with appropriate adjuvants. Sixteen 8-12-week-old outbred female and male kittens (n=4/group) were randomly assigned into four treatment groups: Group 1, Antigen alone; Group 2, Sepivac SWE™ adjuvant; Group 3, aluminum hydroxide adjuvant; Group 4, PBS administered control animals. All animals were vaccinated twice at day 0 and 14, intramuscularly in a volume of 0.5 mL (Groups 1-3: 5 µg of Spike protein). On days 0 and 28 serum samples were collected to evaluate anti-spike IgG, inhibition of spike binding to angiotensin-converting enzyme 2 (ACE-2), neutralizing antibodies to Wuhan-01 SARS-CoV-2 D614G (wild-type) and Delta variant viruses, and whole blood for hematology studies. At day 28, all groups were challenged with SARS-CoV-2 wild-type virus 106 TCID50 intranasally. On day 31, tissue samples (lung, heart, and nasal turbinates) were collected for histology, viral RNA detection and virus titration. Parameters evaluated in this study included safety, immunogenicity, and protection from infection with wild-type SARS-CoV-2 virus. After two immunizations, both vaccines induced high titers of serum anti-spike IgG, ACE-2 binding inhibitory and neutralizing antibodies against both wild-type and Delta variant virus in the juvenile cats. Both subunit vaccines provided protection of juvenile cats against virus shedding from the upper respiratory tract, and against viral replication in the lower respiratory tract and hearts. These promising data warrant ongoing evaluation of the vaccine’s ability to protect cats against SARS-CoV-2 Delta variant and in particular to prevent transmission of the infection to naïve cats, before proceeding with large-scale field trials.

scholarly journals Comparative Analysis of the Serological Reactivity of Individuals with Clinical History of Malaria using Two Different ELISA Tests

Diagnostics ◽  
2019 ◽  
Vol 9 (4) ◽  
pp. 168
Author(s):  
Yorleydy Ruiz Moreno ◽  
Silvia Tavares Donato ◽  
Fátima Nogueira ◽  
Marcelo Sousa Silva
Keyword(s):  
Clinical History ◽  
Negative Control ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serological Response ◽  
Serum Samples ◽  
Serological Tests ◽  
Serological Reactivity ◽  
History Of ◽  
Low Levels ◽  
Antigenic Reactivity

Early diagnosis of malaria reduces disease, prevents deaths, and contributes to decreased malaria transmission. The use of specific and sensitive antigens in the execution of serological diagnostics may have an impact on the transmission of the disease. However, many individuals cannot be easily diagnosed by serological tests due to low levels of antibodies in the serum. Using two different Enzyme-Linked Immunosorbent Assay (ELISA) tests (a commercial and an in-house ELISA), a total of 365 serum samples from individuals with a clinical history of malaria were analyzed. From the serum samples analyzed, 192 (53%) samples from the commercial ELISA and 219 (60%) samples from the in-house ELISA presented positive serological reactivity to malaria. The concordance of the samples tested (n = 365) between both ELISAs was of 67% (n = 242), and with the negative control was 100% (n = 17). We demonstrated that the in-house ELISA showed high antigenic reactivity to Plasmodium falciparum antigens when compared with the commercial ELISA. The degree of concordance of both ELISAs suggested the possibility of existence of other P. falciparum antigens present in the crude extract of P. falciparum that are important in the serological response during malaria infection.

scholarly journals Evaluation of Eight Serological Tests for Diagnosis of Imported Schistosomiasis

2012 ◽  
Vol 19 (6) ◽  
pp. 948-953 ◽  
Author(s):  
Hans-Friedemann Kinkel ◽  
Sabine Dittrich ◽  
Britta Bäumer ◽  
Thomas Weitzel
Keyword(s):  
Antibody Test ◽  
Parasitic Infections ◽  
Serum Samples ◽  
Serological Tests ◽  
Content Type ◽  
Commercial Kits ◽  
Few Data ◽  
Indirect Immunofluorescent ◽  
Positive Results ◽  
Immunofluorescent Antibody Test

ABSTRACTThe diagnosis of schistosomiasis in individuals from countries where the disease is not endemic is challenging, and few data are available on the accuracy of serological diagnosis in those patients. We evaluated the performance of eight serological assays, including four commercial kits, in the diagnosis of imported schistosomiasis in individuals from areas where the disease is not endemic, including six enzyme-linked immunosorbent assays using three different antigens, an indirect hemagglutination assay, and an indirect immunofluorescent-antibody test. To analyze the assays, we used a total of 141 serum samples, with 121 derived from patients with various parasitic infections (among which were 37 cases of schistosomiasis) and 20 taken from healthy volunteers. The sensitivity values for detection of schistosomiasis cases ranged from 41% to 78% and were higher forSchistosoma mansonithan forS. haematobiuminfections. Specificity values ranged from 76% to 100%; false-positive results were most frequent for samples from patients with cestode infections. By combining two or more tests, sensitivity improved markedly and specificity decreased only moderately. Serological tests are useful instruments for diagnosing imported schistosomiasis in countries where the disease is not endemic, but due to limitations in test sensitivities, we recommend the use of two or more assays in parallel.

scholarly journals Structural and functional comparison of SARS-CoV-2-spike receptor binding domain produced in Pichia pastoris and mammalian cells

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Keyword(s):  
Pichia Pastoris ◽  
Receptor Binding ◽  
Mammalian Cells ◽  
Neutralizing Antibodies ◽  
Large Scale ◽  
Binding Domain ◽  
Size Exclusion ◽  
Scale Production ◽  
Receptor Binding Domain ◽  
Serological Tests

AbstractThe yeast Pichia pastoris is a cost-effective and easily scalable system for recombinant protein production. In this work we compared the conformation of the receptor binding domain (RBD) from severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) Spike protein expressed in P. pastoris and in the well established HEK-293T mammalian cell system. RBD obtained from both yeast and mammalian cells was properly folded, as indicated by UV-absorption, circular dichroism and tryptophan fluorescence. They also had similar stability, as indicated by temperature-induced unfolding (observed Tm were 50 °C and 52 °C for RBD produced in P. pastoris and HEK-293T cells, respectively). Moreover, the stability of both variants was similarly reduced when the ionic strength was increased, in agreement with a computational analysis predicting that a set of ionic interactions may stabilize RBD structure. Further characterization by high-performance liquid chromatography, size-exclusion chromatography and mass spectrometry revealed a higher heterogeneity of RBD expressed in P. pastoris relative to that produced in HEK-293T cells, which disappeared after enzymatic removal of glycans. The production of RBD in P. pastoris was scaled-up in a bioreactor, with yields above 45 mg/L of 90% pure protein, thus potentially allowing large scale immunizations to produce neutralizing antibodies, as well as the large scale production of serological tests for SARS-CoV-2.

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