scholarly journals Development and Evaluation of a Set of Spike and Receptor Binding Domain-Based Enzyme-Linked Immunosorbent Assays for SARS-CoV-2 Serological Testing

Diagnostics ◽  
2021 ◽  
Vol 11 (8) ◽  
pp. 1506
Author(s):  
Rosa Camacho-Sandoval ◽  
Alejandro Nieto-Patlán ◽  
Gregorio Carballo-Uicab ◽  
Alejandra Montes-Luna ◽  
María C. Jiménez-Martínez ◽  
...  
Keyword(s):  
Performance Test ◽  
Viral Infections ◽  
Third Party ◽  
Receptor Binding Domain ◽  
Serological Testing ◽  
Rt Pcr ◽  
Serum Samples ◽  
Sanitary Authority ◽  
Secondary Antibodies ◽  
Igg1 Isotype

The implementation and validation of anti-SARS-CoV-2 IgG serological assays are reported in this paper. S1 and RBD proteins were used to coat ELISA plates, and several secondary antibodies served as reporters. The assays were initially validated with 50 RT-PCR positive COVID-19 sera, which showed high IgG titers of mainly IgG1 isotype, followed by IgG3. Low or no IgG2 and IgG4 titers were detected. Then, the RBD/IgG assay was further validated with 887 serum samples from RT-PCR positive COVID-19 individuals collected at different times, including 7, 14, 21, and 40 days after the onset of symptoms. Most of the sera were IgG positive at day 40, with seroconversion happening after 14–21 days. A third party conducted an additional performance test of the RBD/IgG assay with 406 sera, including 149 RT-PCR positive COVID-19 samples, 229 RT-PCR negative COVID-19 individuals, and 28 sera from individuals with other viral infections not related to SARS-CoV-2. The sensitivity of the assay was 99.33%, with a specificity of 97.82%. All the sera collected from individuals with infectious diseases other than COVID-19 were negative. Given the robustness of this RBD/IgG assay, it received approval from the sanitary authority in Mexico (COFEPRIS) for production and commercialization under the name UDISTEST-V2G®.

scholarly journals Evaluation of in-house ELISA and 11 commercial ELISA kits in the serological diagnosis of COVID-19

2021 ◽  
pp. 39-52
Author(s):  
Waldemar Rastawicki ◽  
Klaudia Płaza ◽  
Adam Pietrusiński
Keyword(s):  
Viral Infections ◽  
Clinical Symptoms ◽  
High Sensitivity ◽  
Diagnostic Methods ◽  
Molecular Diagnostic ◽  
Rt Pcr ◽  
Serum Samples ◽  
Serological Tests ◽  
Igm Antibodies ◽  
Epidemiological Surveys

Introduction: ELISA-Immunoassays can complement the molecular diagnostic methods, and can be one of the important tools of sero-surveillance and vaccine evaluation. The aim of the presented study was to develop in-house ELISA and evaluate 11 commercial ELISA tests for detection of anti-SARS-CoV-2 antibodies in serum samples collected from COVID patients. Methods: In total, 237 serum samples obtained from 165 people with COVID-19 with RT-PCR confirmed SARS-CoV-2 virus infection were used for the study. The specificity of the developed in-house ELISA kit was tested using 170 serum samples obtained from patients with various bacterial and viral infections. The study used an in-house ELISA and 11 commercial ELISA kits developed by various manufacturers. Results: The presented study showed high sensitivity (81.0%) and specificity (97.2%) of the developed in-house kit in relation to the RT-PCR method. The sensitivity of the inhouse test significantly increased (98.1%) when only convalescents - persons at least 3 weeks after COVID-19 were examined. Commercial ELISA kits most frequently detected IgG antibodies (from 44.9% to 89.4%), especially in samples obtained later in the disease, and the least frequent detection of IgM antibodies (from 4.2% to 42.4%). Conclusions: All the presented ELISA kits may be used in serodiagnosis of COVID-19 however the detection of antibodies in individual tests differed quite significantly and was dependent on the period of the disease, on the class of immunoglobulins and the type of antigen used. The sensitivity of serological tests in the IgG class is clearly higher when examining samples obtained at least 2-3 weeks from the onset of clinical symptoms. Searching for IgA antibodies may be useful mainly in the early phase of the disease while IgM antibodies does not provide significant additional information. In the case of asymptomatic or mild infection, the level of antibodies is low which may be the cause problems with the correct interpretation of epidemiological surveys

scholarly journals SARS-CoV-2 IgG antibody responses in rt-PCR-positive cases: first report from India

2021 ◽  
Vol 3 (10) ◽  
Author(s):  
Girish Chandra Dash ◽  
Debaprasad Parai ◽  
Hari Ram Choudhary ◽  
Annalisha Peter ◽  
Usha Kiran Rout ◽  
...  
Keyword(s):  
Antibody Response ◽  
Clinical Utility ◽  
Antibody Responses ◽  
Serological Testing ◽  
Rt Pcr ◽  
Serum Samples ◽  
Igg Antibody ◽  
Significant Difference ◽  
The Mean ◽  
The Relationship

Introduction. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antibody responses remain poorly understood and the clinical utility of serological testing is still unclear. Aim. To understand the relationship between the antibody response to SARS-CoV-2 infection and the demographics and cycle threshold (C t) values of confirmed RT-PCR patients. Methodology. A total of 384 serum samples were collected from individuals between 4–6 weeks after confirmed SARS-CoV-2 infection and tested for the development of immunoglobulin class G (IgG) against SARS-CoV-2. The C t values, age, gender and symptoms of the patients were correlated with the development of antibodies. Results. IgG positivity was found to be 80.2 % (95 % CI, 76.2–84.2). Positivity increased with a decrease in the C t value, with the highest (87.6 %) positivity observed in individuals with C t values <20. The mean (±sd) C t values for IgG positives and negatives were 23.34 (±6.09) and 26.72 (±7.031), respectively. No significant difference was found for demographic characteristics such as age and sex and symptoms and antibody response. The current study is the first of its kind wherein we have assessed the correlation of the RT-PCR C t with the development of IgG against SARS-CoV-2. Conclusion. Although C t values might not have any relation with the development of symptoms, they are associated with the antibody response among SARS-CoV-2-infected individuals.

scholarly journals Seroprevalence analysis of SARS-CoV-2 in pregnant women along the first pandemic outbreak and perinatal outcome

PLoS ONE ◽  
2020 ◽  
Vol 15 (11) ◽  
pp. e0243029
Author(s):  
Cecilia Villalaín ◽  
Ignacio Herraiz ◽  
Joanna Luczkowiak ◽  
Alfredo Pérez-Rivilla ◽  
María Dolores Folgueira ◽  
...  
Keyword(s):  
Pregnant Women ◽  
Horizontal Transmission ◽  
Perinatal Outcomes ◽  
Study Groups ◽  
Serological Testing ◽  
Igg Antibodies ◽  
Rt Pcr ◽  
Serum Samples ◽  
Specific Igg ◽  
Specific Igg Antibodies

Objectives To evaluate the progression of the seroprevalence of SARS-CoV-2 in the pregnant population of the south of Madrid during the first wave of the COVID-19 pandemic. Secondarily we aimed to evaluate maternal and perinatal outcomes. Study design Retrospective cohort study conducted at Hospital Universitario 12 de Octubre during weeks 10 to 19 of 2020, coinciding with the Spanish lockdown. We tested 769 serum samples obtained from routine serological testing during the first and third trimesters of pregnancy for specific IgG anti SARS-CoV-2 RBD and S proteins. RT-PCR tests were performed in suspected cases according to clinical practice. We compared maternal and perinatal outcomes in those with delivered pregnancies (n = 578) according to the presence or absence of specific IgG antibodies. Those with positive IgG were subdivided by the presence or absence of Covid-19 related symptoms at any time and the results of RT-PCR testing if performed. Therefore, we had 4 study groups: G1 (IgG negative), G2 (IgG positive, asymptomatic, RT-PCR testing negative or not done), G3 (IgG positive, symptomatic, RT-PCR testing negative or not done), and G4 (IgG positive, symptomatic, RT-PCR positive). Results Seropositivity increased from 0% to 21.4% (95% CI 11.8–31.0) during the study period, of which 27.9% had an asymptomatic course. Overall outcomes were favorable with a significant increased rate of preterm birth in G4 vs G1 (21.4% vs 6.7%) and cesarean/operative delivery (50% vs 26.9%). Asymptomatic and mild cases did not have differences regarding pregnancy course when compared to seronegative women. There were no documented cases of vertical or horizontal transmission. Conclusion Seroprevalence in pregnant women in southern Madrid went up to 21.4% of which 27.9% had an asymptomatic course. Overall perinatal results were favorable, especially in those asymptomatic.

scholarly journals Clinical validation and performance evaluation of the automated Vitros Total Anti-SARS-CoV-2 Antibodies assay for screening of serostatus in COVID-19

2020 ◽  
Author(s):  
Emily Garnett ◽  
Joanna Jung ◽  
Estella Tam ◽  
Deepthi Rajapakshe ◽  
Stephen Cheney ◽  
...  
Keyword(s):  
Viral Infections ◽  
Acute Infection ◽  
Antibody Test ◽  
Cross Reactivity ◽  
Medical Professional ◽  
List Type ◽  
Serological Testing ◽  
Rt Pcr ◽  
Serological Tests ◽  
Few Data

AbstractObjectivesEvaluation of serostatus against SARS-CoV-2 has emerged as an important tool in identification of exposure to COVID-19. We report on the validation of the Vitros Anti-SARS-CoV-2 Total (CoV2T) assay for qualitative serological testing of SARS-CoV-2 antibodies.MethodsWe performed validation studies according to COLA guidelines, using samples previously tested for SARS-CoV-2 by RT-PCR. We evaluated precision, analytical interferences, and cross-reactivity with other viral infections. We also evaluated concordance with molecular and other serological testing, and evaluated seroconversion.ResultsThe Vitros CoV2T assay exhibited acceptable precision, was resistant to analytical interference, and did not exhibit cross-reactivity with samples positive for other respiratory viruses. The CoV2T assay exhibited 100% negative predictive agreement (56/56) and 71% positive predictive agreement (56/79) with RT-PCR across all patient samples, and was concordant with other serological assays. Concordance with RT-PCR was 97% > 7 days after symptom onset.ConclusionsThe Vitros CoV2T assay was successfully validated in our laboratory. We anticipate it will be a useful tool in screening for exposure to SARS-CoV-2, however, the use of the CoV2T and other serological assays in clinical management of COVID-19 patients is yet unknown, and must be evaluated in future studies.Key pointsWhat issue or core problem does the study address?Multiple serological assays for detection of anti-SARS-CoV-2 antibodies have received FDA Emergency Use Authorizations, but few data have been published on the performance of these assays.What would one take-home point for the working medical professional be?The Vitros Anti-SARS-CoV-2 Total assay is a total antibody test to be used as a serological screen for exposure to COVID-19. This assay performs well, and is comparable to other serological tests.What is the most significant or most interesting finding of the study?We confirmed that the Vitros Anti-SARS-CoV-2 Total assay, like other serological tests, is not suitable for diagnosis of acute infection, as it is not sensitive to infection <7 days post-onset.

Detection of Nucleic Acids in Cells or Tissue Sections Using In situ Polymerase Chain Reaction (PCR)

1996 ◽  
Vol 54 ◽  
pp. 16-17
Author(s):  
J. R. Hully ◽  
K. R. Luehrsen ◽  
K. Aoyagi ◽  
C. Shoemaker ◽  
R. Abramson
Keyword(s):  
Nucleic Acids ◽  
Viral Infections ◽  
Anatomical Location ◽  
Rt Pcr ◽  
Reverse Transcription Pcr ◽  
Cellular Source ◽  
Gene Transcripts ◽  
Initial Description ◽  
In Cells

The development of PCR technology has greatly accelerated medical research at the genetic and molecular levels. Until recently, the inherent sensitivity of this technique has been limited to isolated preparations of nucleic acids which lack or at best have limited morphological information. With the obvious exception of cell lines, traditional PCR or reverse transcription-PCR (RT-PCR) cannot identify the cellular source of the amplified product. In contrast, in situ hybridization (ISH) by definition, defines the anatomical location of a gene and/or it’s product. However, this technique lacks the sensitivity of PCR and cannot routinely detect less than 10 to 20 copies per cell. Consequently, the localization of rare transcripts, latent viral infections, foreign or altered genes cannot be identified by this technique. In situ PCR or in situ RT-PCR is a combination of the two techniques, exploiting the sensitivity of PCR and the anatomical definition provided by ISH. Since it’s initial description considerable advances have been made in the application of in situ PCR, improvements in protocols, and the development of hardware dedicated to in situ PCR using conventional microscope slides. Our understanding of the importance of viral latency or viral burden in regards to HIV, HPV, and KSHV infections has benefited from this technique, enabling detection of single viral copies in cells or tissue otherwise thought to be normal. Clearly, this technique will be useful tool in pathobiology especially carcinogenesis, gene therapy and manipulations, the study of rare gene transcripts, and forensics.

scholarly journals Investigations, actions and learning from an outbreak of SARS-CoV-2 infection among healthcare workers in the United Kingdom

2020 ◽  
pp. 175717742097679
Author(s):  
Kordo Saeed ◽  
Emanuela Pelosi ◽  
Nitin Mahobia ◽  
Nicola White ◽  
Christopher Labdon ◽  
...  
Keyword(s):  
Real Time ◽  
Healthcare Workers ◽  
Healthcare Facility ◽  
Rt Pcr ◽  
Serum Samples ◽  
The United Kingdom ◽  
Key Issues ◽  
Current Infection ◽  
Polymerase Chain ◽  
A Current

Background: We report an outbreak of SARS coronavirus-2 (SARS-CoV-2) infection among healthcare workers (HCW) in an NHS elective healthcare facility. Methodology: A narrative chronological account of events after declaring an outbreak of SARS-CoV-2 among HCWs. As part of the investigations, HCWs were offered testing during the outbreak. These were: (1) screening by real-time reverse transcriptase polymerase chain reaction (RT- PCR) to detect a current infection; and (2) serum samples to determine seroprevalence. Results: Over 180 HCWs were tested by real-time RT-PCR for SARS-CoV-2 infection. The rate of infection was 15.2% (23.7% for clinical or directly patient-facing HCWs vs. 4.8% in non-clinical non-patient-facing HCWs). Of the infected HCWs, 57% were asymptomatic. Seroprevalence (SARS-CoV-2 IgG) among HCWs was 13%. It was challenging to establish an exact source for the outbreak. The importance of education, training, social distancing and infection prevention practices were emphasised. Additionally, avoidance of unnecessary transfer of patients and minimising cross-site working for staff and early escalation were highlighted. Establishing mass and regular screening for HCWs are also crucial to enabling the best care for patients while maintaining the wellbeing of staff. Conclusion: To our knowledge, this is the first UK outbreak report among HCWs and we hope to have highlighted some key issues and learnings that can be considered by other NHS staff and HCWs globally when dealing with such a task in future.

scholarly journals Glycosylation is a key in SARS-CoV-2 infection

2021 ◽  
Author(s):  
Celso A. Reis ◽  
Rudolf Tauber ◽  
Véronique Blanchard
Keyword(s):  
Adaptive Immune Response ◽  
Protective Role ◽  
Host Cells ◽  
Target Cells ◽  
Receptor Binding Domain ◽  
Serological Testing ◽  
Virus Binding ◽  
Adaptive Immune ◽  
Cytokine Cascade ◽  
Different Levels

AbstractSARS-CoV-2 causes the respiratory syndrome COVID-19 and is responsible for the current pandemic. The S protein of SARS-CoV-2-mediating virus binding to target cells and subsequent viral uptake is extensively glycosylated. Here we focus on how glycosylation of both SARS-CoV-2 and target cells crucially impacts SARS-CoV-2 infection at different levels: (1) virus binding and entry to host cells, with glycosaminoglycans of host cells acting as a necessary co-factor for SARS-CoV-2 infection by interacting with the receptor-binding domain of the SARS-CoV-2 spike glycoprotein, (2) innate and adaptive immune response where glycosylation plays both a protective role and contributes to immune evasion by masking of viral polypeptide epitopes and may add to the cytokine cascade via non-fucosylated IgG, and (3) therapy and vaccination where a monoclonal antibody-neutralizing SARS-CoV-2 was shown to interact also with a distinct glycan epitope on the SARS-CoV-2 spike protein. These evidences highlight the importance of ensuring that glycans are considered when tackling this disease, particularly in the development of vaccines, therapeutic strategies and serological testing.

scholarly journals Frequency of Hepatitis B, C and HIV Infections among Transfusion-Dependent Beta Thalassemia Patients in Dhaka

2021 ◽  
Vol 13 (1) ◽  
pp. 89-95
Author(s):  
Golam Sarower Bhuyan ◽  
Aftab Uz Zaman Noor ◽  
Rosy Sultana ◽  
Farjana Akther Noor ◽  
Nusrat Sultana ◽  
...  
Keyword(s):  
Hepatitis B ◽  
Viral Infections ◽  
Rapid Test ◽  
Hiv Infections ◽  
Beta Thalassemia ◽  
Nucleic Acid Testing ◽  
Serological Testing ◽  
Elisa Method ◽  
B Virus ◽  
Immunodeficiency Virus

Transfusion transmitted infections have remained a major deterrent to public health, particularly among the patients with transfusion-dependent Beta thalassemia in developing countries. Although proper donor selection through adoption of WHO-advised infection panel has lowered the rate of infections, the multi-transfused patients are not free of risk. In this study, we screened 148 transfusion-dependent Beta thalassemia patients to determine the frequency of Hepatitis C Virus (HCV), Hepatitis B Virus (HBV) and Human Immunodeficiency Virus (HIV) using the ELISA method. Among them, infected cases with HCV, HBV and HIV were 13.51%, 3.37% and 0%, respectively. Moreover, 2% of the patients were found to be co-infected with both HBV and HCV. The percentage of infections in the patients with frequent transfusion interval (≤30 days) was significantly higher (p < 0.0005) than that in the patients with less frequent transfusion intervals (>30 days). Immunochromatography (ICT)-based rapid test kits are usually used to screen and confirm these infections in the blood of the patients. However, ICT-based tests are not sensitive enough to detect the infections. So, a combination of both Nucleic Acid testing (NAT) and serological testing are suggested to significantly reduce the risk of viral infections during blood transfusion.

scholarly journals COVID-19 serological testing for Healthcare Workers in Lombardy, Italy

2020 ◽  
Vol 30 (Supplement_5) ◽  
Author(s):  
G Del Castillo ◽  
A Castrofino ◽  
F Grosso ◽  
A Barone ◽  
L Crottogini ◽  
...  
Keyword(s):  
Healthcare Workers ◽  
Geographical Area ◽  
Healthcare Facility ◽  
Nasopharyngeal Swab ◽  
Infection Prevalence ◽  
Serological Testing ◽  
Prevention Policy ◽  
Rt Pcr ◽  
Serological Tests ◽  
Screening Process

Abstract Issue COVID-19 pandemic began in Italy on February 20th, 2020. Since the beginning of the emergency Healthcare Workers' (HCWs) involvement was prominent, mainly due to direct assistance to COVID-19 patients. Therefore, we implemented a prevention policy for HCW screening through serological and RT-PCR testing. Description of the problem HCW screening for SARS-CoV-2 infection is essential for prevention and control of the pandemic. Lombardy's Healthcare authorities settled a screening process for HCWs divided into three steps: 1) body temperature assessment at the beginning and the end of work shift, if fever &gt; 37.5 °C was present the HCW was sent back home and a nasopharyngeal swab was performed; 2) progressive recruitment for serological testing; 3) on those positive to IgG a nasopharyngeal swab was performed and tested for viral RNA by RT-PCR. Results Among 79185 HCW tested, 9589 (12%) were positive on serological IgG testing. Of the 9589 positive a nasopharyngeal swab was performed on 6884. Of these 358 (5%) tested positive and the remaining 6526 (95%) negative to RT-PCR. We calculated a Positive Predictive Value of 5.2%. The rate of positive serological tests for each Healthcare facility varied between 0% and 78%. Five percent of all facilities, belonging to Brescia, Bergamo and Cremona area, reported a positivity rate higher than 40% in HCWs. A second cluster (18% of all facilities), involving the same geographical area, reported a rate between 20% and 40%, whereas the remaining facilities (76%) of the region a rate &lt;20%. Lessons Serological IgG testing can be, if followed by immediate nasopharyngeal swab testing, a valid screening intervention on asymptomatic HCWs especially in a high infection prevalence setting. Key messages Serological IgG testing can be, if followed by immediate nasopharyngeal swab testing, a valid screening intervention on asymptomatic HCWs. Infection prevention in HCW may benefit from a screening campaign especially in high prevalence settings.

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