scholarly journals From Multiplex Serology to Serolomics—A Novel Approach to the Antibody Response against the SARS-CoV-2 Proteome

Viruses ◽  
2021 ◽  
Vol 13 (5) ◽  
pp. 749
Author(s):  
Julia Butt ◽  
Rajagopal Murugan ◽  
Theresa Hippchen ◽  
Sylvia Olberg ◽  
Monique van Straaten ◽  
...  
Keyword(s):  
Antibody Response ◽  
High Throughput ◽  
Large Population ◽  
High Sensitivity ◽  
Population Based ◽  
Antibody Responses ◽  
Hek293 Cells ◽  
Novel Approach ◽  
Assay Performance ◽  
Multiplex Serology

The emerging SARS-CoV-2 pandemic entails an urgent need for specific and sensitive high-throughput serological assays to assess SARS-CoV-2 epidemiology. We, therefore, aimed at developing a fluorescent-bead based SARS-CoV-2 multiplex serology assay for detection of antibody responses to the SARS-CoV-2 proteome. Proteins of the SARS-CoV-2 proteome and protein N of SARS-CoV-1 and common cold Coronaviruses (ccCoVs) were recombinantly expressed in E. coli or HEK293 cells. Assay performance was assessed in a COVID-19 case cohort (n = 48 hospitalized patients from Heidelberg) as well as n = 85 age- and sex-matched pre-pandemic controls from the ESTHER study. Assay validation included comparison with home-made immunofluorescence and commercial enzyme-linked immunosorbent (ELISA) assays. A sensitivity of 100% (95% CI: 86–100%) was achieved in COVID-19 patients 14 days post symptom onset with dual sero-positivity to SARS-CoV-2 N and the receptor-binding domain of the spike protein. The specificity obtained with this algorithm was 100% (95% CI: 96–100%). Antibody responses to ccCoVs N were abundantly high and did not correlate with those to SARS-CoV-2 N. Inclusion of additional SARS-CoV-2 proteins as well as separate assessment of immunoglobulin (Ig) classes M, A, and G allowed for explorative analyses regarding disease progression and course of antibody response. This newly developed SARS-CoV-2 multiplex serology assay achieved high sensitivity and specificity to determine SARS-CoV-2 sero-positivity. Its high throughput ability allows epidemiologic SARS-CoV-2 research in large population-based studies. Inclusion of additional pathogens into the panel as well as separate assessment of Ig isotypes will furthermore allow addressing research questions beyond SARS-CoV-2 sero-prevalence.

scholarly journals From multiplex serology to serolomics – a novel approach to the antibody response against the SARS-CoV-2 proteome

2020 ◽  
Author(s):  
Julia Butt ◽  
Rajagopal Murugan ◽  
Theresa Hippchen ◽  
Sylvia Olberg ◽  
Monique van Straaten ◽  
...  
Keyword(s):  
Antibody Response ◽  
High Throughput ◽  
Large Population ◽  
High Sensitivity ◽  
Population Based ◽  
Antibody Responses ◽  
Hek293 Cells ◽  
Novel Approach ◽  
Assay Performance ◽  
Multiplex Serology

ABSTRACTBackgroundThe emerging SARS-CoV-2 pandemic entails an urgent need for specific and sensitive high-throughput serological assays to assess SARS-CoV-2 epidemiology. We therefore aimed at developing a fluorescent-bead based SARS-CoV-2 multiplex serology assay for detection of antibody responses to the SARS-CoV-2 proteome.MethodsProteins of the SARS-CoV-2 proteome and protein N of SARS-CoV-1 and common cold Coronaviruses (ccCoVs) were recombinantly expressed in E. coli or HEK293 cells. Assay performance was assessed in a Covid-19 case cohort (n=48 hospitalized patients from Heidelberg) as well as n=85 age- and sex-matched pre-pandemic controls from the ESTHER study. Assay validation included comparison with home-made immunofluorescence and commercial Enzyme-linked immunosorbent (ELISA) assays.ResultsA sensitivity of 100% (95% CI: 86%-100%) was achieved in Covid-19 patients 14 days post symptom onset with dual sero-positivity to SARS-CoV-2 N and the receptor-binding domain of the spike protein. The specificity obtained with this algorithm was 100% (95% CI: 96%-100%). Antibody responses to ccCoVs N were abundantly high and did not correlate with those to SARS-CoV-2 N. Inclusion of additional SARS-CoV-2 proteins as well as separate assessment of immunoglobulin (Ig) classes M, A, and G allowed for explorative analyses regarding disease progression and course of antibody response.ConclusionThis newly developed SARS-CoV-2 multiplex serology assay achieved high sensitivity and specificity to determine SARS-CoV-2 sero-positivity. Its high throughput ability allows epidemiologic SARS-CoV-2 research in large population-based studies. Inclusion of additional pathogens into the panel as well as separate assessment of Ig isotypes will furthermore allow addressing research questions beyond SARS-CoV-2 sero-prevalence.

scholarly journals Evaluation of commercially available high-throughput SARS-CoV-2 serological assays for serosurveillance and related applications

2021 ◽  
Author(s):  
Mars Stone ◽  
Eduard Grebe ◽  
Hasan Sulaeman ◽  
Clara Di Germanio ◽  
Honey Dave ◽  
...  
Keyword(s):  
High Throughput ◽  
High Sensitivity ◽  
Antibody Responses ◽  
Antibody Detection ◽  
Laboratory Evaluation ◽  
Assay Performance ◽  
Testing Algorithm ◽  
Sensitivity Specificity ◽  
Intended Use

SARS-CoV-2 serosurveys can estimate cumulative incidence for monitoring epidemics but require characterization of employed serological assays performance to inform testing algorithm development and interpretation of results. We conducted a multi-laboratory evaluation of 21 commercial high-throughput SARS-CoV-2 serological assays using blinded panels of 1,000 highly-characterized blood-donor specimens. Assays demonstrated a range of sensitivities (96%-63%), specificities (99%-96%) and precision (IIC 0.55-0.99). Durability of antibody detection in longitudinal samples was dependent on assay format and immunoglobulin target, with anti-spike, direct, or total Ig assays demonstrating more stable, or increasing reactivity over time than anti-nucleocapsid, indirect, or IgG assays. Assays with high sensitivity, specificity and durable antibody detection are ideal for serosurveillance. Less sensitive assays demonstrating waning reactivity are appropriate for other applications, including characterizing antibody responses after infection and vaccination, and detection of anamnestic boosting by reinfections and vaccine breakthrough infections. Assay performance must be evaluated in the context of the intended use.

scholarly journals A large population-based association study between HLA and KIR genotypes and measles vaccine antibody responses

PLoS ONE ◽  
2017 ◽  
Vol 12 (2) ◽  
pp. e0171261 ◽  
Author(s):  
Inna G. Ovsyannikova ◽  
Daniel J. Schaid ◽  
Beth R. Larrabee ◽  
Iana H. Haralambieva ◽  
Richard B. Kennedy ◽  
...  
Keyword(s):  
Association Study ◽  
Large Population ◽  
Population Based ◽  
Antibody Responses ◽  
Measles Vaccine ◽  
Kir Genotypes

scholarly journals Optimisation and standardisation of a multiplex immunoassay of diverse Plasmodium falciparum antigens to assess changes in malaria transmission using sero-epidemiology

2019 ◽  
Vol 4 ◽  
pp. 26 ◽  
Author(s):  
Lindsey Wu ◽  
Tom Hall ◽  
Isaac Ssewanyana ◽  
Tate Oulton ◽  
Catriona Patterson ◽  
...  
Keyword(s):  
Quality Control ◽  
Plasmodium Falciparum ◽  
Serum Sample ◽  
High Throughput ◽  
High Sensitivity ◽  
Positive Control ◽  
Antibody Responses ◽  
Suspension Array ◽  
Epidemiological Surveys ◽  
Positive Controls

Background: Antibody responses have been used to characterise transmission and exposure history in malaria-endemic settings for over a decade. Such studies have typically been conducted on well-standardised enzyme-linked immunosorbent assays (ELISAs). However, recently developed quantitative suspension array technologies (qSAT) are now capable of high-throughput and multiplexed screening of up to hundreds of analytes at a time. This study presents a customised protocol for the Luminex MAGPIX© qSAT using a diverse set of malaria antigens. The aim is to develop a standardised assay for routine serological surveillance that is implementable across laboratories and epidemiological settings. Methods: A panel of eight Plasmodium falciparum recombinant antigens, associated with long- and short-lived antibody responses, was designed for the Luminex MAGPIX© platform. The assay was optimised for key steps in the protocol: antigen-bead coupling concentration, buffer composition, serum sample dilution, and bead storage conditions. Quality control procedures and data normalisation methods were developed to address high-throughput assay processing.  Antigen-specific limits of quantification (LOQs) were also estimated using both in-house and WHO reference serum as positive controls. Results: Antigen-specific bead coupling was optimised across five serum dilutions and two positive controls, resulting in concentrations operational within stable analytical ranges. Coupled beads were stable after storage at room temperature (22⁰C) for up to eight weeks. High sensitivity and specificity for distinguishing positive and negative controls at serum sample dilutions of 1:500 (AUC 0.94 95%CI 0.91-0.96) and 1:1000 (AUC 0.96 95%CI 0.94-0.98) were observed. LOQs were also successfully estimated for all analytes but varied by antigen and positive control. Conclusions: This study demonstrates that developing a standardised malaria-specific qSAT protocol for a diverse set of antigens is achievable, though further optimisations may be required. Quality control and data standardisation methods may also be useful for future analysis of large sero-epidemiological surveys.

scholarly journals Antibody Responses to SARS-CoV-2 Infection—Comparative Determination of Seroprevalence in Two High-Throughput Assays versus a Sensitive Spike Protein ELISA

Vaccines ◽  
2021 ◽  
Vol 9 (11) ◽  
pp. 1310
Author(s):  
Dinesh Mohanraj ◽  
Kelly Bicknell ◽  
Malini Bhole ◽  
Caroline Webber ◽  
Lorna Taylor ◽  
...  
Keyword(s):  
High Throughput ◽  
False Negative ◽  
Roc Curves ◽  
Antibody Responses ◽  
Lower Sensitivity ◽  
Assay Performance ◽  
Increased Sensitivity ◽  
Comparative Determination ◽  
Abbott Architect

Robust assay development for SARS-CoV-2 serological testing requires assessment of asymptomatic and non-hospitalised individuals to determine if assays are sensitive to mild antibody responses. Our study evaluated the performance characteristics of two high-throughput SARS-CoV-2 IgG nucleocapsid assays (Abbott Architect and Roche) and The Binding Site (TBS) Anti-Spike IgG/A/M ELISA kit in samples from healthcare workers (HCWs). The 252 samples were collected from multi-site NHS trusts and analysed for SARS-CoV-2 serology. Assay performance was evaluated between these three platforms and ROC curves were used to redefine the Abbott threshold. Concordance between Abbott and TBS was 66%. Any discrepant results were analysed using Roche, which showed 100% concordance with TBS. Analysis conducted in HCWs within 58 days post-PCR result demonstrated 100% sensitivity for both Abbott and Roche. Longitudinal analysis for >100 days post-PCR led to sensitivity of 77.2% and 100% for Abbott and Roche, respectively. A redefined Abbott threshold (0.64) increased sensitivity to 90%, producing results comparable to TBS and Roche. The manufacturer’s threshold set by Abbott contributes to lower sensitivity and elevated false-negative occurrences. Abbott performance improved upon re-optimisation of the cut-off threshold. Our findings provided evidence that TBS can be used as bespoke alternative for SARS-CoV-2 serology analysis where high-throughput platforms are not feasible on site.

scholarly journals MARS: a tool for haplotype-resolved population-based structural variation detection

2021 ◽  
Author(s):  
Lu Zhang ◽  
Arend Sidow ◽  
Xin Zhou
Keyword(s):  
Structural Variation ◽  
Large Population ◽  
High Sensitivity ◽  
Population Based ◽  
Multiple Alignment ◽  
Structural Variants ◽  
Validation Rate ◽  
Global View ◽  
Genome Wide ◽  
Genome Assemblies

Motivation: Linked-reads enables genome-wide phased diploid assemblies. These haplotype-resolved assemblies allow us to genotype structural variants (SVs) with a high sensitivity and be able to further phase them. Yet, existing SV callers are designed for haploid genome assemblies only, and there is no tool to call SV from a large population of diploid assemblies which can define and refine SVs from a global view. Results: We introduce MARS (Multiple Alignment-based Refinement of Svs) in linked-reads for the detection of the most common SV types - indels from diploid genome assemblies of a large population. We evaluated SVs from MARS based on Mendelian law of inheritance and PacBio HiFi reads and it achieved a high validation rate around 73%-87% for indels that we have selected from 34 assembled samples.

scholarly journals Changes in SARS-CoV-2 Spike versus Nucleoprotein Antibody Responses Impact the Estimates of Infections in Population-Based Seroprevalence Studies

2020 ◽  
Author(s):  
Craig Fenwick ◽  
Antony Croxatto ◽  
Alix T. Coste ◽  
Florence Pojer ◽  
Cyril André ◽  
...  
Keyword(s):  
General Population ◽  
Antibody Response ◽  
Specific Antibody ◽  
Post Infection ◽  
Acute Infection ◽  
Population Based ◽  
Antibody Responses ◽  
S Protein ◽  
N Protein ◽  
Protein Assays

SARS-CoV-2-specific antibody responses to the Spike (S) protein monomer, S protein native trimeric form or the nucleocapsid (N) proteins were evaluated in cohorts of individuals with acute infection (n=93) and in individuals enrolled in a post-infection seroprevalence population study (n=578) in Switzerland. Commercial assays specific for the S1 monomer, for the N protein and a newly developed Luminex assay using the S protein trimer were found to be equally sensitive in antibody detection in the acute infection phase samples. Interestingly, as compared to anti-S antibody responses, those against the N protein appear to wane in the post-infection cohort. Seroprevalence in a ‘positive patient contacts’ group (n=177) was underestimated by N protein assays by 10.9 to 32.2% and the ‘random selected’ general population group (n=311) was reduced up to 45% reduction relative to S protein assays. The overall reduction in seroprevalence targeting only anti-N antibodies for the total cohort ranged from 9.4 to 31%. Of note, the use of the S protein in its native trimer form was significantly more sensitive as compared to monomeric S proteins. These results indicate that the assessment of anti-S IgG antibody responses against the native trimeric S protein should be implemented to estimate SARS-CoV-2 infections in population-based seroprevalence studies. IMPORTANCE In the present study, we have determined SARS-CoV-2-specific antibody responses in sera of acute and post-infection phase subjects. Our results indicate that antibody responses against viral S and N proteins were equally sensitive in the acute phase of infection but that responses against N appear to wane in the post-infection phase while those against S protein persist over time. The most sensitive serological assay in both acute and post-infection phases used the native S protein trimer as binding antigen that has significantly greater conformational epitopes for antibody binding compared to the S1 monomer protein used in other assays. We believe that these results are extremely important in order to generate correct estimates of SARS-CoV-2 infections in the general population. Furthermore, the assessment of antibody responses against the trimeric S protein will be critical to evaluate the durability of the antibody response and for the characterization of a vaccine-induced antibody response.

scholarly journals Optimisation and standardisation of a multiplex immunoassay of diverse Plasmodium falciparum antigens to assess changes in malaria transmission using sero-epidemiology

2020 ◽  
Vol 4 ◽  
pp. 26 ◽  
Author(s):  
Lindsey Wu ◽  
Tom Hall ◽  
Isaac Ssewanyana ◽  
Tate Oulton ◽  
Catriona Patterson ◽  
...  
Keyword(s):  
Quality Control ◽  
Plasmodium Falciparum ◽  
Serum Sample ◽  
High Throughput ◽  
High Sensitivity ◽  
Positive Control ◽  
Antibody Responses ◽  
Suspension Array ◽  
Epidemiological Surveys ◽  
Positive Controls

Background: Antibody responses have been used to characterise transmission and exposure history in malaria-endemic settings for over a decade. Such studies have typically been conducted on well-standardised enzyme-linked immunosorbent assays (ELISAs). However, recently developed quantitative suspension array technologies (qSAT) are now capable of high-throughput and multiplexed screening of up to hundreds of analytes at a time. This study presents a customised protocol for the Luminex MAGPIX© qSAT using a diverse set of malaria antigens. The aim is to develop a standardised assay for routine serological surveillance that is implementable across laboratories and epidemiological settings. Methods: A panel of eight Plasmodium falciparum recombinant antigens, associated with long- and short-lived antibody responses, was designed for the Luminex MAGPIX© platform. The assay was optimised for key steps in the protocol: antigen-bead coupling concentration, buffer composition, serum sample dilution, and bead storage conditions. Quality control procedures and data normalisation methods were developed to address high-throughput assay processing.  Antigen-specific limits of quantification (LOQs) were also estimated using both in-house and WHO reference serum as positive controls. Results: Antigen-specific bead coupling was optimised across five serum dilutions and two positive controls, resulting in concentrations operational within stable analytical ranges. Coupled beads were stable after storage at room temperature (22⁰C) for up to eight weeks. High sensitivity and specificity for distinguishing positive and negative controls at serum sample dilutions of 1:500 (AUC 0.94 95%CI 0.91-0.96) and 1:1000 (AUC 0.96 95%CI 0.94-0.98) were observed. LOQs were also successfully estimated for all analytes but varied by antigen and positive control. Conclusions: This study demonstrates that developing a standardised malaria-specific qSAT protocol for a diverse set of antigens is achievable, though further optimisations may be required. Quality control and data standardisation methods may also be useful for future analysis of large sero-epidemiological surveys.

scholarly journals Role of coronary artery calcium score in risk prediction and therapy guidance of asymptomatic individuals

2020 ◽  
Vol 50 (5) ◽  
pp. 324-329
Author(s):  
Judit Simon ◽  
Lili Száraz ◽  
Béla Merkely ◽  
Pál Maurovich-Horvat
Keyword(s):  
Cardiovascular Risk ◽  
Coronary Artery ◽  
Risk Prediction ◽  
Imaging Modality ◽  
Large Population ◽  
Calcium Score ◽  
High Sensitivity ◽  
Population Based ◽  
Risk Reclassification

Coronary CT angiography (CCTA) has emerged as a gatekeeper to rule out coronary artery disease (CAD), due to its high sensitivity and negative predictive value. Prior to CCTA a native calcium screening scan is acquired, which provides additional information about the coronary artery anatomy and cardiovascular risk prediction by measuring coronary artery calcification (CAC). Based on large population-based and cohort studies, zero CAC score is linked to low probability of cardiovascular events in the future. Moreover, zero CAC score is superior in the discrimination and risk reclassification when compared with other cardiovascular risk factors. CAC score can also help to identify those who are less likely to benefit from statin pharmacotherapy. Since CAC score has an important role in risk stratification and it is a cheap and widely accessible non-invasive imaging modality, the major guidelines have already incorporated CAC score for risk prediction and therapy guiding. However, these guidelines give slightly different recommendations. Therefore, this review aimed to introduce the CAC measurement and to summarize the prognostic role of CAC scoring in individualized risk prediction and guiding preventative therapies.

Sign in / Sign up

Export Citation Format

Share Document