Development and Standardization of a High-Throughput Multiplex Immunoassay for the Simultaneous Quantification of Specific Antibodies to Five Respiratory Syncytial Virus Proteins

Schepp, Rutger M.; Haan, Cornelis A. M. de; Wilkins, Deidre; Layman, Hans; Graham, Barney S.; Esser, Mark T.; Berbers, Guy A. M.

doi:10.1128/msphere.00236-19

Cited by 23 publications

(32 citation statements)

References 27 publications

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“…RSV-specific IgA concentrations were determined in nasal samples by a multiplex immunoassay, essentially as described before (44). Fluorescent beads (Bio-Rad Laboratories) were coated with RSV pre-F, post-F, N, G A , or G B protein, and bound IgA was detected using goat F(ab')2 anti-human IgA conjugated to phycoerythrin (PE) (Southern Biotech).…”

Section: Methodsmentioning

confidence: 99%

Antibody and Local Cytokine Response to Respiratory Syncytial Virus Infection in Community-Dwelling Older Adults

Lakerveld

Imholz

et al. 2020

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Respiratory syncytial virus (RSV) is increasingly recognized for causing severe morbidity and mortality in older adults, but there are few studies on the RSV-induced immune response in this population. Information on the immunological processes at play during RSV infection in specific risk groups is essential for the rational and targeted design of novel vaccines and therapeutics. Here, we assessed the antibody and local cytokine response to RSV infection in community-dwelling older adults (≥60 years of age). During three winters, serum and nasopharyngeal swab samples were collected from study participants during acute respiratory infection and recovery. RSV IgG enzyme-linked immunosorbent assays (ELISA) and virus neutralization assays were performed on serum samples from RSV-infected individuals (n = 41) and controls (n = 563 and n = 197, respectively). Nasal RSV IgA and cytokine concentrations were determined using multiplex immunoassays in a subset of participants. An in vitro model of differentiated primary bronchial epithelial cells was used to assess RSV-induced cytokine responses over time. A statistically significant increase in serum neutralization titers and IgG concentrations was observed in RSV-infected participants compared to controls. During acute RSV infection, a statistically significant local upregulation of beta interferon (IFN-β), IFN-λ1, IFN-γ, interleukin 1β (IL-1β), tumor necrosis factor alpha (TNF-α), IL-6, IL-10, CXCL8, and CXCL10 was found. IFN-β, IFN-λ1, CXCL8, and CXCL10 were also upregulated in the epithelial model upon RSV infection. In conclusion, this study provides novel insights into the basic immune response to RSV infection in an important and understudied risk population, providing leads for future studies that are essential for the prevention and treatment of severe RSV disease in older adults. IMPORTANCE Respiratory syncytial virus (RSV) can cause severe morbidity and mortality in certain risk groups, especially infants and older adults. Currently no (prophylactic) treatment is available, except for a partially effective yet highly expensive monoclonal antibody. RSV therefore remains a major public health concern. To allow targeted development of novel vaccines and therapeutics, it is of great importance to understand the immunological mechanisms that underlie (protection from) severe disease in specific risk populations. Since most RSV-related studies focus on infants, there are only very limited data available concerning the response to RSV in the elderly population. Therefore, in this study, RSV-induced antibody responses and local cytokine secretion were assessed in community-dwelling older adults. These data provide novel insights that will benefit ongoing efforts to design safe and effective prevention and treatment strategies for RSV in an understudied risk group.

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Section: Methodsmentioning

confidence: 99%

Antibody and Local Cytokine Response to Respiratory Syncytial Virus Infection in Community-Dwelling Older Adults

Lakerveld

Imholz

et al. 2020

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“…We used a multiplex immunoassay to quantify the concentration of IgG specific for the RSV attachment (G) and fusion (F) proteins . G protein from both an A‐ and B‐strain was included, as these sequences are highly variable between the two RSV subtypes .…”

Section: Resultsmentioning

confidence: 99%

“…Also the combination of antibody binding titres and NK cell activation does not provide a clear separation between disease groups. Since pre‐F‐specific antibody titres strongly correlate with in vitro neutralisation titres, it is expected that the combination of neutralisation and NK cell activation will reveal a similar pattern.…”

Section: Discussionmentioning

confidence: 99%

“…We used a multiplex immunoassay to quantify the concentration of IgG specific for the RSV attachment (G) and fusion (F) proteins. 31 G protein from both an A-and B-strain was included, as these sequences are highly variable between the two RSV subtypes. 32 Both the preand post-fusion conformations of the F protein were included, as these harbour partly overlapping but also distinct epitopes.…”

Section: The Concentration Of Rsv-specific Igg Does Not Differ Betweementioning

confidence: 99%

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Natural killer cell activation by respiratory syncytial virus‐specific antibodies is decreased in infants with severe respiratory infections and correlates with Fc‐glycosylation

et al. 2020

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Objectives Respiratory syncytial virus (RSV) is a major cause of severe lower respiratory tract infections in infants, and there is no vaccine available. In early life, the most important contributors to protection against infectious diseases are the innate immune response and maternal antibodies. However, antibody‐mediated protection against RSV disease is incompletely understood, as both antibody levels and neutralisation capacity correlate poorly with protection. Since antibodies also mediate natural killer (NK) cell activation, we investigated whether this functionality correlates with RSV disease. Methods We performed an observational case–control study including infants hospitalised for RSV infection, hernia surgery or RSV‐negative respiratory viral infections. We determined RSV antigen‐specific antibody levels in plasma using a multiplex immunoassay. Subsequently, we measured the capacity of these antibodies to activate NK cells. Finally, we assessed Fc‐glycosylation of the RSV‐specific antibodies by mass spectrometry. Results We found that RSV‐specific maternal antibodies activate NK cells in vitro. While concentrations of RSV‐specific antibodies did not differ between cases and controls, antibodies from infants hospitalised for severe respiratory infections (RSV and/or other) induced significantly less NK cell interferon‐γ production than those from uninfected controls. Furthermore, NK cell activation correlated with Fc‐fucosylation of RSV‐specific antibodies, but their glycosylation status did not significantly differ between cases and controls. Conclusion Our results suggest that Fc‐dependent antibody function and quality, exemplified by NK cell activation and glycosylation, contribute to protection against severe RSV disease and warrant further studies to evaluate the potential of using these properties to evaluate and improve the efficacy of novel vaccines.

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“…To quantify the concentration of RSV-specific IgG and IgA, a multiplex immunoassay was performed as described before (30). Briefly, diluted plasma samples were incubated with RSV antigen-coupled beads, including G A , G B , post-F, and pre-F (DS-CAV1 mutant) (31).…”

Section: Methodsmentioning

confidence: 99%

Natural killer cell activation by respiratory syncytial virus-specific antibodies is decreased in infants with severe respiratory infections and correlates with Fc-glycosylation

Erp

Lakerveld

Graaf

et al. 2019

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AbstractBackgroundRespiratory syncytial virus (RSV) is a major cause of severe lower respiratory tract infections in infants and there is no vaccine available. In early life, the most important contributors to protection against infectious diseases are the innate immune system and maternal antibodies. However, the mechanisms by which antibodies can protect against RSV disease are incompletely understood, as both antibody levels and neutralization capacity correlate poorly with protection. We therefore asked whether antibody-mediated natural killer (NK) cell activation correlates with RSV disease.MethodsWe performed an observational case-control study including infants hospitalized for RSV infection (n=43, cases), hernia surgery (n=16, controls), or RSV-negative viral respiratory tract infections (n=18, controls). First, we determined RSV antigen-specific antibody levels in infant plasma using a multiplex immunoassay. Subsequently, we measured the capacity of these antibodies to activate NK cells. Finally, we assessed Fc-glycosylation of the RSV-specific antibodies by mass spectrometry.ResultsWe found that RSV-specific maternal antibodies potently activate NK cells in vitro. While the concentrations of RSV-specific antibodies did not differ between cases and controls, antibodies from infants hospitalized for severe lower respiratory tract infections (RSV and/or other) induced significantly less NK cell interferon gamma production than those from uninfected controls. Furthermore, NK cell activation correlated with Fc-fucosylation of RSV-specific antibodies, but their glycosylation status did not significantly differ between cases and controls.ConclusionsOur results suggest that Fc-dependent antibody function and quality, exemplified by NK cell activation and glycosylation, contribute to protection against severe RSV disease and warrant further studies to evaluate the potential of harnessing these activities to develop an effective vaccine.

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Development and Standardization of a High-Throughput Multiplex Immunoassay for the Simultaneous Quantification of Specific Antibodies to Five Respiratory Syncytial Virus Proteins

Cited by 23 publications

References 27 publications

Antibody and Local Cytokine Response to Respiratory Syncytial Virus Infection in Community-Dwelling Older Adults

Antibody and Local Cytokine Response to Respiratory Syncytial Virus Infection in Community-Dwelling Older Adults

Natural killer cell activation by respiratory syncytial virus‐specific antibodies is decreased in infants with severe respiratory infections and correlates with Fc‐glycosylation

Natural killer cell activation by respiratory syncytial virus-specific antibodies is decreased in infants with severe respiratory infections and correlates with Fc-glycosylation

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