Abstract
Antibody responses against the influenza A virus hemagglutinin (HA)-protein are studied intensively because they can protect against (re)infection. Previous studies have focused on antibodies targeting the head or stem domains, while other possible specificities are often not taken into account. To study such specificities, we developed a diverse set of HA-domain proteins based on an H1N1pdm2009-like influenza virus strain, including monomeric head and trimeric stem domain, as well as the full HA-trimer. These proteins were used to study the B cell and antibody responses in six healthy human donors. A large proportion of HA-trimer B cells bound exclusively to HA-trimer probe (54–77%), while only 8–18% and 9–23% were able to recognize the stem or head probe, respectively. Monoclonal antibodies (mAbs) were isolated and three of these mAbs, targeting the different domains, were characterized in-depth to confirm the binding profile observed in flow cytometry. The head-directed mAb, targeting an epitope distinct from known head-specific mAbs, showed relatively broad H1N1 neutralization and the stem-directed mAb was able to broadly neutralize diverse H1N1 viruses. Moreover, we identified a trimer-directed mAb that did not compete with known head or stem domain specific mAbs, suggesting that it targets an unknown epitope or conformation of influenza virus’ HA. These observations indicate that the described method can characterize the diverse antibody response to HA and might be able to identify HA-specific B cells and antibodies with previously unknown specificities that could be relevant for vaccine design.
Highlights
The antibody responses elicited by influenza A virus infections or vaccines provide a key component in the protection against future influenza virus infections
A nonreduced and reduced SDS-PAGE stained with Coomassie blue was performed and all proteins showed the expected size on the gels; full-length HA-trimer around 75 kDa, monomeric head and trimeric stem around 30 kDa and no significant impurities were observed
We report on immune assays involving three different probes representing the HA trimer, head and stem domains to analyze the overall antibody response directed against the influenza A virus’ HA
Summary
The antibody responses elicited by influenza A virus infections or vaccines provide a key component in the protection against future influenza virus infections. HA is highly diverse; to date, eighteen subtypes have been described for both avian and human influenza A, with high genetic and antigenic diversity within each subtype [4,5,6] This diversity is a challenge for the human immune system and it is important to understand which domains are targets for (neutralizing) antibodies and what the conservation of these antibody targets is within and between different HA subtypes. Antibodies elicited by conventional vaccines target primarily the immunodominant globular head domain of HA [10,11,12] These antibodies generally bind the receptor binding site (RBS) and the adjacent (highly variable) antigenic sites [13,14] and they usually neutralize the virus by obstructing its binding to the receptor on the cell surface [15]. While the HA head and stem domains feature prominently in antibody and vaccine studies, much less is known about other targets such as quaternary epitopes on the HA trimer
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
