Structure and applications of novel influenza HA tri-stalk protein for evaluation of HA stem-specific immunity

PLoS One. 2018 Sep 27;13(9):e0204776. doi: 10.1371/journal.pone.0204776. eCollection 2018.


Long alpha helix (LAH) from influenza virus hemagglutinin (HA) stem or stalk domain is one of the most conserved influenza virus antigens. Expression of N-terminally extended LAH in E. coli leads to assembly of α-h elical homotrimer which is structurally nearly identical to the corresponding region of post-fusion form of native HA. This novel tri-stalk protein was able to differentiate between group 1 and 2 influenza in ELISA with virus-infected mice sera. It was also successfully applied for enzyme-linked immunospot assay to estimate the number of HA stem-reactive antibody (Ab)-secreting cells in mice. An in-house indirect ELISA was developed using a HA tri-stalk protein as a coating antigen for evaluation of HA stem-specific Ab levels in human sera collected in Luxembourg from 211 persons with occupational exposure to swine before the pandemic H1N1/09 virus had spread to Western Europe. Our results show that 70% of these pre-pandemic sera are positive for HA stem-specific Abs. In addition, levels of HA stem-specific Abs have positive correlation with the corresponding IgG titers and neutralizing activities against pandemic H1N1/09 virus.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antibodies, Viral / immunology*
  • Female
  • Hemagglutinin Glycoproteins, Influenza Virus* / chemistry
  • Hemagglutinin Glycoproteins, Influenza Virus* / immunology
  • Humans
  • Influenza A Virus, H1N1 Subtype* / chemistry
  • Influenza A Virus, H1N1 Subtype* / immunology
  • Influenza, Human* / epidemiology
  • Influenza, Human* / immunology
  • Influenza, Human* / pathology
  • Male
  • Mice
  • Mice, Inbred BALB C
  • Pandemics*
  • Plasma Cells / immunology*
  • Plasma Cells / pathology
  • Protein Structure, Secondary


  • Antibodies, Viral
  • H1N1 virus hemagglutinin
  • Hemagglutinin Glycoproteins, Influenza Virus

Grants and funding

The research leading to these results has received funding from the European Union Seventh Framework Programme for research, technological development and demonstration under grant agreement № 602437 (all authors) and from European Regional Development Fund project (AKi, KT and AKa). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.