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Flow-Cytometric Method Measuring B Cell Surface Immunoglobulin Avidity.

Davide Angeletti1, Gregory M Frank1, Jonathan W Yewdell2

  • 1Laboratory of Viral Diseases, National Institutes of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD, USA.

Methods in Molecular Biology (Clifton, N.J.)
|June 8, 2017
PubMed
Summary
This summary is machine-generated.

Researchers developed a simple flow cytometry method to measure antibody avidity against influenza virus hemagglutinin. This technique quantifies B cell avidity, aiding studies of viral immunity and vaccine responses.

Keywords:
AffinityAvidityB cellFlow cytometryGerminal centerHemagglutininImmunoglobulinInfluenza

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Area of Science:

  • Immunology
  • Virology
  • Biotechnology

Background:

  • Antibody affinity maturation is crucial for effective humoral immunity, occurring in germinal centers (GC) via B cell competition.
  • Measuring antibody avidity against complex antigens like viruses in polyclonal sera is challenging.
  • Understanding B cell avidity is key to assessing immune responses to pathogens and vaccines.

Purpose of the Study:

  • To present a straightforward flow cytometry-based method for determining the average avidity of germinal center B cells for influenza A virus hemagglutinin.
  • To establish a quantitative measure (AC50) reflecting B cell population avidity.
  • To provide a tool applicable to basic and clinical studies of viral immunity.

Main Methods:

  • Utilized flow cytometry with fluorescently labeled hemagglutinin and B cell markers.
  • Employed a graded antigen concentration approach to quantify B cell binding.
  • Defined the AC50 as the antigen concentration needed to stain 50% of hemagglutinin-specific B cells within the GC B cell population.

Main Results:

  • The developed method allows for high-throughput, qualitative, and quantitative detection of individual B cells.
  • The calculated AC50 value strongly correlates with the measured avidity of the GC B cell population for hemagglutinin.
  • Demonstrated the method's effectiveness in assessing antibody avidity against a key viral antigen.

Conclusions:

  • The presented flow cytometry assay provides a reliable and accessible means to measure B cell avidity against influenza hemagglutinin.
  • This method can be broadly applied to assess antibody avidity in various viral infections and vaccine studies.
  • Facilitates a deeper understanding of immune responses to medically relevant immunogens.