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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
Summary
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.
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.
