Quantifying the spectrum and dynamics of B-cell binding avidity in response to influenza

PROJECT SUMMARY Activation, clonal expansion, and affinity maturation of antigen-specific B cells are considered the hallmarks of adaptive immunity; hence, the assessment of B cell responses is thought to provide correlates of immune protection. Antigen-specific B cell responses have traditionally been observed indirectly, through the study of the magnitude of the serum antibody pool using ELISA. Direct measurements of antibody-secreting cells have also been done using ELISPOT, or more recently by flow cytometry (FC). None of these methods, however, provide a measure of a B cell's overall binding avidity for its cognate antigen, despite the fact that this measure is frequently acknowledged as prognostic of immune protection and may be critical for the evaluation of functional immunity. The primary goals of this project are to i) develop a microfluidic platform to capture and isolate B cells according to their binding avidity for viral antigens, and ii) use the platform to examine how B cell binding avidity changes in time (affinity maturation) following infection and correlates with functional immunity. Our long-term objective is to develop new tools with which to rapidly and effectively assess immune cell function. This project expands on our previous work with tumor-specific T cells, and specifically examines the B cell response to influenza A in hybridoma and murine model systems. The specific aims of this project are to: 1) Design and optimize a microfluidic device to determine the specificity and binding avidities of influenza virus nuclear protein (NP)- and hemagglutinin (HA)-specific monoclonal B cells; and 2) characterize binding avidity dynamics of HA- and NP-specific peripheral B cells following influenza virus infection. Our platform will demonstrate the functionality of a rapid and easily adaptable system to evaluate the dynamics of the B cell response to influenza but is completely adaptable to host of other pathogens such as SARS-Cov-2. Furthermore, the project will begin to vigorously test long-held dogmas regarding the potential critical importance of affinity maturation in the complex setting of an acute infection that induces polyclonal extrafollicular and germinal center B cell responses, as well as local and systemic B cell immunity, where related earlier studies have failed to demonstrate affinity maturation as a main driver of protective immunity.