antibody repertoire; bioinformatic; B cells; high-throughput; humoral immunity
Khan Tarik A, Friedensohn Simon, Gorter de Vries Arthur R, Straszewski Jakub, Ruscheweyh Hans-Joachim, Reddy Sai T (2016), Accurate and predictive antibody repertoire profiling by molecular amplification fingerprinting., in Science advances
, 2(3), 1501371-1501371.
Greiff Victor, Bhat Pooja, Cook Skylar C, Menzel Ulrike, Kang Wenjing, Reddy Sai T (2015), A bioinformatic framework for immune repertoire diversity profiling enables detection of immunological status., in Genome medicine
, 7(1), 49-49.
Greiff Victor, Miho Enkelejda, Menzel Ulrike, Reddy Sai T (2015), Bioinformatic and Statistical Analysis of Adaptive Immune Repertoires., in Trends in immunology
, 36(11), 738-49.
Menzel Ulrike, Greiff Victor, Khan Tarik A, Haessler Ulrike, Hellmann Ina, Friedensohn Simon, Cook Skylar C, Pogson Mark, Reddy Sai T (2014), Comprehensive evaluation and optimization of amplicon library preparation methods for high-throughput antibody sequencing., in PloS one
, 9(5), 96727-96727.
Schanz Merle, Liechti Thomas, Zagordi Osvaldo, Miho Enkelejda, Reddy Sai T, Günthard Huldrych F, Trkola Alexandra, Huber Michael (2014), High-throughput sequencing of human immunoglobulin variable regions with subtype identification., in PloS one
, 9(11), 111726-111726.
Greiff Victor, Menzel Ulrike, Haessler Ulrike, Cook Skylar C, Friedensohn Simon, Khan Tarik A, Pogson Mark, Hellmann Ina, Reddy Sai T (2014), Quantitative assessment of the robustness of next-generation sequencing of antibody variable gene repertoires from immunized mice., in BMC immunology
, 15, 40-40.
Haessler Ulrike, Reddy Sai T (2014), Using next-generation sequencing for discovery of high-frequency monoclonal antibodies in the variable gene repertoires from immunized mice., in Methods in molecular biology (Clifton, N.J.)
, 1131, 191-203.
The memory B cell response recognizes and responds to pathogens by initiating a series of molecular and cellular reactions, which culminate in the generation of antigen-specific polyclonal antibodies; therefore memory B cells plays an indispensible role in maintaining immunological protection. Following initial encounter and stimulation by antigen, naïve B cells differentiate into either short-lived plasma cells or form germinal centers (GC), which result in the further differentiation of memory B cells or long-lived antibody-secreting plasma cells. Despite their immense importance to immunity, there are still many unknowns related to the differentiation and development of memory B cells. For example, most previous studies have relied on transgenic mouse models and analysis of monoclonal-like B cell responses. Consequentially, such approaches were not able to capture the complexities and repertoire diversity present in a more “natural” immune response, one that leads to the generation of a polyclonal population of antibodies against antigen. The overall goal of this proposal is to provide a quantitative molecular analysis of polyclonal memory B cell responses in wild-type mice following immunization with protein antigens, which will enable us to elucidate mechanisms governing differentiation and development of memory B cells and plasma cells. Our analysis will be conducted with a systems immunology-based approach; specifically this will consist of magnetic-associated sorting coupled to flow cytometry for isolation of antigen-specific IgG+ memory B cells and plasma cells (Aim 1). Next we will use the Illumina miSeq platform for high-throughput DNA sequencing of antibody variable genes from the isolated antigen-specific cells; this will be followed by extensive bioinformatic and statistical analysis for the characterization and quantification of several features related to polyclonal antibody repertoires (Aim 2). In addition to a comprehensive genetic analysis, we will also perform a thorough investigation of the polyclonal antibodies on a protein-based level. This will include recombinant expression of several high-frequency antibodies from each memory B cell and plasma cell subset followed by biophysical characterization of binding affinity towards antigen (Aim 3). The major outcome of all of these aims will be to gain substantial insight into the differentiation and development of memory B cells and plasma cells. For example, we will be able to address whether memory B cells and plasma cells follow a bifurcative or linear differentiation program. Additionally, we will also quantify the clonal diversity of each antigen-specific memory repertoire. We will also determine the relationship between high frequency of antibody expression in a repertoire and it’s binding affinity towards antigen. Importantly, since all of these studies will be performed on a quantitative and systems-based manner, we will be able to generate an unprecedented depth of information on the antigen-specific polyclonal memory B cell and plasma cell response, which will provide valuable insight in the design and development of vaccines and immunotherapeutics.