Abstract
High-throughput sequencing of the B cell receptor (BCR) repertoire can provide rapid characterization of the B cell response in a wide variety of applications in health, after vaccination and in infectious, inflammatory and immune-driven disease, and is starting to yield clinical applications. However, the interpretation of repertoire data is compromised by a lack of studies to assess the intra and inter-individual variation in the BCR repertoire over time in healthy individuals. We applied a standardized isotype-specific BCR repertoire deep sequencing protocol to a single highly sampled participant, and then evaluated the method in 9 further participants to comprehensively describe such variation. We assessed total repertoire metrics of mutation, diversity, VJ gene usage and isotype subclass usage as well as tracking specific BCR sequence clusters. There was good assay reproducibility (both in PCR amplification and biological replicates), but we detected striking fluctuations in the repertoire over time that we hypothesize may be due to subclinical immune activation. Repertoire properties were unique for each individual, which could partly be explained by a decrease in IgG2 with age, and genetic differences at the immunoglobulin locus. There was a small repertoire of public clusters (0.5, 0.3, and 1.4% of total IgA, IgG, and IgM clusters, respectively), which was enriched for expanded clusters containing sequences with suspected specificity toward antigens that should have been historically encountered by all participants through prior immunization or infection. We thus provide baseline BCR repertoire information that can be used to inform future study design, and aid in interpretation of results from these studies. Furthermore, our results indicate that BCR repertoire studies could be used to track changes in the public repertoire in and between populations that might relate to population immunity against infectious diseases, and identify the characteristics of inflammatory and immunological diseases.
Highlights
An effective humoral immune response is in part dependent on having a diversity of B cells with different B cell receptors (BCRs) capable of recognizing and binding to many different antigens
We show high reproducibility in the methods used to sample the BCR repertoire, and not exhaustively sampling the entire repertoire at the individual B cell clone level, calculate comparable global repertoire metrics, and routinely detect abundant clusters in repeat samples
In the absence of an immune stimulus given during the study, there was considerable variation in the BCR repertoire over time in a single individual, highlighting that it is a highly dynamic system that is constantly subject to immune stimulus and selective pressures
Summary
An effective humoral immune response is in part dependent on having a diversity of B cells with different B cell receptors (BCRs) capable of recognizing and binding to many different antigens. The sum of all B cells with distinct BCRs is termed the BCR repertoire, and in humans has the theoretical potential to reach a size of up to 1011 unique variants [1]. BCRs consist of paired heavy and light chains, with primary diversity generated by the somatic recombination of V, D (heavy chain only), and J gene segments during B cell development to form the functional genes [2]. Upon B cell activation and proliferation, there is a secondary diversification step mediated by somatic hypermutation, and selection of B cells with increasing affinity for the antigen [3]. The BCR repertoire is often considered as a single entity, it is a mixture of different B cell subpopulations, each of which may have a distinct repertoire structure [4, 5]
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