The J chain is shown in magenta and the SC in green. receptor repertoire. H chain genes are assembled from one functional variable (V), one diversity (D), and one joining (J) segment encoded within a single locus to produce a single open reading frame expressed with an H-chain constant (C) segment that determines the type (isotype) of antibody produced, i.e., IgM, IgD, IgE, IgG, or IgA. Within these antibody isotypes, IgG includes four subclasses, and IgA includes two. L chains are encoded by one of two genes, either kappa or lambda, and are similarly rearranged from genetic loci that include V, J, and C segments. In addition to the diversity generated by the combinatorial strategy of gene assembly and junctional diversity, which is the addition of nucleotides to the V(D)J during rearrangement, antigen receptors can be further altered by somatic hypermutation, a process that introduces point mutations during B-cell activation and maturation. The combination of these processes can theoretically generate a staggering number of antibodies with distinct amino acid sequences. Despite extensive studies, we still do not fully understand many aspects of antibody production, for example, the sites of resident B cells responsible for the secretion of antibodies in different biofluids, how B cells enter these sites, and to what extent antibodies within these biofluids share a common origin. In this study [1], the authors describe a proteome-based assessment of the IgA1 clonal repertoire in two biofluids, blood and breast milk. Specifically, NPPB top-down liquid chromatographymass spectrometry (LCMS) profiling of serum and milk IgA1 repertoires was used to determine the most dominant clones in the respective fluids and assess their stability over time. Using well-controlled experiments and specific standards, the authors found a relatively small number of dominant clones in serum and milk IgA1, with many clones shared between the two biofluids. This study demonstrates the power of top-down LCMS for profiling antibody repertoires and provides a protein complement to single-cell NPPB transcriptomic profiling approaches. Functionally, NPPB immunoglobulins consist of two parts: (1) Fab, the antigen binding segment, consisting of the L chain and the N-terminal portion of the H chain, and (2) the Fc segment, the C-terminal parts of the H chain. In the Fab, each H or L chain contains three hypervariable regions called complementarity determining regions that contact antigen epitopes. Whereas Fab parts interact with Eng antigens, based on the specificities imparted by the variable segments of the H and L chains, Fc segments control the effector functions of antibodies by mediating interactions with other components of the human immune system. IgM and IgA may have additional component chains that are important for their functions: the joining (J) chain and a secretory component. The J chain can combine antibody molecules into larger structures, connecting two or more IgA molecules to form polymeric IgA and promoting the oligomerization of IgM into a pentamer. Secretory components are found on secretory immunoglobulins, i.e., polymeric IgA and polymeric IgM molecules on mucosal surfaces. IgA in humans consists of two subclasses, IgA1 and IgA2. IgA1 is the predominant subclass in circulation, mainly present as a monomer (~8090%) with a small proportion of polymeric IgA1 (~1020%; mostly dimeric). Secretory IgA1, however, is present in substantial amounts in human milk. Structures of monomeric IgA1, dimeric IgA1, and secretory IgA1 are depicted in Fig.1[24]. == Fig. 1. == Models of 3-D structures of monomeric, dimeric, and secretory IgA1. Surface renderings of humanAmonomeric IgA1,Bdimeric IgA1, andCdimeric IgA1 bound to the secretory component (SC) of the polymeric immunoglobulin receptor. Models were generated based on the solution structure of IgA1 [PDB ID: 1IGA, [2]] and the structure of the dimeric Fc/J chain/SC complex [PDB ID: 6UE7, [3]], as noted in [4]. Heavy chains are shown in cyan or pale cyan. Light chains are shown in yellow or pale yellow. The J chain is usually shown in magenta and the SC in green. The NPPB model of monomeric IgA1 shows frequently utilized sites forO-glycosylation (T225, T228, S230, S232, T233, T236; shaded in red) within the hinge region of IgA1. The site of cleavage by theO-glycan-dependent protease, OgpA, is usually denoted by an arrow for one Fab. This site is usually N-terminal of the IgA1.