Supplementary Material for: Postnatal and Adult Immunoglobulin Repertoires of Innate-Like CD19⁺CD45R^lo B Cells

The diversity in antibody repertoire relies on different B cell populations working efficiently to fulfil distinct specific functions. We recently described an innate-like CD19⁺CD45R^-/lo (19⁺45R^lo) cell population in postnatal unstimulated adult mice, a heterogeneous population containing cells expressing immunoglobulin M (IgM) and others behaving as differentiated mature B lymphocytes (intracytoplasmic IgG1, AID⁺, Blimp-1⁺RAG2^-). In the present study, we characterized the Ig repertoire expressed by splenic 19⁺45R^lo cells, assuming that they would bear a restricted repertoire biased for germline rearrangements and low mutation rates similar to other innate-like cells. Sequences from 19⁺45R^lo cells displayed a variety of V, D and J regions, and the analysis of the CDR-H3 region revealed an intermediate overall CDR-H3 length and moderate hydrophobicity. Both IgM and switched sequences of PD15 19⁺45R^lo cells had shorter CDR-H3 region and fewer non-template N nucleotides than adult sequences, as expected for profiles that correspond to an immature phenotype. Regarding the mutation rate in the VH regions, IgG1 sequences already carried a high rate of replacement mutations at PD15, which increased further in the sequences obtained from adult mice. Moreover, statistical models suggest that a proportion of the switched sequences in adult 19⁺45R^lo cells had experienced antigen selection, unlike other innate-like B cell compartments.