2E and Supporting Information Fig. 1). To ensure that acid-stripping of passively bound IgE is not affecting the detection of IgE, we repeated the Nb infection with CD23−/− IgEki/ki. Again, WT mice express IgG1 (19.9%), whereas only little chimeric IgE can be detected in the mesenteric lymph nodes of CD23−/− IgEki/ki mice (2.76%). Similar low level IgE expression was

seen in spleen and bone marrow of IgEki/ki Pirfenidone concentration mice. PCR reveals the presence of membrane IgE-IgG1 transcripts in spleen (Fig. 2G) without IgE+ cells (Fig. 2F). These findings suggest that IgE-membrane expressing B cells in mice are either rare or cannot be expressed in vivo, even in the IgE-IgG1 chimeric form. In the case of Nb infection, this is fundamentally different from IgG1+ B cells, which can readily be detected in Akt inhibitor lymph nodes of infested mice. In this context, Achatz-Straussberger et al. [31]. demonstrated that IgE-secreting B cells, from a different IgE-IgG1 chimeric knock-in mouse called KN1, do migrate more efficiently toward the chemokine CXCL12 into plasma cell niches in the bone marrow. However, in our model bone marrow is not populated more efficiently by IgE+ B cells after Nb infestation. The knock-in strategy allows the natural recombination and selection process for the generation of an antigen-specific polyclonal immunoglobulin response. Immunization with the model antigen TNP-OVA, adsorbed to the Th-2 response favoring

adjuvant alum, allowed the comparison of IgE and IgG1 production (Fig. 3A). IgEwt/wt mice produced very little TNP-OVA-specific IgE, but high titers of TNP-OVA-specific total IgG, including IgG1 (Fig. 3B). In contrast, we observed a strong increase of antigen-specific IgE titers in the IgEki mice compared with that of WT littermates, an 11-fold increase for IgEwt/ki and a 42-fold increase for IgEki/ki, respectively. IgG1 was not significantly reduced

in IgEwt/ki, but is absent in IgEki/ki mice (Fig. 3B). The genetic deficiency of IgG1 may account for the reduction of total IgG in IgEki/ki mice, despite the relative increase in IgG2a, IgG2b, and IgG3 levels (Fig. 3B). One further difference between WT and IgEki is a significant increase in antigen-specific IgM (data not shown). One possible explanation is that CD23-mediated uptake Pregnenolone of antigen and a novel mechanism of an antigen transporting capacity by B cells to dendritic cells enhance the specific antigen response, which leads to a stronger immunoglobulin response [30]. This hypothesis is supported by the observation that the changes in CD23−/− IgE knock-in mice for IgG3 and IgG2b are less distinct or absent, respectively (Fig. 3C). Taken together, the IgE knock-in strategy leads to complete lack of IgG1 in homozygous IgEki mice, reduction of IgG and a very strong specific IgE response in both IgE knock-in genotypes. It was recently suggested that basophils have a crucial role in IgG1-mediated anaphylaxis [9].