The anti-ovalbumin IgG was purified from sera of OVA immunized mice

The anti-ovalbumin IgG was purified from sera of OVA immunized mice. the bFcRn Tg mice. Our Traditional western blot analysis showed strong expression of the bFcRn transgene in peritoneal macrophages and bone marrow derived dendritic cells; and a quantitative PCR analysis demonstrated that the expression ratios of the bFcRn to mFcRn were 2.6- and 10-fold in these cells, respectively. We also found that overexpression of bFcRn enhances the phagocytosis of Ag-IgG immune complexes (ICs) GSK461364 by both macrophages and dendritic cells and significantly improves Ag presentation by dendritic cells. Finally, we determined that immunized bFcRn mice produce a much greater diversity of Ag-specific IgM, whereas only the levels, but not the diversity, of IgG is increased by overexpression of bFcRn. We suggest that the increase in diversity of IgG in Tg mice is prevented by a selective bias towards immunodominant epitopes of ovalbumin, which was used in this study as a model antigen. These results are also in line with our previous reports describing a substantial increase in the levels of Ag-specific IgG in FcRn Tg mice immunized with Ags that are weakly GSK461364 immunogenic and, therefore, not affected by immunodominance. Introduction The production of monoclonal antibodies (mAbs) using hybridoma technology has allowed significant advances in biomedical research and has greatly improved our capacity for clinical diagnostics and therapeutics. Currently, more than 25 immunoglobulins GSK461364 have been approved for therapeutical use in humans and over 240 antibodies are in development targeting a wide variety of diseases, including autoimmunity, cancer, infectious diseases and cardiovascular diseases (reviewed by [1]). In recent years, there has been an increasing demand for the development of cheaper, faster and more efficient technologies for the production of high-affinity and high-specificity mAbs. One approach to improve the efficiency of hybridoma production is to enhance humoral immune response against various antigens (Ags), including weakly immunogenic targets to which mAbs are generally difficult to generate. Another approach is to create a higher diversity of Ag-specific antibodies, allowing for the development of a larger variety of hybridomas, which can be screened for their ability to bind native epitopes and to produce functionally relevant mAbs [2]. To achieve these goals, our group has recently created transgenic (Tg) mice that overexpress the bovine neonatal Fc receptor (bFcRn) [3] and show a greatly augmented humoral immune response. Our previous analyses have shown that the bFcRn Tg mice offer major advantages for hybridoma production and could serve as important tools for the development of new therapeutic mAbs [4]. In addition, we have recently generated Tg rabbits that overexpress the rabbit FcRn and observed similarly improved IgG protection and enhanced humoral immune response as described for bFcRn Tg mice [5]. The neonatal Fc receptor (FcRn) is a MHC Class I-related receptor composed of an -chain and 2-microlobulin (2m) [6] and was originally identified as the protein that mediates the transport of IgG from maternal milk to the small intestine of newborn rodents [7]. FcRn has proven to be a key player in regulating the transport of IgG within and across cells of diverse origins and it also serves to rescue IgG and albumin from degradation, thereby prolonging their half-lives [8]. IgG protection was originally thought to be mediated by capillary endothelial cells [9] TYP but recent findings suggest that this process also occurs in hematopoietic cells [10], [11] and in mammary epithelial cells during lactation [12]. More recently, several publications have shown that FcRn plays major roles in Ag-IgG immune complex (IC) phagocytosis by neutrophils [13], and in Ag presentation of IgG ICs by professional Ag presenting cells (APCs) [14], [15], [16], [17]. We have recently shown that overexpression of bFcRn in Tg mice leads to increased levels of IgG in the serum as a result of a reduction in IgG catabolism. In addition, we found that expression of bFcRn in Tg mice causes an increase in the levels of Ag-specific IgG and IgM during the secondary immune response and leads to an enhanced expansion of Ag-specific B cells and plasma cells in their spleen [18], [19]. We also observed that, upon immunization, bFcRn Tg mice develop enlarged spleens that contain higher numbers of neutrophil granulocytes and dendritic cells (DCs) as compared to wild-type (wt) mice [18], [20]. This augmented immune response is also reflected in the ability of bFcRn Tg mice to produce high levels of Ag-specific antibodies, B cells and plasma cells to weakly immunogenic targets [20] and to produce elevated numbers of Ag-specific hybridomas [19]. To better understand the mechanisms underlying the augmented humoral immune response observed in bFcRn Tg mice, we further characterized the profile of bFcRn transgene expression in different cells of the immune system. In addition, we investigated the effects of overexpression of bFcRn in the phagocytic.