Characterization of the Rabbit Neonatal Fc Receptor (FcRn) and Analyzing the Immunophenotype of the Transgenic Rabbits That Overexpresses FcRn

The neonatal Fc receptor (FcRn) regulates IgG and albumin homeostasis, mediates maternal IgG transport, takes an active role in phagocytosis, and delivers antigen for presentation. We have previously shown that overexpression of FcRn in transgenic mice significantly improves the humoral immune response. Because rabbits are an important source of polyclonal and monoclonal antibodies, adaptation of our FcRn overexpression technology in this species would bring significant advantages. We cloned the full length cDNA of the rabbit FcRn alpha-chain and found that it is similar to its orthologous analyzed so far. The rabbit FcRn - IgG contact residues are highly conserved, and based on this we predicted pH dependent interaction, which we confirmed by analyzing the pH dependent binding of FcRn to rabbit IgG using yolk sac lysates of rabbit fetuses by Western blot. Using immunohistochemistry, we detected strong FcRn staining in the endodermal cells of the rabbit yolk sac membrane, while the placental trophoblast cells and amnion showed no FcRn staining. Then, using BAC transgenesis we generated transgenic rabbits carrying and overexpressing a 110 kb rabbit genomic fragment encoding the FcRn. These transgenic rabbits – having one extra copy of the FcRn when hemizygous and two extra copies when homozygous - showed improved IgG protection and an augmented humoral immune response when immunized with a variety of different antigens. Our results in these transgenic rabbits demonstrate an increased immune response, similar to what we described in mice, indicating that FcRn overexpression brings significant advantages for the production of polyclonal and monoclonal antibodies

Binding and Vesiculation of Rabbit IgG by Rabbit Yolk Sac Membrane

Abstract Studies are described which establish the glycocalyx as the morphologic site for the binding of IgG to the membrane of the rabbit yolk sac and which indicate that micropinocytotic vesiculation is the mechanism for its cellular uptake. Yolk sac membranes were exposed in vivo and in vitro to rabbit antihorse ferritin antibody. The membranes were then treated with ferritin and examined by electron microscopy for deposition of ferritin. Deposits of IgG-bound ferritin were found localized at the glycocalyx coat of the microvilli, intermicrovillar pits, and incipient micropinocytotic vesicles. Stages in the micropinocytotic process are shown, depicting internalization of microvillial membrane surfaces, indicating that this is the mechanism by which IgG is taken up by the fetal yolk sac from the maternal uterine lumen for transfer to the fetus. These findings are discussed in the context of the specificity and kinetic characteristics of the transfer process.</jats:p