Molecular analysis of B cell receptor activation by Fab-bridge

本研究的目的是建立一个可以调节B细胞受体交联条件的实验系统。为此，我们尝试创建“Fab-bridge”。Fab桥的基本概念是用不同长度的连接体连接抗小鼠IgM单克隆抗体的两个Fab部分。首先，我们计划培养针对骨髓瘤IgM fc部分的单克隆抗体。虽然人们认为用木瓜蛋白酶消化法制备IgM的Fc部分比较困难，但Dombrink-Kurzman和Voss报道了用低温木瓜蛋白酶消化法制备Fc （Molecular Immunology, 25, 1309-1320(1988)）。然而，我们无法从我们的IgM中通过木瓜蛋白酶消化得到IgM的Fc。在Fab部分的生成过程中，Fc部分被完全降解。因此，我们制备了抗全IgM的单克隆抗体。我们成功地获得了四种单克隆抗体。目前，我们正在评估这些抗体刺激小鼠脾B细胞的能力。

Aim of this study is to set up an experimental system in which we can able to regulate condition of crosslinking of B cell receptor. For this purpose we tried to create "Fab-bridge". Basic concept of Fab-bridge is to connect two Fab portions of anti-mouse IgM monoclonal antibody using linker with different length. Firstly, we planned to raise monoclonal antibody to Fc-portion of myeloma IgM. Although it is considered to be difficult to prepare Fc portion of IgM by papain digestion, Dombrink-Kurzman and Voss reported the Fc preparation by low-temperature papain digestion (Molecular Immunology, 25, 1309-1320(1988)). However, we could not get Fc of IgM by papain-digestion from our IgM. During generation of the Fab portion, Fc portion was completely degraded. So we raised monoclonal antibody against whole IgM. We succeeded in obtaining four monoclonal antibodies. Currently, we are evaluating the ability of these antibody to stimulate mouse splenic B cells.