Viral envelope protein gp64 transgenic mouse facilitates the generation of monoclonal antibodies against exogenous membrane proteins displayed on baculovirus
Viral envelope protein gp64 transgenic mouse facilitates the generation of monoclonal antibodies against exogenous membrane proteins displayed on baculovirus
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DOI：
10.1016/j.jim.2007.02.005
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发表时间：
2007-04-30
影响因子：
2.2
通讯作者：
Hamakubo, Takao
中科院分区：
文献类型：
--
作者：
Saitoh, Ryoichi;Ohtomo, Toshihiko;Hamakubo, Takao
本研究以出芽杆状病毒（budded baculovirus，BV）为研究对象，利用病毒囊膜蛋白gp 64转基因小鼠直接免疫BV，研究BV上的转运蛋白、G蛋白偶联受体等多通道膜蛋白的功能展示。与野生型BALB/ c小鼠相比，gp 64转基因小鼠对病毒的反应较弱。用表达肽转运蛋白PepT 1的BV免疫gp 64转基因小鼠，获得47株单克隆抗体。通过荧光细胞分析，这些mAb对胰腺癌细胞AsPC-1上的PepT 1具有特异性，并且对AsPC-1表现出抗体依赖性细胞毒性或补体依赖性细胞毒性。我们还通过用表达趋化因子受体CCR 2的BV与部分纯化的CCR 2一起免疫CCR 2缺陷背景下的gp 64转基因小鼠产生了7种rnAb。这些单克隆抗体具有特异性结合CCR 2在CHO细胞的荧光分析，并表现出中和活性的配体依赖性抑制环AMP的生产。该方法为产生针对膜蛋白的治疗/诊断mAb提供了强有力的工具。(C)2007 Elsevier B. V.保留所有权利。
We have been investigating the functional display of multipass membrane protein such as transporter or G-protein coupled receptor on the budded baculovirus (BV).We tested the use of a viral envelope protein gp64 transgenic mouse for the direct immunization of these membrane proteins displayed on BVs. The gp64 transgenic mice showed only a weak response to virus compared to wild type BALB/ c mice. Immunizing gp64 transgenic mice with the BV expressing peptide transporter PepT1, we obtained 47 monoclonal antibodies (mAbs). These mAbs were specific to the PepT1 on the pancreatic cancer cells AsPC-1 by fluorocytometric analysis and exhibited antibody-dependent cellular cytotoxicity or complement-dependent cytotoxicity to AsPC-1. We also generated 7 rnAbs by immunizing gp64 transgenic mice on a CCR2-deficient background with the BV expressing chemokine receptor CCR2 together with partially purified CCR2. These mAbs possessed specific binding to CCR2 in CHO cells on fluorocytometric analysis, and exhibited neutralizing activities for ligand-dependent inhibition of cyclic AMP production. This method provides a powerful tool for the generation of therapeutic/diagnostic mAbs against membrane proteins. (C) 2007 Elsevier B.V. All rights reserved.