Production of Monoclonal Antibody to 2′,3′‐Cyclic Nucleotide 3′‐Phosphodiesterase from Bovine Cerebral White Matter
Production of Monoclonal Antibody to 2′,3′‐Cyclic Nucleotide 3′‐Phosphodiesterase from Bovine Cerebral White Matter
复制标题
牛脑白质2′,3′-环核苷酸3′-磷酸二酯酶单克隆抗体的制备
DOI：
10.1111/j.1471-4159.1986.tb02849.x
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发表时间：
1986
影响因子：
4.7
通讯作者：
Y. Tsukada
中科院分区：
文献类型：
--
作者：
M. Fujishiro;S. Kohsaka;K. Nagaike;Y. Tsukada
用部分纯化的牛脑白色物质2′，3 ′-环核苷酸3′-磷酸二酯酶（CNE）免疫刘易斯大鼠，取脾细胞与小鼠骨髓瘤细胞系SP-2融合。通过（1）酶联免疫吸附试验、（2）牛大脑免疫组织化学染色、（3）蛋白质印迹分析和（4）CN结合试验检测单克隆抗体的产生。获得了与CNPase分子特异性结合的单克隆抗体。然而，抗体不抑制酶活性。Western blotting分析表明，单克隆抗体结合CNa（Wla）和CNb（Wlb）。该单克隆抗体被鉴定为IgG 2c亚类。免疫组织化学检查显示，在几个物种（牛、小鼠、大鼠和人）中，CNS中的髓鞘被单克隆抗体严重染色。与此相反，周围神经系统髓鞘没有染色，即使在牛组织。这些结果表明，在本研究中获得的单克隆抗体特异性地识别CNS中的CN-4分子。
Lewis rats were immunized with partially purified 2′,3′‐cyclic nucleotide 3′‐phosphodiesterase (CNPase) from bovine cerebral white matter and the spleen cells were fused with cell of a mouse myeloma cell line (SP‐2). The production of monoclonal antibody was detected by (1) enzyme‐linked immunoadsorbent assay, (2) immunohistochemical staining of bovine cerebrum, (3) Western blotting analysis, and (4) CNPase binding assay. Monoclonal antibody that specifically binds CNPase molecules was obtained. However, the antibody did not suppress the enzyme activity. Western blotting analysis demonstrated that the monoclonal antibody binds both CNa (Wla) and CNb (Wlb). The monoclonal antibody was identified as being of the IgG2c subclass. Immunohistochemical examination revealed that the myelin sheath in the CNS was heavily stained with the monoclonal antibody in several species (bovine, mouse, rat, and human). In contrast, peripheral nervous system myelin was not stained even in bovine tissue. These results suggest that the monoclonal antibody obtained in the present study specifically recognizes the CNPase molecules in the CNS.