Preparation of Monoclonal Antibody against Major Capsid Protein of Largemouth Bass Iridovirus and Its Epitope Identification

doi:10.15933/j.cnki.1004-3268.2024.10.016

Abstract

Abstract: The major capsid protein(MCP)was selected as the target protein of largemouth bass iridovirus(LMBV),MCP recombinant protein was expressed by prokaryotic expression system and used to immunize BALB/c mice.The immunized mice spleen B lymphocytes were fused with mouse myeloma cells(SP/20),and monoclonal antibodies were obtained through subclonal screening,abdominal inoculation and collection of ascites.The specificity of the monoclonal antibody and its epitopes were studied by Western blot and immunohistochemistry.The results showed that one monoclonal antibody strain 2C9⁃6 was obtained.The specificity analysis results suggested that 2C9⁃6 could specifically recognize eukaryotically expressed LMBV⁃MCP and LMBV virus particles.Epitope analysis results showed that the epitope recognized by 2C9⁃6 was located in the peptide segment composed of amino acids at positions 1—20 of the N terminal of LMBV⁃MCP protein.

Key words: Largemouth bass iridovirus（LMBV）, Main capsid protein（MCP）, Monoclonal antibody, Epitope

摘要： 以大口黑鲈虹彩病毒（Largemouth bass iridovirus，LMBV）主衣壳蛋白（Major capsid protein，MCP）为检测靶蛋白，利用原核表达系统表达MCP重组蛋白并免疫BALB/c小鼠，将免疫小鼠脾B淋巴细胞与小鼠骨髓瘤细胞（SP/20）融合，经亚克隆筛选、小鼠腹腔接种、收集腹水等过程获得单克隆抗体；利用免疫印迹技术对所制备的单克隆抗体免疫识别的特异性以及其识别的抗原表位进行研究分析，为研制适于养殖现场快速检测LMBV的胶体金检测试纸条奠定基础。结果表明，获得了1株单克隆抗体2C9-6。特异性分析结果显示，单克隆抗体2C9-6可以特异性识别真核表达的LMBV-MCP和LMBV病毒颗粒。抗原表位分析结果表明，单克隆抗体2C9-6识别LMBV-MCP的抗原表位位于LMBV-MCP蛋白N端第1—20位氨基酸。

关键词: 大口黑鲈虹彩病毒, 主衣壳蛋白, 单克隆抗体, 抗原表位

CLC Number:

S948

SONG Zhanlin, MING Shengli, ZENG Lei, PAN Jiajia, ZHAO Liming, LIU Taoxue, WANG Jiang, LIU Zhonghu, . Preparation of Monoclonal Antibody against Major Capsid Protein of Largemouth Bass Iridovirus and Its Epitope Identification[J]. Journal of Henan Agricultural Sciences, 2024, 53(10): 149-158.

宋占林, 明胜利, 曾磊, 潘佳佳, 赵黎明, 刘桃雪, 王江, 刘忠虎. 大口黑鲈虹彩病毒主衣壳蛋白单克隆抗体制备与抗原表位鉴定[J]. 河南农业科学, 2024, 53(10): 149-158.

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