Preparation of monoclonal antibody against lignin

抗木质素单克隆抗体的制备

• 批准号: 17580143
• 负责人: YOSHINAGA Arata
• 金额: $ 2.34万
• 依托单位: Kyoto University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2005
• 资助国家: 日本
• 起止时间: 2005 至 2007
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-17580143/
• 关键词: Tree cell wall; lignin; lignification; Immunolabeling; monoclonal antibody; 8-O-4 linkage; 8-5 linkage; 8-8 linkage; ヒノキ; 分化中木部; イチョウ; 8-0-4結合

To understand complex chemical structure of lignin in wood cell walls, monoclonal antibodies that recognize specific structure of lignin will be very important. The purpose of the present study is to develop the monoclonal antibody against lignin model compounds-protein conjugates. Three kinds of model compounds including three kinds of linkage (8-O-4, 8-5and8-8) of lignin were selected and conjugated with bovine serum albumin via p-aminohippuric acid. Monoclonal antibodies were raised with mice againstthree kinds of conjugates. As a result, 4hybridoma clones from 8-O-4 oligomer, 8 clones from 8-5 dimer and 8 clones from 8-8 dimer were developed.Enzyme-linked immunosorbent assay showed that the monoclonal antibody against8-O-4 oligomer conjugates did not react with p-aminohippuric acid-BSA conjugates and that the antibody against 8-5 and 8-8 dimers did not react with both p-aminohippuric acid-BSA conjugates and 8-O-4 dimer (guaiacyl glycerol-beta-guaiacyl ether)-p-aminohippuric acid-BSA conjugates. These results suggested that these monoclonal antibodies could recognize different linkage structure of lignin.Immunogold labeling of differentiating xylems of Japanese cypress, oak and poplar showed that the monoclonal antibody from at least one hybridoma clone from the three different antigens reacted with lignifying and lignified cell walls. Since the labeling sites were different among the three kinds of antigens, these monoclonal antibodies could recognize different chemical structure of lignin in wood cell walls. Although the specificity of these antibodies must be analyzed in detail, these monoclonal antibodies will be very effective probes to clarify complex structure of lignin and its formation process in wood cell walls.

为了了解木质细胞壁中木质素的复杂化学结构，识别木质素特定结构的单克隆抗体将非常重要。本研究的目的是开发针对木质素模型化合物蛋白结合物的单克隆抗体。选择了三种模型化合物，包括木质素的三种链接（8-O-4，8-5和8-8），并通过P-氨基磷酸与牛血清白蛋白共轭。用小鼠对三种结合物饲养单克隆抗体。结果，开发了来自8-O-4低聚物，8-5二聚体的8个克隆和8-8二聚体的8个克隆的4杂交瘤克隆。酶联免疫吸附测定表明，与8-O-4寡聚物的单克隆抗体与8-O-4寡聚物的8-氨基糖和8-氨基酸盐和8-氨基抗体 - 抗氨基甲基酸盐和8-抗酸性抗体 - 二聚体与p-氨基酸酸-BSA偶联物和8-O-4二聚体（鸟酰基甘油-beta-guaiacyl ether） - P-氨基硫酸盐酸-BSA偶联物没有反应。这些结果表明，这些单克隆抗体可以识别木质素的不同链接结构。日本柏树，橡木和杨体区分木质部的不含木质部标记表明，来自至少一种来自三种不同抗原的杂交抗体的单克隆抗体与木质抗原与木质化和木质化细胞壁反应。由于标记位点在三种抗原中有所不同，因此这些单克隆抗体可以识别木质细胞壁中木质素的不同化学结构。尽管必须详细分析这些抗体的特异性，但这些单克隆抗体将是非常有效的探针，以阐明木质素的复杂结构及其在木细胞壁中的形成过程。