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Detection of Novel Amyloid-Associated Protein by Monoclonal Antibody and its Gene Analysis

新型淀粉样蛋白相关蛋白的单克隆抗体检测及其基因分析

基本信息

批准号：
01570363
负责人：
IMAI Kohzoh
金额：
$ 1.41万
依托单位：
Sapporo Medical College
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (C)
财政年份：
1989
资助国家：
日本
起止时间：
1989 至 1990
项目状态：
已结题

项目摘要

We have found that monoclonal antibody AM34 (IgGl) recognizes a novel antigen which exists in amyloid deposits in various organs of the rheumatoid arthritis.In addition, anti-beta-protein monoclonal antibodies (TB1, TB2, TB3, TB4 and TB5) were generated from the fusion of mouse myeloma X63-Ag8.653 cells with splenocytes obtained from a BALB/c mouse that had been immunized with a synthetic peptide of beta-protein consisting of 24 amino acids. All these antibodies bound specifically to the amyloid changes of senile plaque and amyloid angiopathy of the brain tissues of patients with Alzheimer's disease (AD) or with senile dementia Alzheimer type (SDAT) , but not at all to protein in the normal brain or other tissues by the immunoperoxidase method. Formic acid pretreatment of brain tissue enhanced this reactivity. Positive reactions were clearly visible even in small deposits of plaque, senile plaque crown amyloid and amyloid deposits of capillary vessels under this condition.Western blot analysis revealed that monoclonal antibodies TB2 and TB3 reacted mainly with 116 kD band when extract of SDAT brain was used. They failed to react, however, with 116kD band when extract of normal brain derived from patients with other diseases was used. Two dimensional electrophoresis revealed that the 116 kD band migrated at around pl8.5. MoAbs TB1, TB2 and TB4 strongly reacted with the 1st part (7 amino acids near to N-terminus) of the synthetic peptide (24 amino acids) of the beta-protein, which consists of hydrophilic amino acids.

我们已经发现单克隆抗体AM34（IgGl）识别存在于类风湿性关节炎的各种器官中的淀粉样蛋白沉积物中的新抗原。（TB1，TB2，TB3，TB 4和TB 5）由小鼠骨髓瘤X63-Ag8.653细胞与从BALB/c小鼠获得的脾细胞的融合产生。c用由24个氨基酸组成的β-蛋白的合成肽免疫的小鼠。所有这些抗体特异性结合阿尔茨海默病（AD）或阿尔茨海默型老年性痴呆（SDAT）患者脑组织的老年斑和淀粉样血管病的淀粉样改变，但通过免疫过氧化物酶方法，这些抗体完全不结合正常脑或其它组织中的蛋白质。甲酸预处理的脑组织增强了这种反应性。Western blot分析表明，单克隆抗体TB 2和TB 3在SDAT脑提取液中主要与116 kD条带反应。但用其他疾病患者的正常脑组织提取液作对照时，116 kD区带无反应。双向电泳显示116 kD条带在p18.5左右迁移。单克隆抗体TB 1、TB 2和TB 4与β-蛋白合成肽（24个氨基酸）的第1部分（靠近N-末端的7个氨基酸）发生强烈反应，所述合成肽由亲水性氨基酸组成。