Studies on Diversity of Anti-TSH Receptor Antibody by Gene Technology.

利用基因技术研究抗TSH受体抗体多样性。

• 批准号: 63570546
• 负责人: ICHIKAWA Yoichi
• 金额: $ 1.34万
• 依托单位: Keio University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (C)
• 财政年份: 1988
• 资助国家: 日本
• 起止时间: 1988 至 1990
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-63570546/
• 关键词: TSH; TSH receptor; cDNA; EF-2; Graves' disease; monoclonal antibody; idiopathic hypothyroidism; TSH受容体; 抗TSH受容体抗体; 遺伝子工学; クローニング; 自己抗体

A cDNA library made of human thyroid carcinoma inserted to lamda gt 11 bacteriophage was cloned with anti-TSH receptor monoclonal antibodies. The cDNA clones obtained had the size of 1020 to 1900 bp and hybridized each other on Southern blott analysis.The fusion protein expressed with IPTG was used for the Western blott analysis. Sera of patients with Graves' disease or those of idiopathic hypothyroidism showed positive staining of the fusion protein, whereas the sera of normal subjects or patients with Hashimoto's disease did not. The cDNA was analyzed for the nucleotide sequence by dideoxy method. Computer search revealed that the cDNA showed strong homology with elongation factor 2(EF-2) of hamster. The reasons of unsuccessful cloning of TSH receptor clone were considered due to cDNA library synthesized with olige dT primer and the limited length (2 kb) of cDNA library.Another cDNA library made of normal human thyroid tissue with ramdom primer method was cloned with anti-TSH receptor monoclonal anti bodies. One of the cloned cDNA had the reading frame of about 2000 Kb, which was almost completely coincided with the nucleotide sequence of TSH receptor published recently. Two identical 4 amino acids sequences were found in EF-2 and TSH receptor, which might be the common epitopes of these proteins.

用抗TSH受体单克隆抗体克隆了人甲状腺癌λ gt 11噬菌体cDNA文库。获得大小为1020 ~ 1900 bp的cDNA克隆，经Southern blotting杂交，IPTG诱导表达的融合蛋白用于Western blotting分析。Graves病和特发性甲状腺功能减退症患者的血清显示融合蛋白的阳性染色，而正常人或桥本病患者的血清则没有。用双脱氧法分析其核苷酸序列。计算机检索表明，该cDNA与金黄地鼠的延伸因子2（EF-2）有很强的同源性。考虑到用寡聚dT引物合成的cDNA文库及cDNA文库的长度（2kb）限制性，不能成功克隆TSH受体克隆，用抗TSH受体单克隆抗体，用随机引物法制备了正常人甲状腺组织cDNA文库。其中一个cDNA的阅读框约为2000 Kb，与最近发表的TSH受体的核苷酸序列几乎完全一致。在EF-2和TSH受体中发现了两个相同的4个氨基酸序列，可能是这两种蛋白的共同表位。

项目成果

市川 陽一: "作用物貭別レセプタ-論,TSH" 日本臨床 増刊レセプタ-. 47. 197-200 (1989)

Yoichi Ichikawa：“作用辨别受体理论，TSH”日本临床受体特别版 47. 197-200 (1989)。

Ichikawa Y.: "Antiーthgrotropic hormore receptor antibody and thgroid direase" Japanese J Medicine. 27. 217-219 (1988)

Ichikawa Y.：“抗促甲状腺激素受体抗体和促甲状腺疾病”日本医学杂志 27. 217-219 (1988)

Ichikawa Y: "Antiーthyrotropic hormone receptor antibody and thyroid disease" Jap J Medicine. 27. 217-219 (1988)

Ichikawa Y：“抗促甲状腺激素受体抗体和甲状腺疾病”Jap J Medicine 27. 217-219 (1988)。

市川 陽一: "作用物値別レセプタ-論、TSH" 日本臨床 増刊レセプタ-. 47. 197-200 (1989)

Yoichi Ichikawa：“效应值的受体理论，TSH”日本临床特刊受体。 47. 197-200 (1989)

市川陽一,松下庸次: "作用物質別レセプタ-論:TSH" 日本臨床. 47(増刊). 197-200 (1989)

Yoichi Ichikawa、Yoji Matsushita：“活性物质的受体理论：TSH”日本临床杂志 47（特刊）。