Molecular pathological research of gene abnormalities using monoclonal antibodies

使用单克隆抗体进行基因异常的分子病理学研究

• 批准号: 61480138
• 负责人: SUGIYAMA Taketoshi
• 金额: $ 3.71万
• 依托单位: Kobe University School of Medicine
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (B)
• 财政年份: 1986
• 资助国家: 日本
• 起止时间: 1986 至 1988
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-61480138/
• 关键词: monoclonal antibody; gene product; pathology; ras; abl; B-11; insulin; 分子異常; 組織化学; ホルモン; 甲状腺癌

Many kinds of monoclonal antibodies(MoAbs) are widely used for routine pathological diagnosis. However, MoAds against oncogenes are new tools and their specicicity and usefulness are still contro-versial. Although MoAbs against c-Ha-ras protein p21 had been reported to be specifically positive for cancer cells, we confirmed those MoAbs were positively reacted with both cancer and normal cells by the western blot method. Parallel relationship was also confirmed between the immuno-histochemical staining and the expression in messenger RNA detected by northern blot method. In another project, MoAbs against synthetic oligopeptides corresponding partially to v-abl protein sequence were generated in our laboratory and the abl-protein products were shown to localize on cell membrane by electron microscope. MoAbs were also generated by the immunization of human squamous cell or renal cell carci-nomas. B-11 MoAb detected 34kd protein and was reacted with squamous and transitional cell carconoma … More s but not with glandular cell carcinomas. When the grade of malignance of human bladder cancer increased, the positivity for B-11 decreased. Methodological improvement was made by using non-isotope labeled DNA probe for in situ hybridization. 5'-bromodeoxyuridine(brdurd) was incorporated into plasmid DNA by ino-culating E.coli and after hybridization with BrdUrd labeled DNA probe, the hybridized probe was detected by FITC-conjugated monoclonal antibody. Polymorphism in the 5'-flanking region of the human insulin gene in Japanese non-insulin-dependent diabetic subjects was analyzed and lower frequency of 5'-flanking in-sertion was observed than in non-Japanese reaces. We also analysed the mechanism of DNA degradation with formalin fixation in order to use the fixed samples for analysis of insulin or other genes. Use of buffered formalin at low temperature and EDTA were shown to block DNA degradation.Further study is necessary to understand the usefulness and specificity of MoAbs against oncogenes and other genes products for molecular pathological studies of cellular process. Less

多种单克隆抗体广泛应用于常规病理诊断。然而，抗癌基因的MoAds是一种新的工具，其特异性和有效性仍存在争议。虽然抗c-Ha-ras蛋白p21的单克隆抗体已被报道对癌细胞呈特异性阳性，但我们通过western blot方法证实这些单克隆抗体对癌细胞和正常细胞均呈阳性反应。免疫组化染色结果与北方印迹法检测的mRNA表达量之间存在平行关系。在另一个项目中，我们实验室制备了针对与v-abl蛋白序列部分对应的合成寡肽的单克隆抗体，并通过电子显微镜显示abl蛋白产物定位于细胞膜上。单克隆抗体也通过免疫人鳞状细胞癌或肾细胞癌产生。B-11单抗检测34 kd蛋白，与鳞状细胞癌和移行细胞癌反应 ...更多信息 但与腺细胞癌无关。随着膀胱癌恶性程度的增加，B-11的阳性率逐渐降低。采用非同位素标记的DNA探针进行原位杂交。将5 ′-溴脱氧尿苷（BrdUrd）掺入质粒DNA中，与BrdUrd标记的DNA探针杂交后，用FITC标记的单克隆抗体进行检测。分析了日本非胰岛素依赖型糖尿病受试者中人胰岛素基因5 '侧翼区的多态性，观察到5'侧翼插入频率低于非日本受试者。我们还分析了福尔马林固定的DNA降解机制，以便将固定的样品用于胰岛素或其他基因的分析。低温福尔马林缓冲液和EDTA可阻断DNA的降解，进一步研究MoAbs对癌基因和其他基因产物的特异性和有效性对细胞过程的分子病理学研究有重要意义。少

项目成果

青山伸郎: 病理と臨床. 5(2). 165-172 (1987)

青山伸夫：病理学和临床研究 5(2)（1987）。

杉山武敏;前田盛;北沢荘平: 臨床検査. 31. 1389-1393 (1987)

Taketoshi Sugiyama；Mori Maeda；Shohei Kitazawa：临床检查。 31. 1389-1393 (1987)

J.Ishikawa; S.Maeda; T.Sugiyama; S.Kamidono.: "Restriction fragment length polymorphism and activation of c-Ha-ras gene in urothelial cancer." Anticancer Research.

J.石川;

荒樋栄宣: 神戸大学医学部紀要. 43. 15-25 (1988)

荒日秀信：神户大学医学院通报。43. 15-25 (1988)。

J.Ishikawa; R.Nishimura; S.Maeda; G.Hamami; T.Sugiyama; S.Kamidono.: "Primary choriocarcinoma of the urinary bladder." Acta Pathol. Jpn.

J.石川;