Molecular pathological research of gene abnormalities using monoclonal antibodies

多种单抗(MoAbbs)被广泛用于常规病理诊断。然而，针对癌基因的MOAD是一种新的工具，其特异性和有效性仍然是相反的。虽然针对c-Ha-ras蛋白p21的MoAbs已报道对癌细胞呈特异性阳性，但我们通过Western印迹方法证实这些MoAbs与癌细胞和正常细胞均呈阳性反应。免疫组织化学染色与Northern印迹法检测到的信使RNA的表达也证实了平行关系。在另一个项目中，本实验室制备了与v-ABL蛋白序列部分对应的合成寡肽的MoAbs，并通过电子显微镜显示ABL-蛋白产物定位于细胞膜上。人肾鳞状细胞癌或肾细胞癌免疫后也可产生MoAbs。B-11单抗检测到34kd蛋白并与鳞状细胞癌和移行细胞癌…反应更多的S，但不是腺细胞癌。随着膀胱癌恶性程度的增加，B-11的阳性表达逐渐降低。采用非同位素标记DNA探针进行原位杂交，对方法进行了改进。将5‘-溴脱氧尿嘧啶核苷(Brdud)连接到DNA中，与BrdUrd标记的DNA探针杂交后，用FITC标记的单抗进行检测。本文分析了日本非胰岛素依赖型糖尿病患者胰岛素基因5‘-侧翼区的多态性，发现5’-侧翼插入频率低于非日本人群。我们还分析了福尔马林固定对DNA降解的机理，以便将固定的样本用于胰岛素或其他基因的分析。低温下使用缓冲福尔马林和EDTA可以阻断DNA的降解。为了进一步了解抗癌基因和其他基因产物的MoAbs对细胞过程的分子病理学研究的有效性和特异性，有必要进行进一步的研究。较少

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