Development of cancer diagnosis using monoclonal antibodies specific for ras gene products

本研究旨在生产针对c-Ha-ras基因产物的抗体。这些抗体可用于癌症的诊断。为了制备c-Ha-ras蛋白，我们合成了编码正常和活化c-Ha-ras蛋白的基因片段，编码12-gly、12-val或61-lau。合成的基因通过表达载体连接到trp启动子的下游，在大肠杆菌中表达。通过离子交换层析和凝胶过滤对表达蛋白进行了定量分离。通过将这些蛋白注射到小鼠体内，并将其脾脏细胞与骨髓瘤细胞杂交，产生单克隆抗体。筛选并克隆抗体产生细胞。从识别c-Ha-ras （val-12）和c-Ha-ras （gly-12）的杂交瘤中获得的抗c-Ha-ras （val-12）。这种抗体将用于开发一种方法来检测过量产生c-Ha-ras蛋白的癌症。

The present invesigation intends a production of antibodies against products of c-Ha-ras genes. These antibodies can be applied to diagonosis of cancers. In order to prepare c-Ha-ras proteins, genes coding for normal and activated c-Ha-ras proteins were synthesized gene fragments coding for either 12-gly, 12-val or 61-lau. The synthetic genes were joined to downstream of the trp promoter in an expression vector and expressed in E.coli. The expressed proteins were isolated by ion-exchange chromatography and gel filtration in quantity. These proteins were used to produce monoclonal antibodies by injecting to mice and hybridization of their spleen cells with myeloma cells. Antibody producing cells were screened and cloned. Anti c-Ha-ras (val-12) obtained from a hybridoma recognized c-Ha-ras (val-12) as well as c-Ha-ras (gly-12). This antibody are to be used to develop a method to detect cancers overproducing c-Ha-ras proteins.