肿瘤相关糖抗原sTn的提纯及其单克隆抗体的制备

提纯肿瘤相关糖抗原sTn并研制针对它的单克隆抗体(mAb).取羊颌下腺经溴棕三甲铵沉淀,经Sepharose CL-4B等获取羊颌下腺粘蛋白(OSM),将OSM采用脾内注射、完全佐剂2种方法免疫Balb/C小鼠.常规方法细胞融合,用ELISA间接法筛选阳性细胞并测定效价,双抗体竞争抑制试验识别sTn表位,采用免疫组化检测sTn在胃癌组织中的表达.最终获得2株可分泌sTn mAb细胞株(2F6,7E4),均为小鼠IgG1亚类,2株mAb抗相同抗原表位.免疫组化染色显示胃癌组织中sTn表达阳性率较高,可达71.4%,且与胃癌进程相关.本文成功提纯了sTn抗原并制备了2株杂交瘤细胞株,在胃癌的诊断和治疗中有潜在的应用价值.

Purification of Tumor-associated antigen sTn and Preparation of Monocional Antibodies against sTn

To purify tumor-associated antigen sTn and prepare monoclonal antibody against it,o- vine submaxillary mucin(OSM) after precipitation by C_(19)H_(42)NBr was purified and concentrated by a Sepharose CL-4B column.OSM was used as the antigen to immunize Balb/C mice in two different ways,direct spleen injection and the routine immunization.After cell fusion,indirect ELISA was used to screen positive ceils and determine the titre of the antibodies.Double anti- body competitive inhibition experiment was used to identify the sTn epitope.Immuno-histo- chemical analysis was employed to locate sTn's expression in gastric carcinoma tissues.The re- sults showed that two strains of monoclonal antibodies (2F6 and 7E4) were obtained.Both of them belong to the IgG1 subclass and target to sTn epitope in OSM.Immuno-histochemical an- alyses showed that sTn were widely expressed in gastric carcinoma tissues,with a positive rate of 71.4%.The high positive rates of sTn are closely related to the stages of the disease.The results suggested that two strains of monoclonal antibodies against sTn were successfully pre- pared;they could possibly be used in the diagnosis and treatment of gastric cancer.