人类β3-N-乙酰氨基葡萄糖基(β3半乳糖)转移酶β3GnT8/GalT7的抗体制备、免疫组化分析及其亚细胞定位  

Sub-cellular Localization and Immunochemistry Analysis of Human β3-N-acetylglucosaminyltransferase(β3GnT8)/β3-Galactosyltransferase (β3GalT7) by Preparation of β3GnT8 Antiserum

作　　者：周嘉梁[1,2] 姜智[1,3] 孙其昌[1] 王修珍[4] 仇灏[1,3] 吴士良[1,3] 

机构地区：[1]苏州大学医学部生物化学与分子生物学系,江苏苏州215123 [2]苏州大学附属第四医院肿瘤科,江苏无锡214062 [3]苏州大学生化工程研究所,江苏苏州215123 [4]苏州大学医学部病理学与病理生理学系,江苏苏州215123

出　　处：《苏州大学学报（医学版）》2009年第5期806-809,F0002,共5页Suzhou University Journal of Medical Science

基　　金：国家自然科学基金资助项目(30670462)

摘　　要：目的探讨β3GnT8(β3GalT7,AY277592,EC2.4.1.-)基因(人类β3-糖基转移酶家族中的一个新成员)的功能。方法原核表达该蛋白,以纯化的蛋白免疫新西兰大白兔,制备多克隆抗血清;以Westernblot和免疫组化分析其在多种组织细胞的表达情况;并通过荧光免疫细胞化学进行亚细胞定位。结果获得了较纯的表达蛋白和高效价、高特异性的抗血清,并且发现该蛋白大部分位于核膜周围的胞浆中;免疫组化分析显示肿瘤细胞和组织中该酶有较高的表达,与正常组织比较具有一定的差异性。结论该实验获得了β3GnT8/β3GalT7基因的蛋白质和相应抗体,并且对该蛋白进行了亚细胞定位,为对其生物学功能的进一步研究奠定了基础。Objective A novel member of human β3-glycosyltransferase family,the 3GnT8 gene （β3GalT7,AY277592,EC2.4.1.-）is first isolated and cloned by our lab. To make further study on the roles of this gene. Methods We constructed a prokaryotic expression system of 3GnT8 and obtained anti-3GnT8 polyclonal antiserum by immunizing rabbit with purified 3GnT8 protein. The expression of 3GnT8 protein in vatious tissues and cell lines was analyzed by Western blot and immunochemical as-says. The Sub-cellular location of 3GnT8 was detected using fluoroimmunocytochemistry. Results The rabbit antiserum was obtained and 3GnT8 was observed in cytoplasm around nuclear membrane. Immunochemistry analysis showed the enzyme was expressed significantly higher in some tumor tissues than in normal tissues. This revealed its biofunction in tumorogenisi. Conclusion The β3GnT8/ β3GalT7 protein and polyclonal antiserum prepared in this study has laid a foundation for further study of its biological function.

关 键 词：β3GnT8/β3GalT7 抗血清 免疫组化 亚细胞定位 

分 类 号：R730.231[医药卫生—肿瘤]