一种新的垂体分泌蛋白Mimecan原核表达、抗体制备及其在垂体瘤组织中的表达(英文)

Mimecan osteoglycin是一种分泌型蛋白质 ,目前其功能尚不明确 .从人垂体cDNA中克隆到OIF基因并构建成重组表达质粒pGEX 5X 2 mimecan ,将此重组表达质粒转化大肠杆菌BL2 1(DE3)后用IPTG诱导 ,成功表达了一种分子量约为 38kD融合蛋白 ,约占菌体总蛋白的 30 %～ 4 0 % .此融合蛋白经纯化分离后免疫新西兰大白兔以制备多克隆抗体 .用Western印迹法检测兔的抗血清 .结果显示 ,该多克隆抗体有较好的针对mimecan蛋白的专一性并且效价较高 ,可用于对mimecan的功能研究 .用此多克隆抗体检测到在某些种类的人垂体瘤组织中mimecan表达极高 ,提示可能存在新的垂体瘤类型 .

Antibody Preparation and Expression Analysis of a New Protein Mimecan in Pituitary Tumors

Mimecan belongs to a family of leucine rich proteoglycans that are secreted into the extracellular matrix. In order to investigate the function of mimecan, the coding region of mimecan was amplifed from a human pituitary cDNA by PCR and the recombinant prokaryotic expression vector pGEX M was constructed. The vector was transformed into E.coli BL21(DE3) and the GST M fusion protein of 38 kD was expressed in the bacteria under induction of IPTG. After purification, the fusion protein was injected into New Zealand rabbits to prepare polyclonal antibody. The antibody was tested by Western blotting for their specificity and sensitivity. Using the antibody it was found that mimecan was expressed highly in certain types of human pituitary tumor tissues. These results make it possible for studying the biological function of mimecan.