摘要
为研究sIgM在吉富罗非鱼体内的组织分布及其在亚细胞水平上的定位,本实验利用纯化的SIGM融合蛋白,按照常规方法免疫新西兰大白兔,制备了兔抗吉富罗非鱼SIGM多克隆抗体;并对吉富罗非鱼肠、脾脏及鳃组织进行了免疫组织化学分析以及胶体金免疫电镜分析。结果显示,ELISA检测所获得的抗血清效价为1∶256 000,该血清能与SIGM融合蛋白发生特异性免疫反应。在肠和鳃组织中,阳性信号存在于富含黏液细胞的上皮细胞层表面,在杯状细胞和黏液细胞中则不存在;在脾脏组织中,阳性信号存在于特定的细胞中。免疫电镜结果显示SIGM主要存在于肠上皮细胞膜附近,并且在肠组织上皮细胞膜表面的微绒毛处含量较多;在鳃上皮组织中,SIGM主要存在于包围鳃丝和鳃小片的上皮细胞及部分红细胞内;而在脾脏组织淋巴细胞内部高尔基体的分泌小泡处存在大量的胶体金颗粒。研究表明,sIgM在鱼体黏膜免疫中具有重要作用,这为探讨鱼类sIgM的特性及功能奠定了基础,并丰富了鱼类黏膜免疫的研究内容。
In order to determine the distribution of sIgM in immune organs and the location of SIGM at the subcellular level in Nile tilapia,polyclonal antibody against SIGM fusion protein was prepared in a New Zealand pedigree white rabbit immunized with the purified SIGM fusion protein. For further study on the location of SIGM in the intestine,gill and spleen in Oreochromis niloticus,immunohistochemistry analysis and the immunoelectron microscopy technique were carried out using the prepared polyclonal antibody. The result of the Western blotting revealed that specific antigen-antibody reaction occurred between the antiserum and its corresponding recombinant fusion protein and the antibody titer reached 1∶ 256 000. In the intestine and gill tissues,the positive signals existed in the epithelial cell surface,but not existed in the goblet cells and the mucous cells; while in spleen tissues,the SIGM protein was mainly distributed in some cells in this organ. The immunoelectron microscopy results showed that SIGM mainly existed in the vicinity of intestinal epithelial cell membrane and the surface membranes of the epithelial cells in intestinal microvilli. In the gill epithelial tissue,SIGM mainly existed in the part of the red blood cells and the epithelial cells which surrounded the gill filaments and gill-amella. In the spleen tissue,a large number of colloidal gold particles existed in the secretory vesicles at the internal Golgi apparatus. All these results suggested that sIgM played a critical role in the muscosal immunity of teleost. These provide insights into the roles of fish sIgM in the mucosal immunity.
出处
《水产学报》
CAS
CSCD
北大核心
2014年第9期1211-1220,共10页
Journal of Fisheries of China
基金
国家自然科学基金青年基金(31302226)
国家科技支撑计划(2012BAD17B02
2012BAD17B03)
广东省科技计划农业攻关项目(2012B020308009)
广东省2012年鱼病防治专项(2130108)
关键词
吉富罗非鱼
SIGM
多克隆抗体
组织定位
GIFT strain of Nile tilapia
SIGM
polyclonal antibody
tissue localization
作者简介
通信作者：简纪常，E-mail：jianjc@gmail．com
