鲤鱼小清蛋白的抗原性研究
发布时间：2019-04-25 17:58
【摘要】： 鱼类食物是人类获取优质蛋白的重要途径,但有部分人对鱼类蛋白过敏,健康受到极大威胁。有研究表明,鲤鱼和鳕鱼等鱼类中主要过敏原是小清蛋白,因而研究鲤鱼小清蛋白的抗原性将有助于预防食物过敏和生产低致敏性产品。 本研究的主要内容包括鲤鱼小清蛋白的分离纯化,兔抗鲤鱼小清蛋白多克隆抗体的制备及青鱼、草鱼、鲢鱼蛋白的粗提物与鲤鱼小清蛋白之间免疫交叉反应的研究。 通过实验,建立了以磷酸盐缓冲液粗提结合DEAE-Sepharose Fast Flow离子交换层析分离纯化鲤鱼小清蛋白的方法;SDS-PAGE和PAGE鉴定结果表明,用这种方法可以获得纯度＞90%的鲤鱼小清蛋白样品。 本实验建立了间接ELISA检测兔抗鲤鱼小清蛋白抗血清效价的检测方法及制备兔抗鲤鱼小清蛋白多克隆抗体的方法。免疫两只日本雄性大白兔分别获得了间接ELISA检测效价高达1:560,000和1:720,000的抗血清。结合间接竞争ELISA实验对鲤鱼小清蛋白的抗原性进行了分析和评价,研究结果表明,鲤鱼小清蛋白具有很好的抗原性。 通过免疫印迹和间接竞争ELISA实验,证实了鲤鱼小清蛋白和青鱼、草鱼、鲢鱼粗提蛋白之间存在免疫交叉反应,发生交叉反应的主要蛋白分子量约为12kDa。
[Abstract]:Fish food is an important way for human to obtain high-quality protein, but some people are allergic to fish protein, and their health is seriously threatened. Studies have shown that the major allergen in fish such as carp and cod is small white protein, so studying the antigenicity of carp small albumin will help to prevent food allergy and produce hypoallergenic products. The main contents of this study include the isolation and purification of carp albumin, the preparation of rabbit anti-carp albumin polyclonal antibody, and the study of immune cross-reaction between the crude extracts of carp, grass carp and silver carp proteins and carp albumin. A method for the separation and purification of carp small white protein with phosphate buffer solution and DEAE-Sepharose Fast Flow ion exchange chromatography was established. The results of SDS-PAGE and PAGE showed that the purity of carp small white protein was more than 90% by this method. An indirect ELISA method for the detection of rabbit anti-carp serum titer and preparation of rabbit anti-carp albumin polyclonal antibody was established. Two Japanese male rabbits were immunized with anti-ELISA sera with the titers of 1? 560 000 and 1? 720 000 respectively. The antigenicity of carp small white protein was analyzed and evaluated by indirect competitive ELISA assay. The results showed that carp small white protein had good antigenicity. By means of immunoblotting and indirect competitive ELISA test, it was confirmed that there was an immune cross-reaction between carp albumin and the crude protein extracted from Cyprinus Carpio, grass carp and silver carp. The molecular weight of the main cross-reaction protein was about 12kda.
【学位授予单位】：南昌大学
【学位级别】：硕士
【学位授予年份】：2008
【分类号】：R392
本文编号：2465339
[Abstract]:Fish food is an important way for human to obtain high-quality protein, but some people are allergic to fish protein, and their health is seriously threatened. Studies have shown that the major allergen in fish such as carp and cod is small white protein, so studying the antigenicity of carp small albumin will help to prevent food allergy and produce hypoallergenic products. The main contents of this study include the isolation and purification of carp albumin, the preparation of rabbit anti-carp albumin polyclonal antibody, and the study of immune cross-reaction between the crude extracts of carp, grass carp and silver carp proteins and carp albumin. A method for the separation and purification of carp small white protein with phosphate buffer solution and DEAE-Sepharose Fast Flow ion exchange chromatography was established. The results of SDS-PAGE and PAGE showed that the purity of carp small white protein was more than 90% by this method. An indirect ELISA method for the detection of rabbit anti-carp serum titer and preparation of rabbit anti-carp albumin polyclonal antibody was established. Two Japanese male rabbits were immunized with anti-ELISA sera with the titers of 1? 560 000 and 1? 720 000 respectively. The antigenicity of carp small white protein was analyzed and evaluated by indirect competitive ELISA assay. The results showed that carp small white protein had good antigenicity. By means of immunoblotting and indirect competitive ELISA test, it was confirmed that there was an immune cross-reaction between carp albumin and the crude protein extracted from Cyprinus Carpio, grass carp and silver carp. The molecular weight of the main cross-reaction protein was about 12kda.
【学位授予单位】：南昌大学
【学位级别】：硕士
【学位授予年份】：2008
【分类号】：R392
【引证文献】
相关期刊论文 前1条
1 李阳;刘楚怡;薛文通;;响应面优化鳜鱼致敏蛋白的木瓜蛋白酶水解工艺[J];食品科学;2012年19期
相关硕士学位论文 前1条
1 沈海旺;章鱼过敏原的研究[D];集美大学;2012年
，本文编号：2465339
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