牛奶过敏原β-乳球蛋白抗体的制备及免疫检测方法的建立  

Preparation of Antibodies and Preliminary Establishment of Immunoassay Method against Allergen β-Lactoglobulin from Milk

作　　者：李亚璞[1] 麻云莲 樊丹 张小兵[1] LI Ya-pu;MA Yun-lian;FAN Dan;ZHANG Xiao-bing(Institute of Biology,Hebei Academy of Sciences,Shijiazhuang 050081,China;Institute of Chemical Technology,Hebei University of Technology,Tianjin 300132,China)

机构地区：[1]河北省科学院生物研究所,河北石家庄050081 [2]河北工业大学化工学院,天津300132

出　　处：《现代食品科技》2018年第10期225-230,共6页Modern Food Science and Technology

基　　金：河北省高层次人才资助项目(A2017002024)

摘　　要：本文将牛奶过敏原β-乳球蛋白抗原分别免疫新西兰大白兔和BALB/c小鼠制备其多克隆抗体和单克隆抗体,并对所获得的抗体进行了效价等特性测定;采用蛋白A亲和层析法将多克隆抗体进行了纯化,采用辛酸-硫酸铵法对单抗细胞株3A7进行了纯化;实验建立了双抗体夹心的ELISA检测方法,并对一些蛋白样品进行了初步的检测。实验结果表明,所制备的抗β-乳球蛋白兔多克隆抗体效价达到了1:6.56×106;Western-blotting鉴定结果显示所制备的多克隆抗体能够与β-乳球蛋白特异性反应;3株单抗的效价均在106以上,纯化后多抗仅与酪蛋白有一定的交叉反应,而纯化后单抗与乳中主要蛋白及其它过敏原蛋白基本没有交叉反应,特异性很好;利用所建立的方法对一些蛋白样品进行检测,准确率100%。The β-lactoglobulin antigen was used with injection to immunonize the New Zealand white rabbit and the BALB/c mice so as to get the rabbit polyclonal antibody serum （PcAb） and the monoclonal antibody （McAb）, and the titers of antibodies were analyzed; The polyclonal antibody was purified by protein A affinity chromatography and the 3A7 monoclonal antibody was purified by Caprylic acid precipitation. The experiment was operated by the double-antibody sandwich procedure and some samples were detected. The results showed that the titer of polyclonal antibody was over 1：6.56×10^6, and the polyclonal antibody serum could bind to β-lactoglobulin specifically, and its specificity was low reactivity with casein. The titers of the three strains of monoclonal antibodies were above 106 and the 3A7 monoclonal antibody had almost no cross reactivities with other proteins in milk and other allergen proteins. The accuracy was 100% for the detection of samples.

关 键 词：Β-乳球蛋白 酶联免疫吸附法 多克隆抗体 单克隆抗体 

分 类 号：TS252.7[轻工技术与工程—农产品加工及贮藏工程]