花生球蛋白鼠源多克隆抗体的制备及免疫学特性鉴定Preparation of murine polyclonal antibody against arachin and identification of its immunological characteristics

为建立花生球蛋白致敏原的免疫学快速检测方法,采用碱溶酸沉法提取花生球蛋白，经SDS-PAGE检测蛋白纯度后，免疫5只6~8周龄的Balb/c雌性小鼠制备花生球蛋白鼠源多克隆抗体（多抗）血清，并进行免疫学特性鉴定。结果表明：免疫结束后得到的5个多抗血清效价均在1∶ 51 200 以上，均具有一定的敏感性，其中2号小鼠的多抗血清敏感性较好，半数抑制浓度为320 ng/mL；制备的多抗血清特异性较强，2号小鼠的多抗血清与伴花生球蛋白、大豆球蛋白、β-伴大豆球蛋白、牛血清白蛋白、麦朊蛋白、乳清蛋白的交叉反应率均较低，在0.5%以下；Western blot鉴定结果表明成功制备出了花生球蛋白鼠源多克隆抗体。综上，获得了效价高、敏感性强、特异性优良的花生球蛋白鼠源多克隆抗体，为其单克隆抗体的制备及免疫学快速检测方法研究奠定了基础。 In order to establish a rapid immunological detection method for the arachin allergens,the arachin was extracted by alkali solution and acid precipitation method, and its purity was detemined by SDS-PAGE, and 5 Balb/c female mice aged 6 to 8 weeks were immunized to prepare the arachin mouse-derived polyclonal antiserum,the polyclonal antiserum immunological characteristics were identified. The results showed that the titers of the polyclonal antiserum obtained after the end of immunization were all above 1∶ 51 200, and all of them had certain sensitivities, among which the polyclonal antiserum from 2# mouse had the best sensitivities, with a half-inhibitory concentration of 320 ng/mL.The polyclonal antiserum from 2# mouse was not sensitive to conarachin, glycinin, β-conglycinin, bovine serum albumin, gluten protein, and skimmed milk powder,and the cross-reaction rate was low(all below 05%),indicating that the specificity of the polyclonal antiserum prepared was relatively strong. Western blot experiment results indicated the successful preparation of arachin mouse-derived polyclonal antiserum. In conclusion, a arachin mouse-derived polyclonal antibody with high titer, strong sensitivity and excellent specificity is obtained, which provide the material foundation for the preparation of monoclonal antibodies and the research of rapid immunological detection methods.