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〔目的〕研究一种相思子毒素抗原、抗体的制备技术。〔方法〕根据相思子毒素对半乳糖的特异性结合能力,采用亲和层析技术将相思子种子中大部分杂质除去;经凝胶过滤层析,进一步去除植物种子中与蛋白毒素并存的凝集素,获得电泳纯度天然相思子毒素纯品。〔结果〕利用大肠埃希菌表达重组相思子毒素A链蛋白,经金属螯合层析法纯化,得到重组相思子毒素A链蛋白抗原。重组抗原免疫大耳白兔、昆明小鼠,采用辛酸-硫酸铵盐析法和亲和层析法纯化兔血清和鼠腹水获得多抗;2种抗体与天然毒素特异性结合的活性效价均为106。〔结论〕抗原、抗体的制备为建立相思子毒素检测方法奠定了基础。
[Objective] To study the preparation technology of a kind of acacia seed antigens and antibodies. [Method] According to the specific binding capacity of acacia to galactose, most of the impurities in the seeds of Acacia seed were removed by affinity chromatography. By gel filtration chromatography, agglutination of plant seeds with protein toxin was further removed Prime, pure electrophoresis purity of natural Acacia seed pure. [Results] The recombinant Aciton toxin A chain protein was expressed in Escherichia coli and purified by metal chelate chromatography to obtain a recombinant Aciton toxin A chain protein antigen. Recombinant antigens were used to immunize rabbits and Kunming mice, and the polyclonal antibodies were purified from rabbit serum and mouse ascites by caprylic acid-ammonium sulfate salting-out and affinity chromatography. The activity titers of specific binding of the two antibodies to natural toxins were 106. [Conclusion] The preparation of antigens and antibodies laid the foundation for the establishment of the method for the determination of acacia toxin.