环丙沙星、恩诺沙星、磺胺二甲嘧啶串联单链抗体的制备及融合表达产物抗原特性的分析

目的建立免疫学快速检测动物制品中的环丙沙星、恩诺沙星、磺胺二甲嘧啶残留,制备融合表达蛋白并进行评价。方法利用传统单克隆抗体制备方法分别制备环丙沙星、恩诺沙星、磺胺二甲嘧啶的单克隆抗体,通过聚合酶链式反应(polymerasechainreaction,PCR)获得3种单抗的单链抗体(single-chainvariable fragment,Sc-Fv),再通过套嵌PCR方法,使用柔性氨基酸Linker将已获得的单链抗体进行串联并通过分子生物学技术获得高效融合表达,以纯化的多联融合单链抗体为探针,结合前期技术建立的免疫学快速检测技术,对融合表达产物的抗原特性进行初步的分析。结果制备的单克隆抗体抗原抗体反应性良好,效价较高,融合表达抗体蛋白能够很好的与3种相应药物反应。结论本研究初步建立了特异性较高的3种药物的融合表达抗体的制备方法,且该抗体具有良好的反应原性。

Preparation of ciprofloxacin, enrofloxacin and sulfadimidine tandem single-chain antibodies and analysis of the antigens characteristics of the fusion expressed products

Objective To establish an immunological rapid detection method for ciprofloxacin, enrofloxacin and sulfadimidine residues in animal products, and to prepare the fusion expression proteins and evaluate them. Methods The ciprofloxacin, enrofloxacin and sulfadimidine monoclonal antibodies were prepared by traditional monoclonal antibody preparation methods respectively, and 3 kinds of single-chain variable fragment (Sc-Fv) were obtained by polymerase chain reaction (PCR), then by a nested PCR method, the obtained single-chain variable fragment were connected in series using a flexible amino acid linker and high-efficiency fusion expression was obtained by using a molecular biology technology. The purified multi-linked fusion Sc-Fv was taken as a probe, and the antigenic characteristics of a fusion expression product were preliminarily analyzed in combination with an immunological rapid detection technology established by a previous technology. Results The monoclonal antibodies were prepared with good antigenic antibody reactivity, high titer and fusion expression of antibody protein had good reaction with 3 corresponding drugs. Conclusion This paper establish the preparation method of the specific high-specificity fusion expression of antibody protein of 3 drugs, and the antibody has good reactivity.