以CpG为佐剂人IL-24多克隆抗体制备.docVIP

以CpG为佐剂人IL-24多克隆抗体制备【摘要】目的:采用在原核表达系统表达的人 IL-24蛋白及人IL-24真核重组质粒免疫新西兰兔,制备兔抗人IL-24多克隆抗体#65377;方法:利用IPTG诱导hIL-24在大肠杆菌中表达,纯化hIL-24重组蛋白,纯化后的蛋白经SDS分析,同时提取hIL-24真核重组质粒,用以免疫新西兰兔,并以CpG为佐剂制备多克隆抗体#65377;Western-blot 鉴定抗体的特异性,ELISA法测定抗体效价#65377;结果:人IL-24经IPTG诱导后可在大肠杆菌中大量表达,表达量占细菌总蛋白的30%,纯化后的蛋白纯度高;原核表达的重组蛋白和真核重组质粒免疫新西兰兔,通过抗体效价测定,获得了抗血清效价达1∶640的多克隆抗体#65377;结论:原核表达的hIL-24蛋白和hIL-24真核重组质粒均能刺激家兔产生抗体,其多克隆抗体的效价较高#65377; 【关键词】人IL-24;原核表达;真核重组质粒;多克隆抗体;CpG 文章编号:1009-5519(2007)11-1583-03 中图分类号:R33 文献标识码:A CpG-ODN is an adjuvant for preparation of the hIL-24 polyclonal antibody HUANG Jun-qiong,SHENG Wei-hua,XIE Yu-feng,et al. (Department of Cell and Molecular Biology,Medical School, Suzhou University,Suzhou 215123,China) 【Abstract】Objective:To immunize rabbits with hIL-24 protein expressed in E.coli and eukaryotic recombinant plasmid to prepare hIL-24 polyclonal antibodies.Methods:The recombinant protein was achieved in E.coli through IPTG induction and was purified. The protein was analyzed by SDS. Rabbits were immunized with hIL-24 protein and eukaryotic recombinant plasmid to prepare hIL-24 polyclonal antibodies.CpG-ODN was used as an adjuvant for preparation of the hIL-24 antibody.The antibodies were detected by western-blot and ELISA.Results:Human IL-24 could be expressed in E.coli after IPTG induction. The recombinant protein was about 30% of total expressed products in E.coli. Purified protein was achieved. The polyclonal antibodies were successfully prepared and had higher titter (1:640). Conclusion:HIL-24 recombinant protein expressed in E.coli and eukaryotic recombinant plasmid can stimulate antibody in rabbits. 【Key words】Human IL-24;Prokaryotic expression;Eukaryotic recombinant plasmid;Polyclonal antibody;CpG 白细胞介素-24(Interleukin-24,IL-24)是Jiang等于1995年利用减数杂交技术从β-干扰素(IFN-β)和蛋白激酶激活剂MEZ诱导分化的人恶性黑色素瘤细胞HO-1中克隆的新基因[1]#65377;人IL-24具有广谱的抗肿瘤活性,通过诱导肿瘤细胞凋亡#65380;抑制血管生成#65380;促进细胞因子分泌而增强抗肿瘤免疫应答等多种机制抑制多种组织来源的肿瘤细胞生长[2~4]#65377;本室构建了高效原核表达载体pET-21a(+)-hIL-24#65377;本实验利用IPT