人IL-37在大肠杆菌表达及多克隆抗体的制备与鉴定  

The expression of human IL-37 in E. coli and preparation and characterization of mouse anti-IL-37 antibody

作　　者：赵付前[1] 何素辉[1] 何玲鸽[1] 林妙芬[1] 王森[1] 陈章权[1,2] 

机构地区：[1]广东医学院广东省医学分子诊断重点实验室,广东东莞523808 [2]广东医学院临床免疫学教研室,广东东莞523808

出　　处：《细胞与分子免疫学杂志》2014年第5期513-516,共4页Chinese Journal of Cellular and Molecular Immunology

基　　金：广东省自然科学基金(S2012040006383);广东省大学生创新训练计划(1057113038)

摘　　要：目的原核表达白细胞介素37(IL-37)及制备其多克隆抗体。方法 PCR扩增IL-37b成熟肽编码区基因,克隆至表达载体pET28a,转化大肠杆菌感受态细胞,IPTG诱导表达,Ni2+-NTA琼脂糖凝胶柱亲和层析纯化重组蛋白,以纯化的重组蛋白IL-37为免疫原免疫BALB/c小鼠制备其特异性抗体,用ELISA、Western blot法和免疫组织化学染色检测抗体的效价和特异性。结果原核表达了重组蛋白IL-37b成熟肽,并获得高效价的小鼠抗IL-37抗体,能特异性识别天然的IL-37抗原。结论成功制备效价高、特异性好的小鼠抗IL-37抗体。Objective To construct a prokaryotic expression system for interleukin-37 （IL-37） and prepare its polyclonal antibody. Methods The gene encoding mature interleukin-37 （IL-37m） was amplified by PCR and subcloned into prokaryotic expression vector pET28a. Then the recombinant plasmid pET28a/IL-37m was transformed into E. coli Rosetta and expressed under IPTG induction. The recombinant IL-37m was purified through Ni^2＋ -NT agarose gel column and the purified recombinant IL-37m was used as immunogen to immunize the BALB/c mouse. The titer and specificity of the mouse anti-lL-37 antibody were analyzed by ELISA, Western blotting and immunohistochemical staining, respectively. Results The recombinant IL-37 was successfully expressed and purified, and the mouse anit-lL-37 antibody was successfully prepared. ELISA showed that the titer of the antiserum was 1：128 000. Western blot analysis revealed that the antibody reacted with IL-37 specifically. Immunohistochemical staining detection manifested the antibody could recognize the native IL-37. Conclusion The mouse anti-lL-37 antibody with high titer and specificity was successfully prepared.

关 键 词：白细胞介素37 表达 抗体 制备 小鼠 

分 类 号：R392.11[医药卫生—免疫学]