CD40单克隆抗体的制备、筛选及功能鉴定

发布时间：2018-02-01 05:11

本文关键词： CD 单克隆抗体筛选功能鉴定　出处：《现代免疫学》2017年05期 　论文类型：期刊论文

【摘要】：CD40是肿瘤坏死因子受体超家族成员之一,属于Ⅰ型跨膜糖蛋白,其自然配体是CD40L。CD40-CD40L信号通路在许多重要的生理和病理过程中都起到关键的调节作用。基于其在激活免疫应答中的重要作用,在肿瘤免疫治疗中,具有激活CD40-CD40L信号的CD40激动剂抗体可用于提高抗肿瘤免疫应答;在移植排斥和炎症反应中,CD40-CD40L信号也起重要作用,阻断CD40信号通路是目前治疗风湿性关节炎、肠炎、系统性红斑狼疮、多发性硬化和糖尿病的一个重要靶点。制备CD40阻断性抗体或激动剂抗体不仅可用于检测CD40分子表达,还可以开发用于炎症性疾病或肿瘤治疗的免疫疗法。本研究表达CD40胞外段蛋白,通过免疫制备14株CD40单克隆抗体,筛选发现全部可作为western blotting检测CD40蛋白表达的抗体,9株CD40抗体可作为一抗用于流式细胞术检测人CD40表达,1株CD40抗体能阻断人CD40L和CD40相互作用。有意义的是,8株CD40抗体能激活人CD40分子信号通路,另外有10株能激活小鼠脾脏细胞CD40下游基因的表达,7株能上调小鼠脾脏B细胞表面共刺激分子CD86的表达。本研究制备一系列CD40抗体,可用于CD40检测、CD40信号的阻断或激活。
[Abstract]:CD40 is a member of the tumor necrosis factor receptor superfamily and belongs to type I transmembrane glycoprotein. Its natural ligand is a key regulator of CD40L.CD40-CD40L signaling pathway in many important physiological and pathological processes, based on its important role in the activation of immune response. In tumor immunotherapy, CD40 agonist antibody with activated CD40-CD40L signal can be used to improve the anti-tumor immune response. CD40-CD40L signal also plays an important role in transplant rejection and inflammation. Blocking the CD40 signaling pathway is the current treatment of rheumatoid arthritis, enteritis, systemic lupus erythematosus. The preparation of CD40 blocking or agonist antibodies can not only be used to detect the expression of CD40 molecules. We can also develop immunotherapy for inflammatory disease or tumor therapy. In this study, we express extracellular segment protein of CD40 and prepare 14 monoclonal antibodies against CD40 by immunization. It was found that all of the 9 CD40 antibodies which could be used as western blotting to detect the expression of CD40 protein could be used as an antibody to detect the expression of human CD40 by flow cytometry. One CD40 antibody could block the interaction between human CD40L and CD40. Significantly, 8 strains of CD40 antibody could activate the signal pathway of human CD40 molecule. In addition, 10 strains could activate the expression of downstream CD40 gene in mouse spleen cells and 7 strains could up-regulate the expression of costimulatory molecule CD86 on the surface of mouse spleen B cells. In this study, a series of CD40 antibodies were prepared. It can be used to detect the blocking or activation of CD 40 signal by CD40.
【作者单位】：上海交通大学医学院中国科学院上海生命科学研究院健康科学研究所;
【基金】：国家重点基础研究发展计划(2014CB541900)
【分类号】：R392
【正文快照】： CD40(cluster of differentiation 40,CD40)是肿瘤坏死因子受体超家族成员之一,表达于多种APC、成纤维细胞、内皮细胞以及肿瘤细胞表面[1]。其配体CD40L可以结合CD40促使其进入细胞内,激活CD40信号,诱导许多下游基因的表达。激活的CD40-CD40L信号通路能活化DC,增强其抗原提呈

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