抗人PD-L1单克隆抗体的制备及其应用

发布时间：2018-06-19 11:16

本文选题：PD-L + 单克隆抗体　；参考：《中国免疫学杂志》2017年06期

【摘要】：目的:获得能应用于临床诊断以及阻断PD-L1与PD-1结合的抗人PD-L1单克隆抗体。方法:采用重组表达的人PD-L1蛋白免疫BALB/c小鼠,通过杂交瘤细胞融合技术获得稳定分泌抗人PD-L1单抗的阳性细胞株,ELISA方法鉴定抗体的特异性、亲和力、亚型等方面特性;免疫印迹、间接免疫荧光方法对肿瘤细胞进行检测;肿瘤杀伤实验验证抗体阻断活性。结果:共获得2株抗人PD-L1单抗,抗体效价分别为1∶2.56×10~6和1∶3×10~5,亲和力分别为1.5×10~9L/mol和2.5×10~8L/mol,均与PD-L2蛋白无交叉反应。免疫印迹、间接免疫荧光证实抗体有诊断作用。杀伤实验显示抗体有阻断作用。结论:共获得两株稳定分泌高效价、高特异性的抗人PD-L1单抗的杂交瘤细胞株,能作为诊断抗体应用于肿瘤表型检测和预后有效性的评估。抗体的阻断功能可应用于联合CIK细胞免疫治疗。
[Abstract]:Objective: to obtain anti-human PD- L1 monoclonal antibody which can be used in clinical diagnosis and block the binding of PD- L1 to PD-1. Methods: BALB / c mice were immunized with recombinant human PD-L1 protein. Immunoblotting was used to identify the specificity, affinity and subtype of antibody by Elisa, which secreted anti-PD-L1 monoclonal antibody stably by hybridoma cell fusion technique. Indirect immunofluorescence assay was used to detect tumor cells, and tumor killing assay was used to verify the blocking activity of antibody. Results: two monoclonal antibodies against human PD-L1 were obtained. The titers of the antibodies were 1: 2.56 脳 10 ~ (-6) and 1:3 脳 10 ~ (5), respectively. The affinity was 1.5 脳 10 ~ (9) L / mol and 2.5 脳 10 ~ (8) L / mol, respectively. Immunoblotting, indirect immunofluorescence confirmed the antibody has diagnostic effect. The killing test showed that the antibody had blocking effect. Conclusion: two hybridoma cell lines secreting stable high titer and high specificity anti-human PD-L1 monoclonal antibodies were obtained, which can be used as diagnostic antibodies for the evaluation of tumor phenotype and prognosis. The blocking function of antibody can be used in combination with CIK cell immunotherapy.
【作者单位】：暨南大学抗体工程研究中心广东省分子免疫与抗体工程重点实验室;暨南大学转化医学转化研究院国际免疫学中心;
【基金】：广东省人才引进创新团队项目《重大疾病的临床诊断试剂研发》(2013S028)
【分类号】：R730.51

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