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  本文選題：hIL-17　切入點：單克隆抗體　出處：《汕頭大學》2010年碩士論文　論文類型：學位論文

【摘要】： 背景與目的:IL17-A是IL17家族(包括IL-17A, IL-17B, IL-17C, IL-17D, IL-17E和IL-17F)的成員,作為Th17細胞亞型因主要分泌的蛋白分子近年來受到廣泛關注。Th17細胞與同屬于CD4T淋巴細胞的Th1、Th2亞型在刺激條件和轉錄因子等方面有顯著不同。在類風濕性關節(jié)炎(RA)、系統(tǒng)性紅斑狼瘡(SLE)、銀屑病等病患中發(fā)現(xiàn)IL17-A表達升高,但并不清楚IL17-A在疾病發(fā)病過程中的具體作用。因此,制備經(jīng)濟有效的IL17-A以及抗hIL-17單克隆抗體(mAb)為今后相關實驗研究具有重要意義。 方法及結果:克隆并原核表達hIL-17蛋白,以之為免疫原免疫BALB/c小鼠,取免疫小鼠脾細胞和小鼠骨髓瘤NS-1細胞融合。通過間接ELISA法篩選可特異性分泌抗hIL-17 mAb的雜交瘤細胞。采用Ig類與亞類鑒定試劑盒鑒定mAb的Ig亞類;通過ELISA、Western blot、免疫組織化學染色法鑒定mAb的特異性。獲得7株可穩(wěn)定分泌抗hIL17-A mAb的雜交瘤細胞,其分泌的mAb六株為IgM,一株為IgG2a。Western blot分析表明,此mAb所識別的靶分子的相對分子質量(Mr)約為15400。免疫組織化學染色表明,mAb與食管癌、胃癌反應呈陽性。 結論:成功地制備IL17-A以及抗hIL-17 mAb,為今后相關實驗研究奠定了基礎。
[Abstract]:Background and objective: IL17-A is a member of the IL17 family (including IL-17A, IL-17B, IL-17C, IL-17D, IL-17E and IL-17F). In recent years, the protein molecules secreted by Th17 cells have attracted extensive attention. Th17 cells are significantly different from Th1T 2 subtypes, which belong to CD4T lymphocytes, in stimulation conditions and transcription factors, and in rheumatoid arthritis. Increased expression of IL17-A was found in patients with systemic lupus erythematosus and psoriasis. However, the role of IL17-A in the pathogenesis of the disease is not clear. Therefore, the preparation of economical and effective IL17-A and anti hIL-17 monoclonal antibody (mAb) is of great significance for the related experimental research in the future. Methods and results: hIL-17 protein was cloned and expressed in prokaryotic cells and immunized BALB/c mice. The hybridoma cells secreting specific anti-#en2# mAb were selected by indirect ELISA method. The immunized spleen cells were fused with mouse myeloma NS-1 cells. The Ig subclasses of mAb were identified by Ig and subclass identification kit. The specificity of mAb was identified by Elisa Western blot and immunohistochemical staining. Seven hybridoma cells secreting hIL17-A mAb stably were obtained, six of which secreted mAb and one was identified by IgG2a.Western blot analysis. The relative molecular weight of the target molecule recognized by this mAb was about 15400.Immunohistochemical staining showed that mAb was positive to esophageal carcinoma and gastric cancer. Conclusion: IL17-A and anti-hIL-17 mAbwere successfully prepared, which laid a foundation for related experimental research in the future.
【學位授予單位】：汕頭大學
【學位級別】：碩士
【學位授予年份】：2010
【分類號】：R392

【共引文獻】

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本文編號：1591463