| 及志恒,何 颖,张俊艳,肖春梅,郑俊斌,邹泽红,陶爱林.重组扁豆过敏原 Len c 1表达纯化及免疫活性鉴定[J].,2017,17(8):1416-1419 |
| 重组扁豆过敏原 Len c 1表达纯化及免疫活性鉴定 |
| Purified Expression and Identification of Immune Activity of the Major Allergen Len c 1 of Lentil |
| 投稿时间：2016-06-11 修订日期：2016-06-27 |
| DOI：10.13241/j.cnki.pmb.2017.08.004 |
| 中文关键词: rLen c 1 纯化 免疫活性 IgE |
| 英文关键词: rLen c 1 Purification Immune activity IgE |
| 基金项目:国家科技重大专项重点课题（2014ZX0801105B）；国家临床重点专科建设项目（520102-110）；广东省产学研项目（2013B090500129） |
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| 中文摘要: |
| 摘要 目的：纯化重组扁豆过敏原Len c 1（rLen c 1）并进行免疫活性鉴定。方法：通过原核表达的方式生产r Len c 1，然后采用亲和层析的方法纯化带有StrepII标签的目的蛋白。将BALB/c小鼠随机分为对照组（注射生理盐水）和过敏原致敏组（注射r Len c 1），通过腹腔注射方式免疫小鼠，建立BALB/c小鼠扁豆致敏模型，利用间接ELISA法检测血清总IgE和过敏原特异性IgE，鉴定重组扁豆过敏原的免疫活性。结果：IPTG成功诱导Len c 1蛋白表达，rLen c 1蛋白主要以包涵体形式表达，通过透析复性和亲和层析获得纯化的复性扁豆过敏原蛋白rLen c 1，成功建立小鼠致敏模型。和对照组相比，致敏组小鼠TIgE及过敏原特异性IgE水平均明显增高，结论：获得纯化的具有免疫活性的rLen c 1过敏原蛋白，为扁豆过敏机制研究、单克隆抗体的制备、以及临床诊断和免疫治疗奠定基础。 |
| 英文摘要: |
| ABSTRACT Objective: To purify recombinant lentils allergens Len c 1 (rLen c 1) and identify the immune activity. Methods: By means of prokaryotic expression system rLen c 1 was produced , then the target protein with Strep Ⅱ tag was purified with the method of affinity chromatography. BALB/c mice were randomly divided into control group (injected by normal saline) and allergen sensitized group (injected by rLen c 1). The mice were immuned by intraperitoneal injection to set up BALB/c mice lentils allergic model, then the total IgE and allergen specific IgE were detected by indirect ELISA to identify the immune activity of recombinant lentil allergen. Results: Len c 1 protein expression was successfully induced by IPTG，which was expressed in the form of inclusion body; with dialysis renaturation and affinity chromatography,the highly purified renatured protein rLen c 1 was gained, and mice sensitization model was established successfully. Compared with the control group of mice, the TIgE and allergen specific IgE significantly increased in sensitized group of mice. Conclusion: Purified rLen c 1 allergen protein with immune activity was obtained to lay a foundation for the preparation of monoclonal antibodies, clinical diagnosis and immunotherapy. |
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