紫癜性肾炎特异性结合肽的筛选与鉴定  

The Selection and Identification of Specifically Binding Peptide with HSPN

作　　者：张晓燕[1] 周南[1] 胡德梅 鱼丽娟 吴守振[2] 

机构地区：[1]陕西省人民医院儿科,陕西西安710068 [2]西安市儿童医院陕西省儿科疾病研究所,陕西西安710003 [3]西安市儿童医院检验科,陕西西安710003

出　　处：《现代生物医学进展》2016年第34期6630-6633,6702,共5页Progress in Modern Biomedicine

基　　金：陕西省自然科学基础研究计划项目(2014JM2-8184)

摘　　要：目的:利用体内噬菌体展示技术筛选并鉴定紫癜性肾炎肾组织特异性结合肽。方法:构建大鼠紫癜性肾炎模型。尾静脉注射噬菌体展示环七肽库,筛选与紫癜性肾炎肾组织特异性结合的噬菌体,经过3轮体内筛选后,提取阳性单克隆噬菌体,并进行DNA测序。利用ELISA方法鉴定单克隆噬菌体对紫癜性肾炎的结合。通过化学合成的方法合成筛选获得的多肽,以流式细胞检测其与肾细胞的结合能力。同时,以合成的多肽体内封闭,检测噬菌体与合成多肽竞争结合紫癜性肾炎肾组织的能力。结果:成功筛选出于大鼠紫癜性肾炎肾组织结合的阳性噬菌体。在随机挑选的43个单克隆噬菌斑中,挑选了其中16个亲和力高的噬菌体,测序后获得了紫癜性肾炎的结合肽CQGPWKLTC。流式结果表明筛选的多肽能够与肾细胞特异性结合。获得的阳性噬菌体能够与紫癜性肾炎肾组织特异性结合,并且与体外合成的多肽具有竞争结合能力。结论:利用体内噬菌体展示技术筛选了与紫癜性肾炎肾组织特异性结合的肽CQGPWKLTC。Objective： To screen and identify the specifically binding peptide with kidney tissue from Henoch-Schonlein purpura nephritis （HSPN） using the technology of Phage display peptide library. Methods： The models of HSPN were constructed, and then the Phage display peptide library were injected the HSPN models i.v. The specifically binding peptides with kidney tissue from HSPN were screened. The positive phages were obtained after 3th selection, and then were sequenced. The positive phages binding with kidney tissue from HSPN were identified by ELISA. The obtaining peptides were synthesized. Then, the binding ability between peptides and kidney cells were analyzed using FCM. Meanwhile, to characterize the competitive binding ability between synthesizing peptides and positive phage, the kidney tissue from HSPN were protected in vivo. Results： The positive phages binding to the kidney tissue from HSPN were screened successfully. The 16 phages with higher affinity were selected randomly from 43 positive phages. The peptide CQGPWKLTC binding with kidney tissue from HSPN was obtained after sequencing. The peptide CQGPWKLTC not only could bind specifically to kidney tissue from HSPN by FCM, but had the competitive binding ability between synthesizing peptides and positive phage. Conclusion： The specifically binding peptide CQGPWKLTC with kidney tissue from HSPN were screened and identified.

关 键 词：噬菌体展示肽技术 紫癜性肾炎 筛选 小分子多肽 

分 类 号：Q959.483[生物学—动物学]