全部 标题 作者
关键词 摘要

OALib Journal期刊
ISSN: 2333-9721
费用:99美元

查看量下载量

相关文章

更多...

TRIP-1抗体的制作及应用

, PP. 139-143

Keywords: 原核表达纯化,TRIP-1,蛋白,动物免疫,抗血清,抗体鉴定

Full-Text   Cite this paper   Add to My Lib

Abstract:

旨在制作TGF-β二型受体结合蛋白TRIP-1的抗体,以便推进对该蛋白在TGF-β信号通路以及癌症发生发展中的功能研究。首先克隆出人的TRIP-1基因,构建其原核重组表达载体pET-His-TRIP-1,将其转化至大肠杆菌BL21(DE3)中,诱导获得TRIP-1蛋白,纯化后将其分别免疫新西兰大耳兔和昆明小鼠(KM),免疫4次后取血清,并对抗血清进行效价和亲和力测定。通过Westernblot和免疫荧光试验鉴定抗血清特异性。最后,利用proteinG初步纯化抗血清。结果表明,通过重组表达获得抗原蛋白,纯化后免疫新西兰大耳兔和昆明鼠可得到效价较高的抗TRIP-1的多克隆抗体,在稀释倍数较高的情况下仍可以用于Westernblot和免疫荧光试验,且有非常好的特异性。

References

[1]  Chen RH, Miettinen PJ, Maruoka EM, et al. A WD-domain protein that is associated with and phosphorylated by the type [J]. Nature, 1995, 377(6549):548-552. [2] Choy L, Derynck R. The type II transforming growth factor(TGF) -β receptor-interacting protein TRIP-1 acts as a modulator of the TGF -β response [J]. J Biol Chem, 1998, 273(47):31455-31462. [3] Perez RE, Navarro A, Rezaiekhaligh MH, et al. TRIP-1 regulates TGF-β1-induced epithelial-mesenchymal transition of human lung epithelial cell line A549 [J]. Am J Physiol-Lung Cell Mol Physiol, 2011, 300(5):L799-L807. [4] Metz-Estrella D, Jonason JH, Sheu TJ, et al. TRIP-1 :A regulator of osteoblast function [J]. Journal of Bone and Mineral Research, 2012, 27(7):1576-1584. [5] Ramachandran A, Ravindran S, George A. Localization of transforming growth factor beta receptor II interacting protein-1 in bone and teeth [J]. J Histochem Cytochem, 2012, 60(4):323-337. [6] Ahlemann M, Zeidler R, Lang S, et al. Carcinoma-associated eIF3i overexpression facilitates mTOR-dependent growth transformation [J]. Molecular Carcinogenesis, 2006, 45(12):957-967. [7] Asano K, Kinzy TG, Merrick WC, et al. Conservation and diversity of eukaryotic translation initiation factor eIF3 [J]. Journal of Biological Chemistry, 1997, 272(2):1101-1109. [8] 罗惠霞, 李敏, 王玉炯. 包涵体蛋白复性的几种方法[J]. 生 物技术通报, 2007(5):96-98. [9] Beatty JD, Beatty BG, Vlahos WG. Measurement of monoclonal antibody affinity by non-competitive enzyme immunoassay [J]. Journal of Immunological Methods, 1987, 100(1-2):173-179. [10] Ding G, Zhu J, Wang Z, et al. Measurement of scFv antibody affinity using non-competitive enzyme-linked immunosorbent assay [J]. 南京医科大学学报(英文版), 2005, 19(2):4. [11] 秦睿玲, 张西臣, 李建华, 等. 柔嫩艾美球虫间接ELISA 检测 方法的建立[J]. 中国兽医科技, 2004, 34(7):60-63. [12] 张小兵, 邸禄芹, 吴萌, 等. 单克隆抗体与多克隆抗体配对 ELISA 方法比较[J]. 生物技术通报, 2009(11):125-129. [13] 陈卓, 刘家驹, 毕亮, 等. NPR1 多肽抗体的制备和应用[J]. 生物技术通报, 2012(1):145-150.

Full-Text

Contact Us

service@oalib.com

QQ:3279437679

WhatsApp +8615387084133