条斑紫菜R-藻红蛋白荧光探针制备条件优化.PDFVIP

生 物 工 程 学 报 Chin J Biotech 2008, January 25; 24(1): 153-158 journals.im.ac.cn Chinese Journal of Biotechnology ISSN 1000-3061 cjb@im.ac.cn ©2008 Institute of Microbiology, CAS CSM, All rights reserved. 研究简报 R- 周 铭, 蔡春尔, 柳俊秀, 吴维宁, 何培民 , 200090 : 通过化学方法使条斑紫菜 R-藻红蛋白与抗体进行交联以制备荧光探针, 并对制备条件进行优化。首先采用异 双功能试剂SPDP (N-琥珀酰亚氨基-3-2-吡啶基二硫丙酸醇)和DTT （二硫苏糖醇）分别使 R-PE(R- 藻红蛋白)衍生化、 IgG （单克隆抗体）巯基化, 其次测定了 SPDP 与R-PE 不同摩尔比对R-PE 衍生化的影响、DTT 与IgG 不同摩尔比对 IgG 巯基化的影响, 结果表明：SPDP 与R-PE 的最佳摩尔比为40 ：1, DTT 与IgG 的最佳摩尔比为500:1。在此基础上, 建 立了R-PE 与IgG 交联的制备技术, 并应用全波长扫描吸收光谱、电泳分析和荧光显微镜观察等监测和分析技术, 证实 了藻红蛋白与抗体已成功交联形成了复合物。 : R- 藻红蛋白, 单克隆抗体, 交联, 荧光探针, 制备, 条件优化 Optimization of Fluorescence Probe Preparation for R-Phycoerythrin in Porphyra yezoensis Ming Zhou, Chuner Cai, Junxiu Liu, Weining Wu, and Peimin He Key Laboratory of Aquatic Genetic Resources and Aquacultural Ecology Certificated by the Ministry of Agriculture, Shanghai Fisheries University, Shanghai 200090, China Abstract: We optimized the chemical conjugation between R-phycoerythrin and antibody. First, the R-PE (R-phycoerythrin) was derived with beterobifunctional reagent SPDP(N-succinimidyl-3-2-pyridyldithio propionate) and antibody was thiolated with DTT(dithiothreitol). Second, we determined the effects of the different molar ratio of SPDP to R-PE for the derivation and DTT to IgG for the thiolation. The results showed that the optimum molar ratio of SPDP to R-PE for derivation was 40:1, and DTT to IgG for thiolation was 500:1. The con