| 题名 | 藻胆蛋白荧光标记的探索 |
| 作者 | 顾铭 |
| 学位类别 | 硕士 |
| 答辩日期 | 2001-06 |
| 授予单位 | 中国科学院研究生院 |
| 导师 | 欧阳藩 |
| 关键词 | R-藻红蛋白 单克隆抗体 蛋白质交联 |
| 其他题名 | Research of phycobiliprotein labeled conjugates |
| 学位专业 | 生化工程 |
| 中文摘要 | 蛋白质交联技术成为了藻胆蛋白应用中的重要环节，制备交联效率、荧光强度和单抗活性市制荧光标记物是提高免疫检测灵敏度的基础.该论文结合国家“八六三”项目“藻胆蛋白荧光标记物和诊断试剂”的研究工作，以R-藻红蛋白和抗HBsAg单克隆抗体为对象，对藻胆蛋白与单抗交联的方法和条件进行了研究. |
| 英文摘要 | Immunochemistry has been widely used in medical biology on the basis of the development of antibody engineering. And with the popularization of flow cytometer and DNA chip, immunofluorescence technique will be greatly improved. Phycobiliproteins are a series of fluorescence probe, including phycoerythrin, phycocyanin, allophycocyanin, phycoerythrincyanin, with high extinction coefficient, strong quantum yield and great Stocks. Phycobiliproteins have many kinds of emission wavelength and can be conjugated with monoclonal antibody to detect several antigens simultaneously. Bioconjugate technique is the key to make phycobiliproteins full use. Sensitiveness can be developed on the basis of good quality of phycobiliprotein labeled conjugate. This thesis was connected to the project, "phycobiliprotein labeled conjugate and diagnosis kit". A monoclonal antibody, anti-HBsAg, was served as a model and the method of conjugation was studied. The thesis include five parts: Preparation of monoclonal antibody: Hybridomas were injected into Balb/c mice to get ascites. Antibody was purified by salting-out, ion exchange and batch affinity chromatography. A simple protocol was chosed for conjugating R-PE and IgG. It was proved that R-phycoerythrin (R-PE) and antibody were stable in a wide range of pH and ironic strength. (3) Reaction patterns of 2-IT and DTT were studied. 2-IT was used to modified IgG instead of DTT. (4) The effect of heterobifunctional reagent concentration on the quality of conjugate was studied. A better protocol was as below: when R-PE:IgG=2:l, SPDP:R-PE=80:1 and 2-IT with the concentration of 2.5mmol/L were advantageous to amplify the fluorescence signal. 100% negative serum and 93.3% positive serum were detected with R-PE conjugated IgG in sandwich method. (5) R-PE conjugated Sheep-anti-Mouse was successfully used in detection of HBsAg. It was proved that the conjugation technique discussed in this thesis could be widely used. |
| 语种 | 中文 |
| 公开日期 | 2013-09-26 |
| 页码 | 90 |
| 内容类型 | 学位论文 |
| 源URL | [http://ir.ipe.ac.cn/handle/122111/1890]  |
| 专题 | 过程工程研究所_研究所（批量导入） |
| 推荐引用方式 GB/T 7714 | 顾铭. 藻胆蛋白荧光标记的探索[D]. 中国科学院研究生院. 2001. |
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