雌酮硫酸钠多克隆抗体的制备与免疫分析法的研究-药剂学专业论文.docxVIP

新疆医科大学医学硕士学位论文 新疆医科大学医学硕士学位论文 — — PAGE 2— — —3— 课题首先建立了半抗原衍生化的方法，即通过化学合成的方法将甾体激素雌酮先合 成为雌酮-17-肟，再将雌酮-17-肟合成为雌酮硫酸钠-17-肟。在此基础上建立了制备 雌酮硫酸钠多克隆抗体的方法，并初步建立了简单、快速、方便的胶体金免疫层析 试纸法和双抗夹心ELISA法检测孕马尿中混合雌激素。 关键词：雌酮硫酸钠；胶体金免疫层析法；酶联免疫吸附法：多克隆抗体 Abstract Preparation of polyclonal antibody of sodium estrone sulfate and study immunoassay specific for sodium estrone sulfate Postgraduate：Gong Yue Hong Supervisor：Prof. Gao Xiao Li Abstract Objective: To establish simple, fast and sensitive immunoassay analytical methods for detecting sodium estrone sulfate (ESS) with preparation of polyclonal antibodies (PcAb) for sodium estrone sulfate. Key techniques such as colloidal gold immunochromato graphic test and ELISA methods would help to develop new tools for detection on great quantity of ESS from pregnant mare urine (PMU) more quickly in fields. Methods：1） ESS and bovine serum albumin (BSA) as key raw materials,. The conjugations of sodium estrone sulfate-17-(O-carboxymethyl)oxime with proteins as a complete antigen were prepared by the mixed acid anhydring method. And the conjugations were analyzed by ultraviolet scanning and SDSNew Zealand white rabbits were immunized by above immunogen of hapten-BSA according to the immune protocol. Then antiserum was collected in order to prepare polyclonal antibodies of sodium estrone sulfate .2) Tri- sodium citrate with aqueous gold chloride were prepared to make gold sol. Colloidal gold were labeled with ESS-PcAb and then were dropped onto conjugate pad after the conjugate were purified. At the same time ESS-PcAb and sheep anti-rabbit IgG were coated ESS-PcAb and sheep anti-rabbit IgG on Nitrocellulose membrane with different dilutions. Nitrocellulose membrane, gold conjugate pad, sample application pad and absorbent pad were assembled onto a plastic back and cut into individual strips (3m m× 5.5mm). 3) A sandwich competition ELISA was developed for detection ESS from PMU. 40ug/ml ESS-PcAb was coated on Use microplate as a capture and HRP conjugate E