脱落酸人工抗原合成及多克隆抗体制备

Synthesis of Antigen and Preparation of Multi-Clonal Antibody Against Abscisic Acide

目的:为了对植物细胞中的脱落酸(ABA)进行定量和定位分析,研究了脱落酸人工抗原的合成以及多克隆抗体的制备。方法:用重氮化法将ABA分别与牛血清白蛋白(BSA)、卵清白蛋白(OVA)联结,得到ABA的免疫抗原和包被抗原,并采用紫外全波长扫描和SDS-PAGE对合成的抗原进行了鉴定。以经过鉴定的抗原免疫白兔,制备出ABA的多克隆抗体;采用间接酶联免疫法(ELISA)对抗血清进行效价检测,通过离子交换层析法获得纯化的抗体。结果:ABA与BSA的平均偶联比为5.3∶1,抗血清效价为1∶16000。结论:人工抗原和多克隆抗体制备成功,为采用ELISA和免疫胶体金技术(ICG)检测ABA提供了有效工具。

Objective： In order to research on the localization and quantization of abscisic acid （ABA） in plant cells, synthesis of antigen and preparation of muhi-clonal antibody against ABA were studied. Methods： ABA was conjugated to bovine serum album（BSA） and ovalbumin（OVA） via diazo coupling. The hapten was identified by UV spectrometric scanning and SDS-PAGE. Rabbits were immunized with antigen to obtain the multi-clonal antibody against ABA. Antibody was identified by enzyme-lined immunosorbent assay（ELISA）, and purified by ion exchange chromatography. Results： Based on the experiments and calculation, the conjugated ratio of ABA and BSA in ABA-C4＇-BSA was 5.3：1, and the titer of the antisera was 1：16 000. Conclusion： The results showed that the synthesis of antigen and preparation of muhi-clonal antibody against abscisic acid were successful, which supplied a good tool for the detection of ABA with ELISA and immune colloidal gold（ICG） methods.