醋酸甲羟孕酮人工抗原的合成及间接ELISA的建立

Synthesis of medroxyprogesterone acetate artificial antigen and development of indirect competitive inhibition ELISA

利用羧甲基羟氨盐酸将醋酸甲羟孕酮(MPA)肟化,制备出带羧基的半抗原,再采用混合酸酐法合成人工抗原MPA-BSA,并经过紫外扫描确定偶联成功。以MPA-BSA免疫新西兰白兔并获得了针对MPA的特异性抗血清。在此基础上建立醋酸甲羟孕酮的间接酶联免疫吸附分析法(ELISA),检测限可达到0.8ng/mL,检测范围为0.4～50ng/mL。

Medroxyprogesterone acetate （ MPA ） was activated by o-carboxymethylhydroxylamine hemi HCl. Then the pure 3-CMO-MPA（ hapten of MPA） was obtained. The hapten was then linked to carrier protein BSA and OVA by using the mixed anhydride method to prepare artificial antigens, which were confirmed by UV spectrophotometry. Then, MPA-BSA was used to immunize the New Zealand white rabbits and the specific antiserum against the antigen was obtained. The indirect competitive inhibition ELISAof medroxyprogesterone acetate has been established. Under optimized conditions, the standard ELISA inhibition curve for the detection of MPA was established which was in a linear range of 0.4 ～ 50 ng/mL and the limit of detection （I20） was 0.8 ng/mL, whereas the half-maximal inhibition （I50） value was 3.5 ng/mL.