兔抗邻苯二甲酸二甲酯的多克隆抗体制备及特性分析

Preparation and property of antibody against environmental hormone dimethyl phthalate

通过偶氮键将邻苯二甲酸二甲酯半抗原衍生物与载体蛋白BSA偶合后作为免疫原,免疫新西兰大白兔,得到合格的邻苯二甲酸二甲酯抗血清,双向琼脂扩散方法测定血清效价为1∶16～1∶32,间接荧光免疫法检测效价为1∶3 200～1∶6 400.通过兔颈部采血、抗血清纯化,得到抗邻苯二甲酸二甲酯多克隆抗体IgG冻干后分装,-20℃低温保存.建立了间接荧光免疫法检测纯化抗体的方法.间接竞争荧光免疫法考察交叉反应试验结果显示：纯化的邻苯二甲酸二甲酯抗体与邻苯二甲酸二乙酯、邻苯二甲酸二丙酯、邻苯二甲酸二丁酯、邻苯二甲酸二环己酯、邻苯二甲酸二戊酯、邻苯二甲酸二（2-乙基己基）酯等邻苯二甲酸酯类环境激素的交叉反应率均＜9%.制备的抗体可应用于环境激素类物质邻苯二甲酸二甲酯的荧光免疫分析.

The hapten of dimethyl phthalate（DMP） was conjugated to bovine serum albumen（BSA） by azo coupling. The conjugate was used as an immunogen to inject New Zealand white rabbits. The final sera had titers of 1 ： 16 - 1 ： 32 with double immunodiffusion, and 1 ： 3 200 - 1 ： 6 400 with indirect competitive fluorescence immunoassay. After an acceptable antibody titer was observed, the animals were exsanguinated and the blood was collected in glass tubes, and purified. Then purified IgG fractions were lyophilized ahquoted into vials, and stored at - 20℃ until use. An indirect fluorescence immunoassay to detect an. tibody was developed. The analytical experiment of anti-DMP by indirect competitive fluorescence immunoassay showed that anti-DMP antibody was cross-reacted with other similar phthalate compounds, such as diethyl phthalate （ DEP）, dipropyl phthalate （ DPrP）, dibutyl phthalate （ DBP）, dicyclohexyl phthalate （ DCHP）, di-n- pentyl phthalate （DPP） and diethylhexyl phthalate（ DEHP）, and the cross reactivity were less than 9%. Anti-DMP antibody was therefore considered interesting for developing the fluorescence immunoassay.