双黄连注射剂中1,3-二咖啡酰奎宁酸致敏家兔后血清中特异性抗体的测定

Determination of Specific Antibodies in Allergic Rabbits Induced by 1,3-Di-caffeoylquinic Acid in Shuanghuanglian Injection

目的:检测双黄连注射剂中1,3-二咖啡酰奎宁酸致敏家兔后血清中特异性抗体效价。方法:双黄连注射剂中疑似小分子半抗原物质1,3-二咖啡酰奎宁酸与正常家兔血清体外孵育制备完全抗原,免疫家兔获得特异性抗体。用ELISA法检测抗血清的抗体效价。间接竞争ELISA法测定血清特异性并绘制抑制曲线。采用兔免疫球蛋白E(Ig E)ELISA试剂盒检测各组家兔抗血清中IgE抗体的含量。结果:1,3-二咖啡酰奎宁酸的抗体效价(A)值高于阴性对照组2倍,说明其具有潜在的致敏性。回归方程I=0.170 6lg C+0.317 5,相关系数r=0.985 4,检测限IC10为57.40μg·ml^(-1),半数抑制浓度IC50=8.732 0mg·ml^(-1)。家兔体内明显产生了外源性IgE型抗体。结论:在本实验条件下双黄连注射剂中疑似半抗原物质1,3-二咖啡酰奎宁酸具有致敏性。

Objective： To determine the antibody titer in the serum of allergic rabbits after the injection of 1, 3-di-caffeoylquinic acid contained in Shuanghuanglian injection. Methods： The complete antigen was prepared by incubating the suspected small molecu- lar hapten 1, 3-di-caffeoylquinic acid contained in Shuanghuanglian injection with the serum from the normal rabbits. The specific anti- body was obtained in the immunized rabbits. The antibody titers of antiserum were measured by ELISA kits. Indirect competitive ELISA was used to determine serological specificity, and the obtained data was used to plot the inhibition curves. The content of IgE antibody in the antiserum of rabbits was detected by rabbit immunoglobulin E （IgE） ELISA kits. Results： The antibody titer （A） of 1,3-di-caffcoylquinie acid was 2 times higher than that of the negative control, which indicated its potential allergenicity. The regres- sion equation was I=0. 170 61gC ＋0.317 5, which was with the correlation coefficient of r =0.985 4, the detection limit of IC10 = 57.40 μg ml - 1 and the half inhibitory concentration of ICs0 = 8. 732 0 mg ml - 1. Furthermore, the exogenous IgE antibody was pro- tinted in the rabbits. Conclusion： The results indicated that the hapten substance 1,3-di-caffeoylquinic acid in Shuanghuanglian injec- tion was allergenic under the present experimental conditions.