对硫磷人工抗原合成及其鉴定

郭剑平; 刘云霞; 高增林; 张伟; 吴启庆; 郭月凤

对硫磷人工抗原合成及其鉴定

Preparation of conjugate antigen and antibody to parathion

摘要

摘要:
目的制备对硫磷人工抗原和抗血清, 为建立酶联免疫吸附法提供技术储备。
方法还原对硫磷硝基成为氨基, 合成半抗原氨基对硫磷; 重氮化偶联大分子载体牛血清白蛋白制备人工抗原; 免疫新西兰白兔, 进行多抗制备。
结果合成的半抗原核磁共振图谱鉴定结果为δ: 6.9~7(m, 2H, CH₂)6.4~6.5(m, 2H, CH₂)4.1~4.2(q, 4H, -CH₂)34(s, 2H, NH₂)13~14(t, 6H, CH₃); 氨基对硫磷与牛血清白蛋白的结合比为35~36:1;抗血清经间接酶联免疫吸附法检测, 效价达1:128000, 双向免疫扩散实验测得抗血清与人工抗原形成沉淀; 间接竞争酶联免疫吸附法测得抗血清对对硫磷最低抑制浓度为0.16ng/mL; 经间接酶联免疫吸附法检测人工抗原与羊抗人血清、兔抗人血清、正常兔血清无交叉反应。
结论合成对硫磷半抗原及人工抗原纯度高、结合比高, 具有较好的免疫原性。

Abstract:
Objective To produce the organophate parathion conjugate antigen antibody.
Methods The hapten, which reduces the parathion changing-NO₂ into-NH₂, is conjugated diazo-linked with the carrier protein BSA to prepare conjugate antigen.By immunizating the New Zealand rabbit, the antisera is obtained.
Results The result by NMR is δ: 6.9-7 (m, 2H, CH₂)6.4-6.5(m, 2H, CH₂)4.1-4.2(q, 4H, -CH2)3.4(s, 2H, NH₂)1.3-1.4(t, 6H, CH₃).The ratio of hapten to BSA is from 35-36 to 1.Test of the antisera by indirected ELISA and immunodiffusion shows that the titer is high and antibody is specific.The titer is 1:128 000.Precipitin bands are observed from the reaction of the antisera with the conjugate by immunodiffusion.The minimum concentration of ELISA inhibition for antisera is 0.16 ng/ml.There is no cross-reactivity between the antisera and serum analogues.The research confirms that the antisera is specific to the parathion.
Conclusion The antigen is in effect.The research provides the preparation for producing McAb of parathion and sets up the ELISA method of detecting parathion.

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