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制备针对乙氧基磷酸酯类有机磷农药的单克隆抗体,以此为基础建立该类农药的快速免疫筛选检测方法.以二乙基磷酸乙酸为通用结构半抗原,分别使之与牛血清白蛋白和鸡卵清蛋白共价偶联,合成免疫原和包被原并对其进行结构鉴定.偶联成功后的免疫原用于免疫Balb/c小鼠.将免疫成功小鼠的脾细胞与小鼠SP2/0骨髓瘤细胞融合,筛选能稳定分泌抗乙氧基磷酸酯类有机磷农药单克隆抗体的杂交瘤细胞株.获得的小鼠腹水用辛酸-硫酸铵法纯化,所得纯化抗体以琼脂双扩散法鉴定其免疫球蛋白类型,间接竞争ELISA方法测定其对半抗原的灵敏度、特异性和亲和性.结果表明,该抗体分泌IgG1亚类的单克隆抗体,且与二乙基磷酸乙酸的亲和性较高(1.4×107L/mol),所得抗体对毒死蜱、对硫磷、丙溴磷、氧化乐果、除线磷、二嗪农、溴硫磷、辛硫磷、喹硫磷、三唑磷等农药有特异性反应.该检测技术可用于上述农药的快速定性或定量检测.
Preparation of a monoclonal antibody against organophosphorus pesticides ethoxyphosphonates, as the basis for the establishment of rapid detection of pesticides screening for this kind of pesticides.With diethylphosphoacetate as the general structure of haptens, respectively, and bovine serum albumin The protein and chicken ovalbumin were covalently coupled to synthesize the immunogen and the original coating and to identify the structure.The immunogen after the coupling was used to immunize Balb / c mice.After immunization, The mouse SP2 / 0 myeloma cells were fused and the hybridoma cell line stably secreting the monoclonal antibody against organophosphorus pesticide of ethoxyethionate was screened, and the ascites obtained from mice was purified by caprylic acid-ammonium sulfate method. The purified antibody Agar double diffusion method to identify the type of immunoglobulin, indirect competitive ELISA method to determine the sensitivity, specificity and affinity of the hapten.The results show that the antibody secreted IgG1 subclass of monoclonal antibodies, and diethyl phosphoric acid (1.4 × 10 7 L / mol). The obtained antibodies against chlorpyrifos, parathion, profenofos, omethoate, fenitrothion, diazinon, phoxim, phoxim, , Triazophos and other pesticides have specific anti- The detection technique may be used to qualitatively or quantitatively detecting the fast pesticides.