有机磷杀虫剂通用结构半抗原的设计及广谱特异性抗体的制备

Use of Generic Hapten in Production of Broad Specificity Anti-Organophosphorous Pesticides Antibody

【目的】制备针对有机磷农药的广谱特异性抗体,用于有机磷农药的免疫快速筛选检测。【方法】设计采用二乙基膦酸乙酸作为通用结构半抗原,用NHS-DCC法和EDC法合成两种人工抗原免疫新西兰大白兔制备抗血清。【结果】NHS-DCC法所制得抗血清的最高滴度为25600,EDC法合成的抗原制得的抗血清滴度最高为6400,均获得免疫应答。以二乙基膦酸乙酸为对象建立的间接竞争ELISA检测方法对二乙基膦酸乙酸最低检测浓度0.0035μg·ml-1,抑制中浓度I50为0.182μg·ml-1。经对12种常见磷酸酯类有机磷农药特异性检测反应试验,结果发现:所得抗体对毒死蜱、氧乐果、二嗪农、乙基对硫磷、丙溴磷、辛硫磷等农药有特异性反应,I50分别为0.12、0.21、0.24、0.78、0.97、3.8μg·ml-1。【结论】该检测技术可用于毒死蜱、氧乐果、二嗪农、乙基对硫磷、丙溴磷等药剂的快速定性或半定量检测。

【Objective】The objective of this study is to develop broad specificity polycolonal antibodies against a group of organophosphorous pesticides.【Method】 Diethylphosphono acetic acid （DPA） was used as a generic hapten, and conjugated to bovine serum albumin （BSA） by the mixed-anhydried method and EDC method. Six New Zealand white rabbits were immunized for preparation of antiserum.【Result】The titers of antiserum by prepared the two methods had reached 25 600 and 6 400, respectively. The CI-ELISA was conducted with the antiserum, the detection limit for DPA was 0.0035 μg·ml^-1 and the I50 was 0.182 μg·ml^-1. The antiserum also showed high affinities to chlorpyrifos, omethoate, diazinon, parathion-ethyl, profenofos, and phoxin, the I50 to these pesticides were 0.12, 0.21, 0.24, 0.78, 0.97, 3.8 μg·ml^-1. 【Conclusion】It is concluded that this assay is a suitable screening tool for quantities and semi-quantities detection of the five organophosphorous pesticides.