甲拌磷人工抗原的合成及多克隆抗体制备

Phorate Synthesis and Production of Its Polyclonal Antibody

以五硫化二磷与无水乙醇为原料，首先制得硫化物中间体，再添加甲醛和2-巯基乙醇进行反应，得到甲拌磷半抗原。在DMAP作为催化剂的条件下，甲拌磷半抗原与丁二酸酐反应，得到甲拌磷半抗原的衍生物。通过碳二亚胺法将甲拌磷半抗原衍生物分别与牛血清白蛋白（BSA）和鸡卵清白蛋白（OVA）共价偶联，得到免疫抗原和包被抗原。用免疫抗原免疫兔子获得了高效价的多克隆抗体，抗血清效价达到了1：50000。通过试验确立甲拌磷标准曲线，检测限为6．383μg／L，检测线性范围为6．383-10000μg／L。

Sulfide was synthesized from phosphorus pentasulfide and ethanol. Phorate hapten was obtained from the sulfide reacted with formaldehyde and 2-mercapto-ethanol. With DMAP as cata lyst,phorate hapten and succinic anhydride synthesized Phorate ramification. The Phorate ramifi cation was conjugated to bovine serum albumin （BSA） and ovalbumin （OVA） with the modified active ester method to form artificial immune antigen and coating antigen. The anti-phorate polyclonal antibodies were obtained by using the artificial immune antigen to immune rabbits. The serum titer was as high as 1 ： 50 000. Experiment determined the standard curves,and the real detec tion limit of the ELISA was 6. 383μg/L and the linear range was 6. 383-10 000μg/L.