烟嘧磺隆人工抗原的合成与鼠源多克隆抗体的制备

Synthesis of Nicosulfuron Artificial Antigen and Preparation of Mouse Polyclonal Antibody

为了制备烟嘧磺隆的多克隆抗体、建立烟嘧磺隆残留的免疫分析方法,以2-氨基磺酰基-N,N-二甲基烟酰胺（ASDM）和琥珀酸酐为原料合成半抗原,采用活泼酯法制备人工抗原并免疫小鼠。结果表明：经紫外可见吸收光谱扫描和聚丙烯酰胺凝胶电泳（SDS-PAGE）鉴定,人工抗原合成成功,半抗原与卵清蛋白（OVA）、牛血清白蛋白（BSA）的偶联比分别为10.5∶1和26.9∶1。5次免疫后小鼠抗血清的效价均达到1∶16 000以上,以ASDM-BSA为免疫原的小鼠产生了抑制烟嘧磺隆的多克隆抗体,建立的间接竞争ELISA方法对烟嘧磺隆的检测范围是1ng·mL-1-10μg·mL-1。为进一步建立更为高效、灵敏和方便的烟嘧磺隆ELISA检测方法提供了参考。

To prepare the polyclonal antibodies against pesticide nicosulfuron and establish an immunochemical assay to detect nicosulfuron residue,haptens were synthesized with 2-aminosulfonyl-N,N-dimethylnicotinamide（ASDM）and succinic anhydride as raw materials.Artificial antigens were synthesized by connecting the hapten to carrier proteins with an active ester method,and then mice were immunized.The two artificial antigens were prepared successfully and identified by UV-visible spectroscopy and SDS-PAGE.The coupling ratios were 10.5∶1and 26.9∶1for ASDM to ovalbumin（OVA）and bovine serum albumin（BSA）,respectively.After quintic animal immunizations,the antibody titer of all immunized mice were over 1∶16 000,whereby polyclonal antibodies against nicosulfuron were acquired from mice immunized by ASDM-BSA.And then an indirect competitive ELISA was established for nicosulfuron content determination with a detection range of 1ng·mL-1-10μg·mL-1.This research could provide reference for further development of more sensitive,effective and feasible ELISA detection method.