烟嘧磺隆人工抗原的合成与鼠源多克隆抗体的制备

摘要

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∶16000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1 ng·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 immunochem-ical assay to detect nicosulfuron residue,haptens were synthesized with 2-aminosulfonyl-N ,N-dimethylnicoti-namide(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 anti-gens were prepared successfully and identified by UV-visible spectroscopy and SDS-PAGE.The coupling ratios were 10.5∶1 and 26.9∶1 for ASDM to ovalbumin(OVA)and bovine serum albumin(BSA),respectively.After quintic animal immunizations,the antibody titer of all immunized mice were over 1∶1 6 000,whereby polyclonal antibodies against nicosulfuron were acquired from mice immunized by ASDM-BSA.And then an indirect com-petitive ELISA was established for nicosulfuron content determination with a detection range of 1 ng·mL-1 ~10 μg·mL-1 .This research could provide reference for further development of more sensitive,effective and fea-sible ELISA detection method.