用酶联免疫吸附试验法检测印楝愈伤组织中的印楝素含量

ELISA Detection of Azadirachtin in the Callus-Induced Azadirachta indica

用双抗体夹心法酶联免疫吸附试验（DAS－ELISA）检测印楝（AzadirachtaindicaA.Juuss）愈伤组织中的印楝素含量，成功建立了DAS－ELISA。以牛血清蛋白为载体，用二环己基碳二亚胺合成免疫原，制备抗血清，获得效价为1：2560。以辣根过氧化物酶为标记酶，用“过碘酸盐氧化法”合成酶标抗体。使用包被抗体的最适浓度为5．0μg／mL，标记抗体浓度为2．5μg／mL，可测抗原最低浓度为10μg/mL。此方法简便，快速，灵敏，专一性强，用量少，且不需要预先纯化，适合于植物组织和细胞培养物中有效活性成分测定，可用于大规模理想细胞系筛选。

The azadirachtin content in the callus cultures of Azadichta indica A. Juss wassuccessfully detected by Double Antibody Sandwich Enzyme Linked Immunosorbent Assay(DAS - ELISA ). The antigen was synthesized with dicyclohexyl carbodiinide (DCCI)throughBSA as carrier. A specific antiserum with a high titre of 1: 2560 was obtained. Enzymelinked antibody was synthesized by NaIO4 oxidation with horseradish peroxidase as themarker enzyme. In the DAS -ELISA procedure,the optimum concentration was 5μg/mlfor IgG coating,but 2. 5 μg/ml for enzyme linked IgG. The data showed that the minimum concentration for detecting azadirachtin was 10μg/ml. This method is simple,easy,sensitive and specific, with which non - purified samples could be rapidly detected at alarge scale. This method is found suitable for screening of lines on mass call cultures.