关键词: 间接酶联免疫吸附法, 斑点免疫法, 碱性磷酸酶, 辣根过氧化物酶, 聚苯乙烯板, 硝酸纤维膜

Abstract: The effects of second antibody conjugated with two enzymes (horseradish peroxidase and alkaline phosphatase) and two solid supports (polystyrene micro-well plate and nitrocellulose membrane) on sensitivity and detection results were investigated. The results show that when the pure Bt insecticidal protein is assayed by a indirect ELISA, there are no significant differences in sensitivity. The sensitivity of second antibody conjugated with alkaline phosphatase is equal to that of the second antibody conjugated with horseradish peroxidase. However, when the Bt insecticidal protein in cotton leaves is assayed on polystyrene plate, the detection value of horseradish peroxidase is higher than that of alkaline phosphatase. The reason is endogenous peroxidase activity in plant. There are some problems caused by chlorophyll in plant extract from leaf tissue when Dot-ELISA on nitrocellulose membrane is used to detect insecticidal protein in plant specimen. The antibody of Bt insecticidal protein prepared in this study is high specific.

Key words: indirect ELISA, Dot-ELISA, alkaline phosphatase, horseradish peroxidase, polystyrene micro-well plate, nitrocellulose membrane