摘要
目的:制备黄瓜细菌性白枯病菌特异性抗体,建立其酶联免疫吸附分析(ELISA)的检测方法。方法:分别以黄瓜细菌性白枯病菌胞内蛋白和菌体为免疫原,制备两种抗体,优化间接ELISA检测条件。结果:间接ELISA法测定两种纯化的菌体抗体(J4)和胞内蛋白抗体(P2)效价分别为1∶32 000和1∶4 000,二抗的最佳稀释度为1∶3 000,方阵试验表明J4的工作浓度为1∶16 000,抗原的最佳包被浓度为106 cfu/ml,P2的工作浓度为1∶2 000,抗原的最佳包被浓度为107 cfu/ml,两种抗体的灵敏度均为105cfu/ml,与黄瓜细菌性角斑病菌等26个菌株无交叉反应,特异性强。结论:成功制备了黄瓜细菌性白枯病菌特异性抗体,建立了ELISA检测方法。
Objective：The specific polyclonal antibodies of Pseudomonas viridiflava on cucumber were prepared,and ELISA method was established.Method：Two antibodies were prepared by immunizing the healthy male rabbits with the intracellular protein and strain of P.viridiflava,and the detection condition of indirect ELISA were optimized.Result：The titers were respectively 1∶32 000 and 1∶4 000 of the two purified antibodies from strain（J4） and the intracellular protein（P2）.The optimum dilution of the secondary antibody was 1∶3 000.It was showed that working concentration of J4 and its antigen optimum coating concentration were 1∶16 000 and 106cfu/ml,and that of P2 were 1∶2 000 and 107 cfu/ml by the checker-board tests.The two antibodies sensitivity were both 105cfu/mL,which had high specific and no cross reaction with Pseudomonas syringae pv.lachrymans and twenty-six strains so on.Conclusion：The specific antibodies of P.viridiflava were obtained successfully in this experiment and a indirect ELISA method for specific detection of P.viridiflava was established.
出处
《生物技术》
CAS
CSCD
北大核心
2011年第2期57-60,共4页
Biotechnology
基金
辽宁省农业科学院博士后项目(200703)资助
