摘要
分离纯化抗副溶血弧菌IgY,并建立一种快速检测副溶血弧菌的间接酶联免疫吸附法(indirect ELISA),分析了该方法的敏感性、特异性和重复性.将水稀释法、乙醇沉淀法和DEAE-Sepharose FF离子交换层析联用以分离纯化抗副溶血弧菌IgY,然后以纯化的IgY为一抗,辣根过氧化物酶标兔抗鸡IgY为二抗,建立检测副溶血弧菌的间接ELISA方法.结果表明:建立的间接ELISA方法,一抗的最佳工作质量浓度为15μg/mL,二抗的最佳稀释度为1∶10 000,副溶血弧菌培养液的检出限为1.0×105cfu/mL,该方法对测定的其他8种菌株没有交叉反应.建立的间接ELISA方法具有良好的灵敏度、特异性和稳定性,能够快速、准确地对副溶血弧菌进行检测.
The objectives of the present study were to establish an indirect ELISA for the detection of Vibrio parahaemololyticus, and analyse the specificity, sensitivity and reliability of the method. Water dilufion method, ethanol precipitation and DEAE-Sepharose FF chromatography were used to extract and purify the anti-V, parahaemololyticus IgY. Then the purified anti-V, parahaemololyticus IgY was used as the first antibody and rabbit anti-chicken IgY-HRP was used as the second antibody. The suitable concentration of anti-V, parahaemololyticus IgY and rabbit anti-chicken IgY-HRP were decided, and the indirect ELISA for the detection of V. parahaemoloIyticus was established. The suitable mass concentration of the first antibody was 15 μg/mL, and the dilution of the second antibody was 1 ： 10 000. The detection limits of this method were 1.0 × 10^5 cfu/mL, and the specificity of the method was also determined, V. parahaemololyticus can be detected, while other bacteria strains can not be detected. The established indirect ELISA is stable, highly specific and sensitive, and it can detect V. parahaemololyticus quickly.
出处
《华南农业大学学报》
CAS
CSCD
北大核心
2008年第4期95-99,共5页
Journal of South China Agricultural University
