柑橘溃疡病菌免疫荧光检测技术研究

Study on detecting Xanthomonas axonopodis pv.citri by immunofluorescence

选取不同浓度的柑橘溃疡病菌SOB培养液,利用载玻片进行固定、封闭、抗体结合、免疫荧光标记及荧光强度检测,以探讨免疫荧光检测技术在柑橘溃疡病菌快速检测方面的应用。结果表明,30℃下用BSA封闭2h,30℃下抗体结合1 h,荧光抗体浓度为4μg/mL,室温放置40 m in后,在荧光显微镜下可以清楚看到绿色的柑橘溃疡病菌体。该方法操作简便,检测过程仅需4 h,抗原抗体反应特异性极强,检测灵敏度可达103cfu/mL。

Citrus bacterial canker disease ,caused by Xanthomonas axonopodis pv. citri,is a serious quarantine disease of most citrus species and cultivars in many citrus-producing areas worldwide. The study tried to use a piece of glass to establish a simple and convenient immunofluorescent technique for a quick identification of Xanthomonas axonopodis pv. citri. Immunofluorescence experiments demonstrate that green Xanthornonas axonopodis pv. citri bacteria bodies can be seen explicitly under fluorescent microscope after enclosed by BSA under 30 C for 2 hours,combined with antibody for 1 hours,with the fluorescent antibody concentration of 4μg/mL in room temperature for 40 minutes while Xanthomonas oryzae pv. oryzae can not. The testing with immunofluorescence antibodies by this means is so simple that only a fluorescent microscope is needed and the whole testing process only takes 4 hours with an extremely strong specificity in antigen-antibody reaction and the assessing delicacy of less than 10^3 cfu/mL. Therefore,this method meets the practical demands of the rapid Citrus bacterial canker quarantine identification quite well in export and import sites.