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基于纳米磁珠标记的免疫层析法,建立快速定量灵敏检测玉米内州萎蔫病菌(Clavibacter michiganensis subsp.nebraskensis,Cmn)的层析试纸条检测方法。用碳二亚胺(EDC)/N-羧基琥珀酰亚胺(NHS)交联法,使玉米内州萎蔫病菌单抗4G12偶联在纳米磁珠表面上,使用喷膜仪将玉米内州萎蔫病菌单抗4H4和羊抗小鼠IgG喷涂于硝酸纤维素膜上,分别作为检测线和质控线,基于双抗夹心法原理建立免疫层析试纸条。通过对检测线上超顺纳米磁珠的磁信号进行检测和结果分析,Cmn抗原最低检测限度为1.0×105 CFU/mL,特异性良好且检测时间缩短为5 min。该方法同时可根据检测线的光学反应进行定性判断,具有操作简便、灵敏度高等优点,并可根据磁信号强度定量检测实际玉米种子样品中Cmn的浓度,为植物病原细菌的快速高效检测提供了一种新型的检测方法。
A rapid and sensitive method for the determination of Clavibacter michiganensis subsp. Nebraskensis (Cmn) was established based on the nanocarrier-labeled immunochromatography. The Maize Wilt 4G4 was coupled to the surface of nanospheres by carbodiimide (EDC) / N-carboxy succinimide (NHS) cross-linking method. Pathogen McAb 4H4 and goat anti-mouse IgG were sprayed on nitrocellulose membrane as test line and control line respectively, and immunochromatographic test strips were established based on the double-antibody sandwich method. By detecting and analyzing the magnetic signal of the supernanometer magnetic beads on the detection line, the minimum detection limit of Cmn antigen was 1.0 × 10 5 CFU / mL, the specificity was good and the detection time was shortened to 5 min. The method can simultaneously judge the optical reaction of the test line qualitatively, has the advantages of simple operation and high sensitivity, and can quantitatively detect the concentration of Cmn in the corn seed sample according to the magnetic signal intensity, thereby providing a rapid and efficient detection of plant pathogenic bacteria A new detection method.