A fluorescent cascade amplification method for sensitive detection of Salmonella based on magnetic Fe3O4 nanoparticles and hybridization chain reaction

We developed a fluorescence sensor based on hybridization chain reaction (HCR) and Fe3O4 nanoparticles for the detection of Salmonella, common pathogen related to numerous food poisoning outbreaks. In this study, a pair of specific primers based on invA of Salmonella was designed and used in asymmetric polymerase chain reaction (aPCR) to produce a long target single-stranded DNA (ssDNA) that was captured by biotin-probe labeled magnetic beads (MBs) forming the MBs-ssDNA. The target ssDNA triggered HCR amplification with the two carboxyfluorescein (FAM) labeled DNA hairpin reporter probes (H1-FAM, H2-FAM) that enabled the measurements of the fluorescence intensities from the resulting ssDNA similar to (H1-FAM similar to H2-FAM)(n). The Fe3O4 nanoparticles were used to isolate the target ssDNA which hybridized with the reporter probes and led to decreased background signal that resulted in a detection limit of 7.4 x 10(1) CFU/mL in buffer and 6.9 x 10(2) CFU/g in spiked lettuce. The method developed also showed excellent selectivity over several common foodborne pathogens. The Fe3O4 nanoparticles capture coupled with fluorescence detection sensor showed promise for the detection of Salmonella.

我们开发了一种基于杂交链式反应（HCR）和四氧化三铁纳米粒子的荧光传感器，用于检测沙门氏菌，沙门氏菌是与众多食物中毒事件相关的常见病原体。在这项研究中，设计了一对基于沙门氏菌invA基因的特异性引物，并将其用于不对称聚合酶链式反应（aPCR），以产生长的靶标单链DNA（ssDNA），该单链DNA被生物素探针标记的磁珠（MBs）捕获，形成MBs - ssDNA。靶标单链DNA触发与两种羧基荧光素（FAM）标记的DNA发夹报告探针（H1 - FAM，H2 - FAM）的HCR扩增，从而能够测量所得类似于（H1 - FAM类似于H2 - FAM）（n）的单链DNA的荧光强度。四氧化三铁纳米粒子用于分离与报告探针杂交的靶标单链DNA，并导致背景信号降低，从而在缓冲液中的检测限为7.4×10¹菌落形成单位/毫升，在加标生菜中的检测限为6.9×10²菌落形成单位/克。所开发的方法对几种常见的食源性病原体也表现出优异的选择性。四氧化三铁纳米粒子捕获结合荧光检测传感器在沙门氏菌检测方面显示出良好的应用前景。