AbstractA fluorometric assay is described for the detection of the food pathogen Pseudomonas aeruginosa (P. aeruginosa). It is based on the hybridization of aptamer and fluorescein-labeled complementary DNA (FAM-cDNA) in combination with magnetic separation. In the absence of P. aeruginosa, FAM-cDNA is assembled on the surface of aptamer modified magnetic particles (MNPs) via hybridization between aptamer and cDNA. Upon addition of P. aeruginosa, FAM-cDNA is replaced by the bacteria and released from the MNPs since the aptamer preferentially binds to bacteria. After magnetic separation, the amount of bacteria can be quantified by determination of the fluorescence intensity (λexc/em = 494/525 nm) of the supernatant containing the released FAM-cDNA. This kind of assay allows for both selective enrichment and sensitive fluorometric determination of bacteria in a single step. The assay has a response to the logarithm of P. aeruginosa concentration that is linear in the range between 10 and 108 cfu·mL−1, with a detection limit as low as 1 cfu·mL−1. The detection process can be finished within <1.5 h. The feasibility of the assay was verified by detecting P. aeruginosa in spiked food samples. Graphical abstractHybridization of aptamer and carboxyfluorescein labeled complementary DNA is combined with magnetic separation for detection of as low as 1 cfu·mL−1Pseudomonas aeruginosa. This kind of assay allows for both selective enrichment and sensitive fluorometric determination of bacteria in a single step.

中文翻译：

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使用适体修饰的磁性纳米粒子选择性捕获和灵敏荧光测定铜绿假单胞菌

摘要描述了一种用于检测食物病原体铜绿假单胞菌 (P. aeruginosa) 的荧光测定法。它基于适体和荧光素标记的互补 DNA (FAM-cDNA) 的杂交以及磁分离。在没有铜绿假单胞菌的情况下，FAM-cDNA 通过适体和 cDNA 之间的杂交组装在适体修饰的磁性颗粒 (MNP) 的表面。添加铜绿假单胞菌后，FAM-cDNA 被细菌取代并从 MNP 中释放，因为适体优先与细菌结合。磁性分离后，可以通过测定含有释放的 FAM-cDNA 的上清液的荧光强度 (λexc/em = 494/525 nm) 来量化细菌的数量。这种检测允许在一个步骤中选择性富集和灵敏地荧光测定细菌。该测定对铜绿假单胞菌浓度对数的响应在 10 到 108 cfu·mL-1 之间呈线性，检测限低至 1 cfu·mL-1。检测过程可在<1.5 h 内完成。通过检测加标食品样品中的铜绿假单胞菌，验证了该测定的可行性。图解摘要适体和羧基荧光素标记的互补 DNA 杂交与磁分离相结合，可检测低至 1 cfu·mL-1 铜绿假单胞菌。这种检测允许在一个步骤中选择性富集和灵敏地荧光测定细菌。铜绿假单胞菌浓度在 10 到 108 cfu·mL-1 之间呈线性，检测限低至 1 cfu·mL-1。检测过程可在 <1.5 h 内完成。通过检测加标食品样品中的铜绿假单胞菌，验证了该测定的可行性。图解摘要适体和羧基荧光素标记的互补 DNA 杂交与磁分离相结合，可检测低至 1 cfu·mL-1 铜绿假单胞菌。这种检测允许在一个步骤中选择性富集和灵敏地荧光测定细菌。铜绿假单胞菌浓度在 10 到 108 cfu·mL-1 之间呈线性，检测限低至 1 cfu·mL-1。检测过程可在 <1.5 h 内完成。通过检测加标食品样品中的铜绿假单胞菌，验证了该测定的可行性。图解摘要适体和羧基荧光素标记的互补 DNA 杂交与磁分离相结合，可检测低至 1 cfu·mL-1 铜绿假单胞菌。这种检测允许在一个步骤中选择性富集和灵敏地荧光测定细菌。图解摘要适体和羧基荧光素标记的互补 DNA 杂交与磁分离相结合，可检测低至 1 cfu·mL-1 铜绿假单胞菌。这种检测允许在一个步骤中选择性富集和灵敏地荧光测定细菌。图解摘要适体和羧基荧光素标记的互补 DNA 的杂交与磁分离相结合，可检测低至 1 cfu·mL-1 铜绿假单胞菌。这种检测允许在一个步骤中选择性富集和灵敏地荧光测定细菌。