Ye Wang, Ming Wang, Hang You, Chao Chen, Jing Zhang, Tianhua Li, Ning Gan
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引用次数: 0
摘要
基于磁性多(噬菌体)编码探针(MPEP)，开发了一种用于筛选和检测活大肠杆菌（E. coli）和副嗜血杆菌（H. paralvei）（水生环境中的两种典型病原体）的双模式检测方法。这些探针是将大量针对不同目标细菌的噬菌体分别嫁接到长链 DNA 结构上制备而成的。它们可以通过磁分离特异性地捕获和富集大肠杆菌和副大肠杆菌。然后，不同长度的 DNA 信号标签与相应的 MPEP-细菌复合物共轭，分别形成两种夹层结构。然后，将捕获的大肠杆菌和帕拉维氏菌裂解，释放出三磷酸腺苷（ATP）和 DNA 信号标签。测量包括两个步骤。首先，使用便携式 ATP 生物发光仪快速筛选含有两种目标细菌中任何一种的阳性样本。其次，只将阳性样本注入微流控芯片，芯片可检测各种 DNA 信号标签，从而准确量化目标细菌。该检测方法具有灵敏度高（大肠杆菌为 3 CFU/mL，副猪嗜血杆菌为 5 CFU/mL）、特异性高（菌株鉴定）、信号放大（20 倍）和时间短（≤ 35 分钟）等特点。只需改变 MPEP 中的相对噬菌体，就可用于检测其他病原体。此外，所提出的双模式检测方法为快速筛查和准确测定活的食源性病原体提供了广阔的前景。临床试验编号：NBDXMS-20240322。
Magnetic poly(phages) encoded probes–based dual-mode assay for rapid determination of live Escherichia coli and Hafnia paralvei based on microfluidic chip and ATP bioluminescence meter
A dual-mode assay was developed for screening and detecting live Escherichia coli (E. coli) and Hafnia paralvei (H. paralvei) (as two typical pathogens in aquatic environments) based on magnetic poly(phages) encoded probes (MPEP). The probes were prepared by grafting a large number of phages targeting different target bacteria on a long-chain DNA structure, respectively. They could specifically capture and enrich E. coli and H. paralvei by magnetic separation. Then, different DNA signal tags with different lengths conjugate with the corresponding MPEP-bacteria complex and form two kinds of sandwich structures, respectively. After that, the captured E. coli and H. paralvei were lysed to release both adenosine triphosphate (ATP) and DNA signal tags. The measurement includes two steps. Firstly, a portable ATP bioluminescence meter was employed to rapidly screen the positive samples that contain either of the two target bacteria. Secondly, only positive samples were injected into the microfluidic chip which could detect various DNA signal tags for accurate quantification of the target bacteria. The assay demonstrated high sensitivity (3 CFU/mL for E. coli and 5 CFU/mL for H. paralvei), high specificity (strain identification), signal amplification (20-fold), and short time (≤ 35 min). It can be applied to detect other pathogens solely by changing the relative phage in MPEP. Furthermore, the proposed dual-mode assay provides a wide prospect for rapid screening and accurate determination of live foodborne pathogens. Clinical Trial Number: nbdxms-20240322.
期刊介绍：
As a peer-reviewed journal for analytical sciences and technologies on the micro- and nanoscale, Microchimica Acta has established itself as a premier forum for truly novel approaches in chemical and biochemical analysis. Coverage includes methods and devices that provide expedient solutions to the most contemporary demands in this area. Examples are point-of-care technologies, wearable (bio)sensors, in-vivo-monitoring, micro/nanomotors and materials based on synthetic biology as well as biomedical imaging and targeting.
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