Carter Darren J, Cary R Bruce
Biosciences Division, Los Alamos National Laboratory, Los Alamos, New Mexico 87545, USA.
Nucleic Acids Res. 2007;35(10):e74. doi: 10.1093/nar/gkm269. Epub 2007 May 3.
Widely used nucleic acid assays are poorly suited for field deployment where access to laboratory instrumentation is limited or unavailable. The need for field deployable nucleic acid detection demands inexpensive, facile systems without sacrificing information capacity or sensitivity. Here we describe a novel microarray platform capable of rapid, sensitive nucleic acid detection without specialized instrumentation. The approach is based on a miniaturized lateral flow device that makes use of hybridization-mediated target capture. The miniaturization of lateral flow nucleic acid detection provides multiple advantages over traditional lateral flow devices. Ten-microliter sample volumes reduce reagent consumption and yield analyte detection times, excluding sample preparation and amplification, of <120 s while providing sub-femtomole sensitivity. Moreover, the use of microarray technology increases the potential information capacity of lateral flow. Coupled with a hybridization-based detection scheme, the lateral flow microarray (LFM) enables sequence-specific detection, opening the door to highly multiplexed implementations for broad-range assays well suited for point-of-care and other field applications. The LFM system is demonstrated using an isothermal amplification strategy for detection of Bacillus anthracis, the etiologic agent of anthrax. RNA from as few as two B. anthracis cells was detected without thermocycling hardware or fluorescence detection systems.
广泛使用的核酸检测方法不太适合在实验室仪器设备有限或无法获取的现场进行部署。对于可现场部署的核酸检测的需求,要求有廉价、简便的系统,同时又不牺牲信息容量或灵敏度。在此,我们描述了一种新型微阵列平台,它能够在无需专门仪器的情况下快速、灵敏地检测核酸。该方法基于一种小型化的侧向流动装置,该装置利用杂交介导的靶标捕获。与传统侧向流动装置相比,侧向流动核酸检测的小型化具有多个优点。十微升的样品体积减少了试剂消耗,并缩短了分析物检测时间(不包括样品制备和扩增),检测时间小于120秒,同时提供亚飞摩尔级别的灵敏度。此外,微阵列技术的使用增加了侧向流动的潜在信息容量。结合基于杂交的检测方案,侧向流动微阵列(LFM)能够进行序列特异性检测,为适用于即时检测和其他现场应用的广泛检测的高度多重化实施方案打开了大门。使用等温扩增策略对炭疽杆菌(炭疽的病原体)进行检测,展示了LFM系统。在没有热循环硬件或荧光检测系统的情况下,检测到了少至两个炭疽杆菌细胞的RNA。