Kim Hanyoup, Kane Michael D, Kim Sol, Dominguez Wilfredo, Applegate Bruce M, Savikhin Sergei
Department of Physics, Purdue University, West Lafayette, IN 47907, USA.
Biosens Bioelectron. 2007 Jan 15;22(6):1041-7. doi: 10.1016/j.bios.2006.04.032. Epub 2006 Jul 11.
A DNA hybridization based optical detection platform for the detection of foodborne pathogens has been developed with virtually zero probability of the false negative signal. This portable, low-cost and real-time assaying detection platform utilizes the color changing molecular beacon as a probe for the optical detection of the target sequence. The computer-controlled detection platform exploits the target hybridization induced change of fluorescence color due to the Förster (fluorescence) resonance energy transfer (FRET) between a pair of spectrally shifted fluorophores conjugated to the opposite ends of a beacon (oligonucleotide probe). Unlike the traditional fluorophore-quencher beacon design, the presence of two fluorescence molecules allows to actively visualize both hybridized and unhybridized states of the beacon. This eliminates false negative signal detection characteristic for the fluorophore-quencher beacon where bleaching of the fluorophore or washout of a beacon is indistinguishable from the absence of the target DNA sequence. In perspective, the two-color design allows also to quantify the concentration of the target DNA in a sample down to < =1 ng/microl. The new design is suitable for simultaneous reliable detection of hundreds of DNA target sequences in one test run using a series of beacons immobilized on a single substrate in a spatial format.
一种基于DNA杂交的食源性病原体光学检测平台已被开发出来，其假阴性信号的概率几乎为零。这个便携式、低成本且实时检测的平台利用变色分子信标作为探针来光学检测目标序列。该计算机控制的检测平台利用了目标杂交引起的荧光颜色变化，这是由于一对与信标（寡核苷酸探针）两端共轭的光谱位移荧光团之间的福斯特（荧光）共振能量转移（FRET）。与传统的荧光团-猝灭剂信标设计不同，两个荧光分子的存在使得能够主动观察信标的杂交和未杂交状态。这消除了荧光团-猝灭剂信标假阴性信号检测的特性，在这种情况下，荧光团的漂白或信标的洗脱与目标DNA序列不存在时无法区分。从长远来看，这种双色设计还能够将样品中目标DNA的浓度定量至<=1 ng/微升。这种新设计适用于在一次测试运行中，使用一系列以空间形式固定在单个底物上的信标，同时可靠地检测数百个DNA目标序列。
