DNA hybridization biosensors using polylysine modified SPCEs.
DNA hybridization biosensors using polylysine modified SPCEs.
复制标题
DOI：
10.1016/j.bios.2007.12.001
复制
发表时间：
2008-04-15
影响因子：
12.6
通讯作者：
Costa-García A
中科院分区：
文献类型：
--
作者：
Díaz-González M;de la Escosura-Muñiz A;González-García MB;Costa-García A
本工作描述了两种用于检测严重急性呼吸道综合征（SARS）病毒特有的30聚体序列的电化学DNA杂交生物传感器（基因传感器）。这两种基因传感器都依赖于寡核苷酸靶与其互补探针的杂交，该互补探针通过静电相互作用固定在带正电荷的聚赖氨酸修饰的丝网印刷碳电极（SPCE）上。在一种设计中，使用生物素化的靶标，并使用碱性磷酸酶标记的链霉亲和素（S-AP）监测杂交反应的检测。这种酶催化底物3-吲哚酚磷酸（3-IP）水解为靛蓝，然后溶解为靛蓝胭脂红，并通过循环伏安法（CV）检测。在另一种设计中，使用Au（I）络合物、硫代金苹果酸钠标记靶，并且通过首次测量由金标记催化的析氢产生的电流来检测双链体形成。使用30分钟的杂交时间，计算出酶基因传感器的检测限为8 pM。虽然这种良好的灵敏度不能用金属标记物（0.5 nM）达到，但使用这种标记物可以大大减少分析时间。两种基因传感器都不需要修饰寡核苷酸探针，并且使用严格的实验条件（60分钟的杂交时间和在杂交缓冲液中50%甲酰胺）可以区分互补寡核苷酸和具有三碱基错配的寡核苷酸。
Two electrochemical DNA hybridization biosensors (genosensors) for the detection of a 30-mer sequence unique to severe acute respiratory syndrome (SARS) virus are described in this work. Both genosensors rely on the hybridization of the oligonucleotide target with its complementary probe, which is immobilized on positively charged polylysine modified screen-printed carbon electrodes (SPCEs), through electrostatic interactions. In one design, a biotinylated target is used and the detection of the hybridization reaction is monitored using alkaline phosphatase labeled streptavidin (S-AP). This enzyme catalyzes the hydrolysis of the substrate 3-indoxyl phosphate (3-IP) to indigo, which is then solubilized to indigo carmine and detected by means of cyclic voltammetry (CV). In the other design, the target is labeled using an Au(I) complex, sodium aurothiomalate, and the duplex formation is detected by measuring, for first time, the current generated by the hydrogen evolution catalyzed by the gold label. Using 30 min of hybridization time, a detection limit of 8 pM is calculated for the enzymatic genosensor. Although this good sensitivity cannot be reached with the metal label (0.5 nM), the use of this label allows a considerable decrease of the analysis time. Both genosensors do not require the modification of the oligonucleotide probe and using stringent experimental conditions (60 min of hybridization time and 50% formamide in the hybridization buffer) can discriminate between a complementary oligonucleotide and an oligonucleotide with a three-base mismatch.
登录
查看更多内容
影响因子：
12.6
作者：
Cai, H;Zhu, NN;Fang, YZ
通讯作者：
Fang, YZ
影响因子：
3
作者：
de la Escosura-Muñiz, A;González-García, MB;Costa-García, A
通讯作者：
Costa-García, A
影响因子：
6.2
作者：
Babkina, SS;Ulakhovich, NA;Zyavkina, YI
通讯作者：
Zyavkina, YI
影响因子：
10.8
作者：
Segura, T;Volk, MJ;Shea, LD
通讯作者：
Shea, LD
影响因子：
7.4
作者：
de-Los-Santos-Alvarez, P;Lobo-Castañón, MJ;Tuñón-Blanco, P
通讯作者：
Tuñón-Blanco, P