Flechsig Gerd-Uwe, Peter Jörg, Hartwich Gerhard, Wang Joseph, Gründler Peter
Institut für Chemie, Universität Rostock, D-18051 Rostock, Germany.
Langmuir. 2005 Aug 16;21(17):7848-53. doi: 10.1021/la051176n.
The detection of DNA hybridization is of central importance to the diagnosis and treatment of genetic diseases. Due to cost limitations, small and easy-to-handle testing devices are required. Electrochemical detection is a promising alternative to evaluation of chip data with optical readout. Independent of the actual readout principle, the hybridization process still takes a lot of time, hampering daily use of these techniques, especially in hospitals or doctor's surgery. Here we describe how direct local electrical heating of a DNA-probe-modified gold electrode affects the surface hybridization process dramatically. We obtained a 140-fold increase of alternating current voltammetric signals for 20-base ferrocene-labeled target strands when elevating the electrode temperature during hybridization from 3 to 48 degrees C while leaving the bulk electrolyte at 3 degrees C. At optimum conditions, a target concentration of 500 pmol/L could be detected. Electrothermal regeneration of the immobilized DNA-probe strands allowed repetitive use of the same probe-modified electrode. The surface coverage of DNA probes, monitored by chronocoulometry of hexaammineruthenium(III), was almost constant upon heating to 70 degrees C. However, the hybridization ability of the probe self-assembled monolayer declined irreversibly when using a 70 degrees C hybridization temperature. Coupling of heated electrodes and highly sensitive electrochemical DNA hybridization detection methods should enhance detection limits of the latter significantly.
DNA杂交检测对于遗传疾病的诊断和治疗至关重要。由于成本限制,需要小型且易于操作的检测设备。电化学检测是一种很有前景的替代方法,可用于替代通过光学读出评估芯片数据。无论实际的读出原理如何,杂交过程仍然需要大量时间,这阻碍了这些技术的日常使用,尤其是在医院或医生诊所。在此,我们描述了对DNA探针修饰的金电极进行直接局部电加热如何显著影响表面杂交过程。当在杂交过程中将电极温度从3℃升高到48℃,而使本体电解质保持在3℃时,对于20个碱基的二茂铁标记靶链,我们获得了交流伏安信号140倍的增加。在最佳条件下,可以检测到500 pmol/L的靶标浓度。固定化DNA探针链的电热再生允许重复使用同一探针修饰电极。通过六氨合钌(III)的计时电量法监测,DNA探针的表面覆盖率在加热到70℃时几乎保持恒定。然而,当使用70℃的杂交温度时,探针自组装单层的杂交能力会不可逆地下降。加热电极与高灵敏度电化学DNA杂交检测方法的结合应能显著提高后者的检测限。