Keighley Simon D, Li Peng, Estrela Pedro, Migliorato Piero
Electrical Engineering Division, Department of Engineering, University of Cambridge, 9 JJ Thomson Avenue, Cambridge CB3 0FA, UK.
Biosens Bioelectron. 2008 Mar 14;23(8):1291-7. doi: 10.1016/j.bios.2007.11.012. Epub 2007 Dec 4.
The ability to immobilize DNA probes onto gold substrates at an optimum surface density is key in the development of a wide range of DNA biosensors. We present a method to accurately control probe DNA surface density by the simultaneous co-immobilization of thiol modified probes and mercaptohexanol. Probe surface density is controlled by the thiol molar ratio in solution, with a linear relationship between thiol molar ratio and probe density spanning (1-9) x10(12)/cm2. The probe surface density per microscopic surface area was determined using chronocoulometry, and a detailed analysis of the method presented. Using this sample preparation method, the effect of probe density and hybridization on the charge transfer resistance with the negatively charged ferri/ferrocyanide redox couple was determined. Above a threshold probe surface density of 2.5 x 10(12)/cm2, electrostatic repulsion from the negatively charged DNA modulates the charge transfer resistance, allowing hybridization to be detected. Below the threshold density no change in charge transfer resistance with probe density or with hybridization occurs. The probe surface density was optimized to obtain the maximum percentage change in charge transfer resistance with hybridization.
以最佳表面密度将DNA探针固定在金基底上的能力是开发多种DNA生物传感器的关键。我们提出了一种通过同时共固定硫醇修饰的探针和巯基己醇来精确控制探针DNA表面密度的方法。探针表面密度由溶液中的硫醇摩尔比控制，硫醇摩尔比与探针密度之间存在线性关系，范围为(1 - 9)×10(12)/cm²。使用计时电量法测定每个微观表面积的探针表面密度，并对所提出的方法进行了详细分析。使用这种样品制备方法，确定了探针密度和杂交对带负电荷的铁氰化铁/亚铁氰化铁氧化还原对的电荷转移电阻的影响。在2.5×10(12)/cm²的阈值探针表面密度以上，带负电荷的DNA产生的静电排斥会调节电荷转移电阻，从而能够检测杂交情况。在阈值密度以下，电荷转移电阻不会随探针密度或杂交而发生变化。优化探针表面密度以获得杂交时电荷转移电阻的最大百分比变化。
