Department of Chemical & Biological Engineering, Polytechnic Institute of NYU, 6 MetroTech Center, Brooklyn, New York 11201, USA.
Langmuir. 2010 Sep 7;26(17):14351-8. doi: 10.1021/la1014384.
Impedance and cyclic voltammetry methods, complemented by Poisson-Boltzmann (PB) modeling, are used to study hybridization of DNA analyte strands to monolayers of morpholino oligomers (MOs) immobilized by one end to mercaptopropanol-passivated gold electrodes. MOs, like peptide nucleic acids (PNAs), are uncharged molecules that recognize nucleic acids following conventional base-pairing rules. The capacitive response to hybridization, determined from real-time impedance measurements, is analyzed with emphasis on understanding the underlying structural changes and on providing a foundation for label-free diagnostics. The capacitive response is correlated with the instantaneous surface molecular populations by labeling DNA and MO strands with ferrocene tags and using cyclic voltammetry to monitor their respective coverages in real-time. This approach allows analysis of hybridization-induced changes in interfacial capacitance as a function of duplex coverage, the DC bias used for readout, buffer molarity, and probe coverage. The results indicate that unhybridized MO layers exist in a compact state on the solid support. For hybridized layers, the intrinsic signal per hybridization event is strongly enhanced at low ionic strengths but, interestingly, does not depend on the readout bias in the sampled range negative of the capacitive minimum. A PB model incorporating an effective medium description of the hybridizing films is used to establish how hybridization-derived changes in dielectric composition and charge distribution at the surface translate into experimentally observed variations in interfacial capacitance.
采用阻抗和循环伏安法，辅以泊松-玻尔兹曼（PB）建模，研究了 DNA 分析物链与通过一端固定在巯基丙醇钝化金电极上的吗啉代寡聚物（MO）单层杂交。MO 与肽核酸（PNA）一样，是不带电荷的分子，它们按照常规碱基配对规则识别核酸。通过实时阻抗测量确定的杂交电容响应，通过强调理解基础结构变化并为无标记诊断提供基础来进行分析。通过用二茂铁标签标记 DNA 和 MO 链，并使用循环伏安法实时监测它们各自的覆盖率，将电容响应与瞬时表面分子种群相关联。这种方法允许分析杂交诱导的界面电容变化作为双链体覆盖率、用于读出的直流偏压、缓冲摩尔浓度和探针覆盖率的函数。结果表明，未杂交的 MO 层在固体支撑物上处于紧凑状态。对于杂交层，在低离子强度下，每个杂交事件的固有信号得到了极大增强，但有趣的是，它不取决于在电容最小值的负采样范围内的读出偏压。采用包含混合膜的有效介质描述的 PB 模型，建立了如何将表面的介电组成和电荷分布的杂交衍生变化转化为实验观察到的界面电容变化。
