In this paper, we report a method to integrate the electrokinetic pre-enrichment of nucleic acids within a bed of probe-modified microbeads with their label-free electrochemical detection. In this detection scheme, hybridization of locally enriched target nucleic acids to the beads modulates the conduction of ions along the bead surfaces. This is a fundamental advancement in that this mechanism is similar to that observed in nanopore sensors, yet occurs in a bed of microbeads with microscale interstices. In application, this approach has several distinct advantages. First, electrokinetic enrichment requires only a simple DC power supply, and in combination with nonoptical detection, it makes this method amenable to point-of-care applications. Second, the sensor is easy to fabricate and comprises a packed bed of commercially available microbeads, which can be readily modified with a wide range of probe types, thereby making this a versatile platform. Finally, the sensor is highly sensitive (picomolar) despite the modest 100-fold pre-enrichment we employ here by faradaic ion concentration polarization (fICP). Further gains are anticipated under conditions for fICP focusing that are known to yield higher enrichment factors (up to 100,000-fold enrichment). Here, we demonstrate the detection of 3.7 pM single-stranded DNA complementary to the bead-bound oligoprobe, following a 30 min single step of enrichment and hybridization. Our results indicate that a shift in the slope of a current–voltage curve occurs upon hybridization and that this shift is proportional to the logarithm of the concentration of target DNA. Finally, we investigate the proposed mechanism of sensing by developing a numerical simulation that shows an increase in ion flux through the bed of insulating beads, given the changes in surface charge and zeta potential, consistent with our experimental conditions.

中文翻译：

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通过离子沿生物共轭珠表面的传导对核酸进行电动富集和无标记电化学检测

在本文中，我们报告了一种将核酸的电动预富集与探针修饰微珠床及其无标记电化学检测相结合的方法。在此检测方案中，局部富集的目标核酸与珠子的杂交调节离子沿珠子表面的传导。这是一个根本性的进步，因为这种机制类似于在纳米孔传感器中观察到的机制，但发生在具有微型间隙的微珠床中。在应用中，这种方法有几个明显的优势。首先，电动富集只需要一个简单的直流电源，结合非光学检测，使这种方法适合即时应用。其次，该传感器易于制造，并包含一个由市售微珠组成的填充床，它可以很容易地用各种探头类型进行修改，从而使其成为一个多功能平台。最后，尽管我们在这里通过法拉第离子浓差极化 (fICP) 进行适度的 100 倍预富集，但该传感器仍然非常灵敏（皮摩尔）。在已知会产生更高富集因子（高达 100,000 倍富集）的 fICP 聚焦条件下，预计会有进一步的收益。在这里，我们展示了在 30 分钟的单步富集和杂交后检测与珠结合寡核苷酸互补的 3.7 pM 单链 DNA。我们的结果表明，电流-电压曲线的斜率在杂交时发生了变化，并且这种变化与靶 DNA 浓度的对数成正比。最后，