The label-free analysis of biomarkers offers important advantages in developing point-of-care (PoC) biosensors. In contrast to label-based methodologies, such as ELISA, label-free analysis enables direct detection of targets without additional steps and labeled reagents. Nonetheless, label-free approaches require high sensitivity to detect the intrinsic features of a biomarker and low levels of nonspecific signals. Electrochemical capacitance, \(C_{\bar \mu }\), is a feature of electroactive nanoscale films that can be measured using electrochemical impedance spectroscopy. \(C_{\bar \mu }\) is promising as an electrochemical transducing signal for the development of high-sensitivity, reagentless and label-free molecular diagnostic assays. We used a proprietary ferrocene (Fc)-tagged peptide that is able to self-assemble onto gold electrodes (thicknesses <2 nm) to which any biological receptor can be coupled. When coupled with biological receptors (e.g., a monoclonal antibody), \(C_{\bar \mu }\) exhibited by the redox-tagged peptide changes as a function of the target concentration. We provide herein the steps for the qualitative and quantitative detection of dengue non-structural protein 1 (NS1) biomarker. Detection of NS1 can be used to diagnose dengue virus infection, which causes epidemics each year in tropical and subtropical regions of the world. Including the pre-treatment of the electrode surface, the analysis takes ~25 h. This time can be reduced to minutes if the electrode surface is fabricated separately, demonstrating that \(C_{\bar \mu }\) is promising for PoC applications. We hope this protocol will serve as a reference point for researchers and companies that intend to further develop capacitive devices for molecular diagnostic assays.

生物标志物的无标记分析在开发护理点 (PoC) 生物传感器方面具有重要优势。与基于标记的方法（如 ELISA）相比，无标记分析无需额外步骤和标记试剂即可直接检测目标。尽管如此，无标记方法需要高灵敏度来检测生物标志物的内在特征和低水平的非特异性信号。电化学电容\(C_{\bar \mu }\)是电活性纳米级薄膜的一个特征，可以使用电化学阻抗谱测量。\(C_{\bar \mu }\)作为一种电化学转导信号，有望用于开发高灵敏度、无试剂和无标记的分子诊断分析。我们使用了一种专有的二茂铁 (Fc) 标记肽，该肽能够自组装到金电极（厚度 < 2 nm）上，任何生物受体都可以与之耦合。当与生物受体（例如单克隆抗体）偶联时，\(C_{\bar \mu }\)由氧化还原标记的肽所表现出的变化是目标浓度的函数。我们在此提供了定性和定量检测登革热非结构蛋白 1 (NS1) 生物标志物的步骤。NS1的检测可用于诊断登革热病毒感染，登革热病毒感染每年都会在世界热带和亚热带地区引起流行病。包括电极表面的预处理，分析时间约为 25 小时。如果电极表面是单独制造的，这个时间可以减少到几分钟，证明\(C_{\bar \mu }\)对于 PoC 应用很有前景。我们希望该协议能够为打算进一步开发用于分子诊断检测的电容设备的研究人员和公司提供参考。