In this work, an amplified electrochemical ratiometric aptasensor for nuclear factor kappa B (NF-κB) assay based on target binding-triggered ratiometric signal readout and polymerase-assisted protein recycling amplification strategy is described. To demonstrate the effect of “signal-off” and “signal-on” change for the dual-signal electrochemical ratiometric readout, the thiol-hairpin DNA (SH-HD) hybridizes with methylene blue (MB)-modified protection DNA (MB-PD) to form capture probes, which is rationally introduced for the construction of the assay platform. On the interface, the probes can specifically bind to target NF-κB and expose a toehold region which subsequently hybridizes with the ferrocene (Fc)-modified DNA strand to take the Fc group to the electrode surface, accompanied by displacing MB-PD to release the MB group from the electrode surface, leading to the both “signal-on” of Fc (I_Fc) and “signal-off” of MB (I_MB). In order to improve the sensitivity of the electrochemical aptasensor, phi29-assisted target protein recycling amplification strategy was designed to achieve an amplified ratiometric signal. With the above advantages, the prepared aptasensor exhibits a wide linear range of 0.1 pg mL⁻¹ to 15 ng mL⁻¹ with a low detection limit of 0.03 pg mL⁻¹. This strategy provides a simple and ingenious approach to construct ratiometric electrochemical aptasensor and shows promising potential applications in multiple disease marker detection by changing the recognition probe.

在这项工作中，描述了一种基于靶标结合触发的比率式信号读数和聚合酶辅助的蛋白质再循环扩增策略的用于核因子κB（NF-κB）测定的扩增电化学比率式适体传感器。为了证明“信号关闭”和“信号开启”变化对双信号电化学比例读数的影响，硫醇-发夹DNA（SH-HD）与亚甲基蓝（MB）修饰的保护DNA（MB- PD）形成捕获探针，为构建检测平台合理引入了捕获探针。在界面上，探针可以特异性结合靶NF-κB并露出脚趾区域，该脚趾区域随后与二茂铁（Fc）修饰的DNA链杂交，将Fc基团带到电极表面，I _Fc）和MB（I _MB）的“信号关闭” 。为了提高电化学适体传感器的灵敏度，设计了phi29辅助靶蛋白回收扩增策略，以实现放大的比例信号。具有上述优点，将制备的适体传感器表现出0.1微克mL的线性范围宽^-1至15纳克毫升^-1与0.03微克mL的低检测限^-1。该策略提供了一种简单而巧妙的方法来构建比率式电化学适体传感器，并通过改变识别探针显示了在多种疾病标记检测中的有希望的潜在应用。