Capturing real-time electron transfer, enzyme activity, molecular dynamics, and biochemical messengers in living cells is essential for understanding the signaling pathways and cellular communications. However, there is no generalizable method for characterizing a broad range of redox-active species in a single living cell at the resolution of cellular compartments. Although nanoelectrodes have been applied in the intracellular detection of redox-active species, the fabrication of nanoelectrodes to maximize the signal-to-noise ratio of the probe remains challenging because of the stringent requirements of 3D fabrication. Here, we report an asymmetric nanopore electrode-based amplification mechanism for the real-time monitoring of NADH in a living cell. We used a two-step 3D fabrication process to develop a modified asymmetric nanopore electrode with a diameter down to 90 nm, which allowed for the detection of redox metabolism in living cells. Taking advantage of the asymmetric geometry, the above 90% potential drop at the two terminals of the nanopore electrode converts the faradaic current response into an easily distinguishable bubble-induced transient ionic current pattern. Therefore, the current signal was amplified by at least 3 orders of magnitude, which was dynamically linked to the presence of trace redox-active species. Compared to traditional wire electrodes, this wireless asymmetric nanopore electrode exhibits a high signal-to-noise ratio by increasing the current resolution from nanoamperes to picoamperes. The asymmetric nanopore electrode achieves the highly sensitive and selective probing of NADH concentrations as low as 1 pM. Moreover, it enables the real-time nanopore monitoring of the respiration chain (i.e., NADH) in a living cell and the evaluation of the effects of anticancer drugs in an MCF-7 cell. We believe that this integrated wireless asymmetric nanopore electrode provides promising building blocks for the future imaging of electron transfer dynamics in live cells.

Solid-state nanopore-based techniques have become a promising strategy for diverse single molecule detections. Owing to the challenge in well and rapid fabrication of solid-state nanopores with the diameter less than 2 nm, small molecule detection is hard to be addressed by existing label-free nanopore methods. In this work, we for the first time propose a metal-coated wireless nanopore electrode (WNE) which offers a novel and generally accessible detection method for analyzing small molecules and ions at the single molecule/ion level. Here, a silver-coated WNE is developed as a proof-of-principle model which achieves the detection the self-generated H, the smallest known molecule, and Ag at single molecule/ion level by monitoring the enhanced ionic signatures. Under a bias potential of -800 mV, the WNE could accomplish the distinction of as low as 14 H molecules and 28 Ag from one spike signal. The finite element simulation is introduced to suggest that the generation of H at the orifice of the WNE results in the enhanced spike of ionic current. As a proof-of-concept experiment, the WNE is further utilized to directly detect Hg from 100 pM to 100 nM by monitoring the frequency of the spike signals. This novel nanoelectrode provides a brand new label-free, ultrasensitive, and simple detection mechanism for various small molecules/ions detection, especially for redox analytes.

Clarifying the hidden but intrinsic feature of single nanoparticles by nanoelectrochemistry could help understand its potential for diverse applications. The uncontrolled interface and bandwidth limitation in the electrochemical measurement put the obstacle in single particle collision. Here, we demonstrate a well-defined 30 nm nanopore electrode with a rapid chemical-electrochemical fabrication method which provides a high reproducibility in both size and performance. A capacitance-based detection mechanism is demonstrated to achieve a high current resolution of 0.6 pA ±0.1 pA (RMS) and a high the temporal resolution of 0.01 ms. By utilizing this electrode, the dynamic interactions of every single particle in the mixture could be directly read during the collision process. The collision frequency is two orders of magnitude higher than previous reports, which helps reveal the hidden features of nanoparticles during the complex and multidimensional interaction processes.

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The nanopipette has been employed for the single molecule analysis due to its advantage of easy fabrication and controllable diameter. Herein, we present that the single molecule immunoreaction could be monitored by using the quartz nanopipette through the discrimination of characteristic blockade current, which reflect the intrinsic character of the individual unlabeled protein molecules due to its heterogeneous motion in solution. Our methods show the ability to monitor the immunoreaction between single α-fetal protein (AFP) and its specific antibody in aqueous solution without any labeling. Our studies may open a new door to comprehensively understand the single molecule immunoreaction, which gain more insight into the molecular dynamic of elementary steps.

Understanding the single molecular protein-protein interaction has great significance in evaluating the affinity of a specific antibody. Herein, the interaction between single α-fetal protein (AFP) and its antibody was monitored via transient ionic current recording by using the antibody functionalized nanopore sensors. More importantly, the kinetic evaluation was performed at the single molecule level to determine the dissociation constant of this interaction. This method enables the monitoring of the kinetic antigen-antibody interaction in their heterogenetic state without any labelling. Our results provided new insights into the evaluation of the antibody's binding affinity and more into the development of immunoassays for diagnostics.

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The instability of organic field-effect transistors (OFETs) is one key obstacle to practical application and is closely related to the unstable aggregate state of organic semiconductors (OSCs). However, the underlying reason for this instability remains unclear, and no effective solution has been developed. Herein, we find that the intrinsic tensile and compressive strains that exist in OSC films are the key origins for aggregate state instability and device degradation. We further report a strain balance strategy to stabilize the aggregate state by regulating film thickness, which is based on the unique transition from tensile strain to compressive strain with increasing film thickness. Consequently, a strain-free and ultrastable OSC film is obtained by regulating the film thickness, with which an ultrastable OFET with a five-year lifetime is realized. This work provides a deeper understanding of and a solution to the instability of OFETs and sheds light on their industrialization.