Asymmetric Nanopore Electrode-Based Amplification for Electron Transfer Imaging in Live Cells

Abstract

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.

Counting and Sizing of Single Vesicles/Liposomes by Electrochemical Events

“…Two types of current transients are considered as the consequence of the balance of electric field force controlled by the applied potential, elastic force originating from the collision, and electroosmotic force related to the charge of the nanopipette. [95,96] Compared with the traditional resistive-pulse technique based on translocation, methods based on the observations in this study could provide more information for particle screening according to the current transients by using one nanopipette. Moreover, for PS particles of the same size, two different current transients could also be obtained by changing the bias potential ( Figure 5C).…”

Single‐Molecule Sensing with Nanopore Confinement: From Chemical Reactions to Biological Interactions

“…More recently,awireless nanopore electrode (WNE)t hat introduces the bipolar electrochemical processes into nanopore was proposed for detection of small redox molecules/ionsa nd monitoro fs ingle-cell metabolism with ultra-sensitivity. [67,68] The WNE coated with am etal layer in the interior surface of nanopore provides ah ighly confined electric field where the bipolar redox reactions can happen. For example, H 2 and Ag + are occurring at the two terminals of silver coatedW NE under as ufficient potential (Figure 4a).…”

Enzyme and AIEgens Modulated Solid‐State Nanochannels: In Situ and Noninvasive Monitoring of H2O2 Released from Living Cells

“…The recent decades have seen the great development of sensing field using functionalized solid‐state nanochannels . So far, the functionalized solid‐state nanochannels have revealed tremendous abilities in the sensing of ions, small biomolecules, biological macromolecules, and even cellular targets with the unique merits of rapid response, high sensitivity, miniaturization and low‐cost . However, to fulfill cellular target detection, the current test platform requires at least two steps ( Scheme a).…”

“…2) Adding into left chamber: When we add the extracted targets into detection device, omissions or pollution of samples may lead the irreversible mistake . Although nanopore electrodes or nanopipette have been successfully utilized in detection of intracellular target without sample pretreatment process, the insertion of nanoprobes to living cells is inevitable. In order to address the above limitations and keep the cells intact, in situ and noninvasive platform is desired.…”