Gap regulation device and method for near-atomic scale electrochemical processing of ion channel proteins and applications

Abstract

This invention relates to a gap control device, method, and application for near-atomic-scale electrochemical machining of ion channel proteins, belonging to the field of electrochemical machining technology. To address the problems of insufficient gap control precision and difficulty in achieving near-atomic-scale localized dissolution and real-time observation in existing micro / nano electrochemical machining, this invention provides a dedicated device and method. The device constructs a precisely controllable nano / sub-nanometer-scale dielectric cavity layer deposited on the surface of the workpiece anode in a TEM window, and seals it with a tool cathode containing micro-perforations to form a closed cavity capable of storing liquid. The method includes: sequentially injecting a phospholipid-containing organic solvent and an electrolyte containing ion channel proteins into the cavity; utilizing the phospholipid's self-assembly at the interface to form a biomimetic confinement layer, allowing the electrolyte to contact the workpiece anode only through the internal channels of the ion channel proteins; and applying an appropriate pulse voltage to achieve localized electrochemical dissolution of the workpiece anode within an ultrathin gap defined by the dielectric layer thickness.

Description

Technical Field

[0001] This invention belongs to the field of electrochemical processing technology, specifically relating to a gap control device, method, and application for near-atomic-scale electrochemical processing of ion channel proteins. Background Technology

[0002] Electrolytic machining is a special processing technology that uses the principle of electrochemical anodic dissolution to remove materials in the form of ion dissolution at the angstrom scale. In principle, it has the potential to manufacture at the atomic level. However, existing micro- and nano-electrochemical machining methods, such as mask micro-electrochemical machining, ultrashort pulse micro-electrochemical machining, and jet micro-electrochemical machining (SPEIDEL A, BISTEROV I, SAXENA KK, et al, 2022. Electrochemical jet manufacturing technology: From fundamentals to application[J]. International Journal of Machine Tools and Manufacture, 180: 103931), often have processing scales only at the micrometer to submicrometer scale. How to limit the dissolution reaction space of electrochemical machining to the near-atomic scale and control the electrochemical reaction process is the key to achieving controllable removal of ions / ion clusters at the atomic scale. Among them, having a nanometer or even sub-nanometer tool cathode-workpiece anode machining gap (inter-electrode gap) is an important basis for achieving reaction space limitation. This is because the charge and discharge of the double layer is very sensitive to the inter-electrode gap. Only an inter-electrode gap below the threshold can achieve effective electrochemical dissolution during electrochemical machining.

[0003] Currently, the gaps in conventional micro / nano electrolytic machining are controlled at hundreds of micrometers. For example, the gap adjustments described in a series of Chinese inventions, such as CN2024210042141, CN2021108497493, CN2020108244207, CN2020111858007, and CN2013105476236, all fall under the category of macroscopic control. However, for electrolytic machining where the removal rate is measured in angstroms, such high gaps lead to a severe attenuation of current density, making normal electrochemical dissolution impossible. This invention, however, uses chemical or physical deposition methods to control the gap at the nanometer to sub-nanometer scale, effectively solving the problem of current density attenuation.

[0004] Generally speaking, researchers have developed an AFM-based electrochemical machining method (EC-AFM) by combining the advantages of atomic force microscopy (AFM). While this method, which relies on complex equipment, has advantages in controlling the nano- to sub-nanometer machining gaps, it still faces a series of problems such as the difficulty in manufacturing nanoscale tips and the easy passivation of the tip surface, resulting in high maintenance costs (LI L, PAN C, SHAN J, et al, 2020. Pit-Induced Electrochemical Layer Dissolution and Wave Propagation on an Au(111) Surface in an Acidic Thiourea Solution[J]. The Journal of Physical Chemistry C, 124(35): 19112-19118.). The more important reason why this invention does not use AFM to control the machining gap is that the phospholipid monomers contained in the organic solvent and the ion channel proteins in the electrolyte are important components of this scheme. These substances are likely to be specifically adsorbed on the AFM probe within such a small machining gap (van der Waals forces), resulting in unpredictable experimental results. Similarly, it is difficult to use such a small processing gap in the electrochemical machining (SECCM) method using nanopipettes (WANG Y, GARCIA-CARRILLO R, REN H, 2025. Kinetics and dynamics of atomic-layer dissolution on low-defect Ag[J]. Chemical Science, 16(3): 1447-1454.), because an excessively small processing gap will cause pipette clogging and difficulty in electrolyte renewal, resulting in the inability to continue machining. The method of this invention does not rely on nanoscale AFM probes or pipettes, but uses ion channel proteins embedded in the phospholipid bilayer as a confinement space. This machining process, which combines the characteristics of the biological and physiological environment, is similar to electrochemical machining with an ultrathin mask, and naturally does not have the problem of electrolyte renewal.

[0005] In addition, another advantage of this method is that the overall thickness of the workpiece anode-tool cathode-electrolyte layer can be flexibly adjusted according to actual needs to meet the requirements of in-situ TEM electrochemical machining for atomic-scale analysis of electrochemical removal mechanisms. Based on this, in order to limit the inter-electrode gap in near-atomic-scale electrochemical machining, ensuring the stable and continuous occurrence of electrochemical reactions within the confined space at the near-atomic scale, while also enabling online in-situ observation of the mechanisms and novel phenomena of electrochemical reactions, this invention proposes a near-atomic-scale electrochemical machining method that synergistically controls the machining gap and cathode thickness. Summary of the Invention

[0006] Technical problem solved: This invention provides a gap control device, method and application for near-atomic scale electrochemical processing of ion channel proteins. By controlling the gap and cathode thickness in nano- to sub-nano-scale processing, it enables the stable and sustainable occurrence of electrochemical reactions within a confined space while also taking into account the function of in-situ electrochemical removal mechanism. This opens up a new method for micro-nano electrochemical processing and releases the ultimate capabilities of electrochemical processing.

[0007] Technical Solution: An apparatus for near-atomic-scale electrochemical processing of ion channel proteins, comprising: a silicon nitride thin film window; a workpiece anode, which is an ultrathin metal film deposited on the window; a dielectric cavity layer formed on the surface of the workpiece anode, the thickness of the dielectric cavity layer being precisely controlled to define the processing gap and patterned to form a cavity for containing liquid; and a tool cathode comprising a supporting substrate and a suspended metal film formed thereon, the tool cathode having a liquid injection channel communicating with the cavity; wherein the suspended metal film side of the tool cathode is aligned and sealed to the dielectric cavity layer, such that the workpiece anode, the sidewall of the dielectric cavity layer, and the suspended metal film of the tool cathode together form a sealed cavity.

[0008] A method for gap control in near-atomic-scale electrochemical machining of ion channel proteins using the aforementioned device includes the following steps: S1: providing a silicon nitride thin film window with a workpiece anode deposited thereon; S2: forming a dielectric cavity layer of controllable thickness on the surface of the workpiece anode, and patterning the dielectric cavity layer to form a cavity; S3: preparing a tool cathode having a suspended metal film and a liquid injection channel; S4: aligning and sealing the tool cathode with the dielectric cavity layer to form a sealed cavity; S5: sequentially injecting a phospholipid-containing organic solvent and an electrolyte containing ion channel proteins into the cavity through the liquid injection channel, so that the electrolyte contacts the workpiece anode interface through the channels of the ion channel proteins; S6: connecting the workpiece anode and the tool cathode to the positive and negative electrodes of a pulse power supply, respectively, for electrochemical machining.

[0009] The thickness of the aforementioned dielectric cavity layer is adjustable in the nanometer to sub-nanometer scale range.

[0010] The material of the dielectric cavity layer mentioned above is silicon nitride, aluminum oxide, or other insulating materials.

[0011] The metal film on the anode of the above-mentioned workpiece is prepared by magnetron sputtering, atomic layer deposition or epitaxial deposition.

[0012] The material of the suspended metal film of the cathode of the above-mentioned tool is gold, platinum or other inert metals that are difficult to polarize in the electrochemical process.

[0013] The total thickness of the suspended metal film of the workpiece anode, dielectric cavity layer and tool cathode is configured to not exceed 50 nm.

[0014] In step S5, after injecting the phospholipid-containing organic solvent, let it stand for 5-10 minutes to allow the phospholipid to self-assemble on the anode surface of the workpiece; when injecting the electrolyte, first inject a small amount of electrolyte into the organic solvent, and after the interface self-assembly is completed, continue to inject until the electrolyte simultaneously contacts the anode of the workpiece and the cathode of the tool.

[0015] The electrical signal parameters output by the aforementioned pulse power supply are adjusted to match the thickness of the dielectric cavity layer.

[0016] The above-described apparatus or method is used in the in-situ observation of electrochemical atomic-scale processing using transmission electron microscopy.

[0017] Beneficial effects: (1) Precise control of processing gap and cathode thickness. The use of chemical or physical deposition methods can strictly control the thickness and deposition uniformity of the dielectric cavity layer and the tool cathode, thereby effectively reducing the inter-electrode gap to the nanometer-sub-nanometer level. The beneficial effect is to reduce the adverse attenuation of the double-layer voltage, while ensuring that the current density at each position of the anode interface of the workpiece is consistent, which is conducive to the uniform electrochemical dissolution. Furthermore, the dielectric cavity sidewalls formed by this method to store organic solvents and electrolytes have atomically smooth characteristics, which is conducive to the weak disturbance of the solution in the dielectric cavity and avoids the mass transfer difficulties caused by the formation of solid-liquid-gas three-phase interface.

[0018] (2) The ultrathin electrolyte layer facilitates the analysis of reaction mechanisms. The ultrathin electrolyte layer encapsulated in the dielectric cavity facilitates the online observation of the phospholipid monomer self-assembly process and the electrochemical removal mechanism and charge accumulation at the anode interface of charged workpieces in in-situ liquid phase TEM. Unlike the traditional single-layer graphene encapsulation liquid phase sample cell, which only seals the ultrathin liquid layer, this scheme combines the dielectric cavity and tool cathode encapsulation method to simultaneously perform a series of important functions such as adjusting the processing gap, in-situ observation, electrochemical reaction and electrolyte storage.

[0019] (3) Improve the signal-to-noise ratio of current signal detection. In near-atomic scale electrochemical machining, the electrochemical dissolution current signal is weak (its interference noise mainly comes from bubble interference, double layer fluctuations and material intrinsic properties). The noise signal is likely to drown out the characteristic current signal used for electrochemical dissolution. However, in this invention, the workpiece anode and the tool cathode are separated by a low-noise dielectric material, which also reduces current noise and is conducive to improving the signal-to-noise ratio of current signal detection.

[0020] (4) Visualization platform for the spread area of ​​phospholipid bilayer mask. Thanks to the ultra-thin characteristics of the entire encapsulation system (with light-transmitting characteristics), the method of the present invention can be well integrated with an inverted fluorescence microscope to detect the spread state of the biophospholipid bilayer between the ultra-thin electrolyte layer and the workpiece anode in real time, ensuring the integrity of the mask. Attached Figure Description

[0021] Figure 1 This is a schematic diagram of the formation of a dielectric cavity.

[0022] Figure 2 This is a flowchart of the anode fabrication process for atomically flat workpieces.

[0023] Figure 3 This is a schematic diagram of the tool cathode structure.

[0024] Figure 4 This is a schematic diagram of the self-assembly of organic solvent-electrolyte droplets at the interface within a dielectric cavity.

[0025] Figure 5 This is a schematic diagram of the formation of electrolyte channels in electrolytic machining.

[0026] Figure 6 This is a high-resolution TEM image of the workpiece anode (2).

[0027] Figure 7 This is a high-resolution TEM image of the tool cathode (5).

[0028] Figure 8 These are electron microscope images of uniformly processed nickel nanopores;

[0029] Figure 9 This is a comparison of the current noise under dielectric cavity conditions and the shielding effect tested in a traditional open environment.

[0030] Figure 10 These are TEM images of phospholipid monomers self-assembling into a phospholipid bilayer and subsequent MspA ion channel protein embedding, collected in situ. Detailed Implementation

[0031] An apparatus for near-atomic-scale electrochemical machining of ion channel proteins includes: a silicon nitride thin film window 1; a workpiece anode 2, which is an ultrathin metal film deposited on the silicon nitride thin film window 1; a dielectric cavity layer 3 formed on the surface of the workpiece anode 2, the thickness of the dielectric cavity layer 3 being precisely controlled to define the machining gap and patterned to form a cavity 7 for containing liquid; and a tool cathode 4 including a support substrate 5 and a suspended metal film formed thereon, the tool cathode 4 having a liquid injection channel 6 communicating with the cavity 7; wherein the suspended metal film side of the tool cathode 4 is aligned and sealed to the dielectric cavity layer 3, such that the workpiece anode 2, the sidewall of the dielectric cavity layer 3, and the suspended metal film of the tool cathode 4 together form the sealed cavity 7.

[0032] A method for gap control in near-atomic-scale electrochemical machining of ion channel proteins using the aforementioned device includes the following steps: S1: providing a silicon nitride thin film window 1 on which a workpiece anode 2 is deposited; S2: forming a dielectric cavity layer 3 of controllable thickness on the surface of the workpiece anode 2, and patterning the dielectric cavity layer 3 to form a cavity 7; S3: preparing a tool cathode 4 having a suspended metal film and a liquid injection channel 6; S4: aligning and sealing the tool cathode 4 with the dielectric cavity layer 3 to form a sealed cavity 7; S5: sequentially injecting a phospholipid-containing organic solvent 10 and an electrolyte 8 containing ion channel proteins 11 into the cavity 7 through the liquid injection channel 6, so that the electrolyte 8 contacts the interface of the workpiece anode 2 through the channel of the ion channel proteins 11; S6: connecting the workpiece anode 2 and the tool cathode 4 to the positive and negative electrodes of a pulse power supply 12, respectively, for electrochemical machining.

[0033] The thickness of dielectric cavity layer 3 is adjustable in the nanometer to sub-nanometer scale range.

[0034] The dielectric cavity layer 3 is made of silicon nitride, aluminum oxide, or other insulating materials.

[0035] The metal film on the workpiece anode 2 is prepared by magnetron sputtering, atomic layer deposition or epitaxial deposition.

[0036] The material of the suspended metal film of the tool cathode 4 is gold, platinum or other inert metals that are difficult to polarize during electrochemical processes.

[0037] The total thickness of the suspended metal film of the workpiece anode 2, dielectric cavity layer 3 and tool cathode 4 is configured to not exceed 50 nm.

[0038] In step S5, after injecting the phospholipid-containing organic solvent 10, let it stand for 5-10 minutes to allow the phospholipid to self-assemble on the surface of the workpiece anode 2; when injecting the electrolyte 8, first inject a small amount of electrolyte 8 into the organic solvent 10, and after the interface self-assembly is completed, continue to inject until the electrolyte 8 simultaneously contacts the workpiece anode 2 and the tool cathode 4.

[0039] The electrical signal parameters output by the pulse power supply 12 are adapted and adjusted according to the thickness of the dielectric cavity layer 3.

[0040] The above-mentioned apparatus or method is used in the in-situ observation of electrochemical atomic-scale processing using transmission electron microscopy.

[0041] Example 1

[0042] refer to Figures 1 to 7 The main steps of the near-atomic-scale electrolytic machining process for achieving 1nm nickel metal holes using this invention are as follows:

[0043] Step 1: A TEM silicon nitride thin film window 1 with an outer frame diameter of 3mm, a thickness of 200μm, and a window size of 50×50μm (e.g., the silicon nitride thickness is 10nm, the thickness needs to be adjusted according to the window size) is used as the substrate for fabricating the workpiece anode 2. The required nickel atoms (e.g., nickel atoms) are sputtered onto the surface of the silicon nitride thin film using a magnetron sputtering process. Figure 6 As shown in the figure, its thickness can be flexibly adjusted according to the deposition time (e.g., 5nm);

[0044] Step 2: After surface cleaning, degreasing, nitrogen drying and plasma cleaning, the TEM silicon nitride thin film window 1 of the nickel-plated workpiece anode 2 is immediately subjected to low-pressure chemical vapor deposition. The deposition thickness of the silicon nitride dielectric cavity layer 3 is controlled according to the deposition time. The thickness of the silicon nitride dielectric cavity layer 3 is the processing gap during electrolytic processing. Further, excess silicon nitride is removed by photolithography and reactive ion etching processes to expose the cavity 7, which serves as a storage space for the phospholipid-containing organic solvent 10 and the electrolyte 8.

[0045] Step 3: Similarly, deposit an inert, non-polarizable metal (e.g., gold) of the required thickness as a tool cathode 4 on the supporting substrate 5 (silicon wafer), such as... Figure 7 As shown, based on the experimental environment requirements of in-situ transmission electron microscopy, the thicknesses of the workpiece anode 2, tool cathode 4, and dielectric cavity layer 3 are reasonably allocated. It is expected that a total thickness of less than 50 nm can achieve better in-situ online observation of the atomic-scale electrochemical dissolution process. The prepared tool cathode 4 and dielectric cavity layer 3 are sealed to adapt to the high vacuum environment of TEM.

[0046] Step 4: Prepare 5 mg / mL organic solvent 10 containing phospholipids (DPhPC phospholipid monomer organic solvent dissolved in hexadecane). Inject the organic solvent into the cavity 7 through the liquid injection channel 6 on the tool cathode 4 using a micropipette. The height should not exceed 1 / 3 of the cavity 7. Wait 5-10 minutes to allow the phospholipid monomers to fully self-assemble at the interface 9 between the workpiece anode 2 and the organic solvent.

[0047] Step 5: Take another 0.5 mol / L NaCl electrolyte 8 containing MspA ion channel protein 11 into a micropipette. First, inject 20 nL of electrolyte 8 into organic solvent 10. Phospholipid monomers self-assemble at the interface between organic solvent 10 and electrolyte 8. Then, add electrolyte 8 until electrolyte 8 contacts the workpiece anode 2 and the tool cathode 4. A phospholipid bilayer 9-1 mask is formed at the organic solvent interface 9 between workpiece anode 2 and electrolyte 8. MspA ion channel protein 11 dissolved in electrolyte 8 self-assembles in phospholipid bilayer 9-1 through hydrophobic interactions. Electrolyte 8 reaches the workpiece anode 2 interface through the internal ion channels of MspA ion channel protein 11.

[0048] Step 6: Using workpiece anode 2 as the positive electrode and tool cathode 4 as the negative electrode, connect ultra-short pulse power supply 12 and apply a pulsed electric field (parameters: pulse width 200 ps, ​​interval 500 ps, ​​amplitude 50 mV). Under the influence of the electric field, metallic nickel undergoes an electrochemical dissolution reaction. After the predetermined time is reached, the processing ends. The processing effect is as follows: Figure 8 As shown.

[0049] Example 2

[0050] This embodiment aims to verify the advantages of the device of the present invention in reducing current noise. It uses the experimental parameters of Example 1 (e.g., pulse width 200 ps, ​​interval 500 ps, ​​amplitude 50 mV) and, under a voltage of 100 mV, collects current noise in both a conventional electrolytic cell and the dielectric cavity of the present invention. Figure 9 As shown, the noise range within the dielectric cavity of this invention is 15±1 pA, while that of the conventional approach is 7.5±6 pA. This indicates that the noise fluctuation range of this invention is significantly lower because the dielectric cavity layer (such as silicon nitride) acts as an insulating layer, effectively shielding external interference (such as bubble disturbances or double-layer fluctuations), thereby improving the signal-to-noise ratio. This low-noise environment is crucial for detecting weak current signals from near-atomic-scale electrochemical dissolution, preventing signal overload and ensuring the stability and observability of the processing.

[0051] Example 3

[0052] This embodiment aims to specifically demonstrate and verify the unique ability of the device of the present invention to achieve in-situ observation of the dynamic process of phospholipid self-assembly under transmission electron microscopy (TEM), which is a key prerequisite for elucidating the electrochemical processing mechanism at the near-atomic scale. The specific implementation process is as follows: Using the same device construction method as in Example 1, an observation unit with a total thickness of no more than 50 nm was prepared. Through the liquid injection channel on the tool cathode, a hexadecane organic solvent containing DPhPC phospholipid monomers was first injected into the dielectric cavity, and allowed to stand for 5-10 minutes to allow it to initially adsorb at the interface on the surface of the workpiece anode (such as a nickel film). Subsequently, an electrolyte containing MspA ion channel protein (such as 0.5 mol / L NaCl) was slowly injected using a microsyringe. At the interface between the two liquids, phospholipid molecules rapidly self-assemble. Taking advantage of the fact that the ultrathin structure of the device of the present invention has minimal impact on electron beam penetration, this dynamic process was successfully captured in-situ under TEM: as follows Figure 10 As shown, it can be clearly observed that phospholipid monomers spontaneously organize into spherical phospholipid vesicles. Subsequently, these vesicles adsorb onto the anode surface of the workpiece, rupture, and eventually spread out to form a continuous and stable phospholipid bilayer confined mask. MspA ion channel proteins embedded in the mask can also be observed.

Claims

1. An apparatus for near-atomic-scale electrochemical processing of ion channel proteins, characterized in that, include: A silicon nitride thin film window (1); a workpiece anode (2), which is a metal film deposited on the silicon nitride thin film window (1); a dielectric cavity layer (3), which is disposed on the surface of the workpiece anode (2), the thickness of the dielectric cavity layer (3) is precisely controlled to limit the processing gap, and is patterned to form a cavity (7) for containing liquid; a tool cathode (4), which includes a support substrate (5) and a suspended metal film formed thereon, and the tool cathode (4) is provided with a liquid injection channel (6) communicating with the cavity (7); wherein, the suspended metal film side of the tool cathode (4) is aligned and sealed with the dielectric cavity layer (3), so that the workpiece anode (2), the sidewall of the dielectric cavity layer (3) and the suspended metal film of the tool cathode (4) together form the sealed cavity (7).

2. A method for gap modulation in near-atomic-scale electrochemical processing of ion channel proteins using the device described in claim 1, characterized in that, Includes the following steps: S1: Provide a silicon nitride thin film window (1) on which the workpiece anode (2) is deposited; S2: Form a dielectric cavity layer (3) with controllable thickness on the surface of the workpiece anode (2), and pattern the dielectric cavity layer (3) to form a cavity (7); S3: Prepare a tool cathode (4) with a suspended metal film and a liquid injection channel (6); S4: Align the tool cathode (4) with the dielectric cavity layer (3) and seal and assemble them to form a sealed cavity (7); S5: Inject a phospholipid-containing organic solvent (10) and an electrolyte (8) containing ion channel protein (11) into the cavity (7) through the liquid injection channel (6), so that the electrolyte (8) contacts the interface of the workpiece anode (2) through the channel of the ion channel protein (11); S6: Connect the workpiece anode (2) and the tool cathode (4) to the positive and negative electrodes of the pulse power supply (12) respectively, and perform electrochemical processing.

3. The method according to claim 2, characterized in that, The thickness of the dielectric cavity layer (3) is adjustable in the nanometer to sub-nanometer scale range.

4. The method according to claim 2 or 3, characterized in that, The dielectric cavity layer (3) is made of silicon nitride or aluminum oxide.

5. The method according to claim 2, characterized in that, The metal film of the workpiece anode (2) is prepared by magnetron sputtering, atomic layer deposition or epitaxial deposition.

6. The method according to claim 2, characterized in that, The material of the suspended metal film of the tool cathode (4) is gold, platinum or other inert metals that are difficult to polarize in the electrochemical process.

7. The method according to claim 2, characterized in that, The total thickness of the suspended metal film of the workpiece anode (2), dielectric cavity layer (3) and tool cathode (4) is configured to not exceed 50 nm.

8. The method according to claim 2, characterized in that, In step S5, after injecting the phospholipid-containing organic solvent (10), let it stand for 5-10 minutes to allow the phospholipid to self-assemble on the surface of the workpiece anode (2); when injecting the electrolyte (8), first inject a portion of the electrolyte (8) into the organic solvent (10), and after the interface self-assembly is completed, continue to inject until the electrolyte (8) simultaneously contacts the workpiece anode (2) and the tool cathode (4).

9. The method according to claim 2, characterized in that, The electrical signal parameters output by the pulse power supply (12) are adapted and adjusted according to the thickness of the dielectric cavity layer (3).

10. The application of the apparatus of claim 1 or the method of claim 2 in in-situ observation of electrochemical atomic-scale processing using transmission electron microscopy.

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