Cd-EDTA detection method based on solid-state nanopore detection technology

Abstract

The present invention discloses a Cd-EDTA detection method based on solid-state nanopore detection technology, comprising: obtaining a nanopore sensor, wherein the nanopores in the nanopore sensor are prepared using electron beam lithography technology; configuring an electrolyte solution; preparing a Cd-EDTA sample and configuring a standard Cd-EDTA solution of different concentrations; after configuring an electrochemical workstation, applying an AC signal, and obtaining a response signal of the nanopore sensor within a preset frequency range to obtain a current blockade signal; and detecting Cu-EDTA based on the current blockade signal corresponding to the Cd-EDTA sample and the standard Cd-EDTA solution of different concentrations. The present invention improves the detection sensitivity of the Cd-EDTA complex and can achieve accurate detection at low concentrations. Real-time, label-free detection is achieved, which is suitable for pollutant monitoring in complex environments in mining areas.

Description

Technical Field

[0001] The present invention belongs to the technical field of rapid detection of environmental pollutants in mining areas, and in particular relates to a Cd-EDTA detection method based on solid-state nanopore detection technology. Background Art

[0002] With the increase in mining activities, heavy metal pollution in the environment, especially pollution from rare metals such as indium (Cd), has gradually become a hot issue in environmental monitoring. Indium is widely used in electronics, photovoltaics, and displays. The complex it forms with ethylenediaminetetraacetic acid (EDTA) (Cd-EDTA) is relatively common in the environment and has strong mobility and bioaccumulation. Once Cd-EDTA enters water bodies, it not only causes water pollution but also poses a long-term threat to ecosystems and human health. Therefore, accurate and real-time detection of Cd-EDTA complexes has become a key task in environmental pollution monitoring in mining areas.

[0003] While traditional detection methods such as spectroscopy, electrochemistry, and chromatography can effectively detect Cd-EDTA in laboratory settings, they often struggle to provide rapid and accurate monitoring results in the complex environment of mining areas, particularly where water contains a wide variety of pollutants and strong background interference. Furthermore, cumbersome procedures and lengthy processing times limit their application in the field. Summary of the Invention

[0004] In order to solve the above technical problems, the present invention proposes a Cd-EDTA detection method based on solid-state nanopore detection technology to solve the problems existing in the above-mentioned prior art.

[0005] To achieve the above objectives, the present invention provides a Cd-EDTA detection method based on solid-state nanopore detection technology, comprising:

[0006] Obtaining a nanopore sensor, wherein the nanopore in the nanopore sensor is fabricated using electron beam lithography;

[0007] Prepare electrolyte solution; prepare Cd-EDTA samples and prepare standard Cd-EDTA solutions of different concentrations;

[0008] After configuring the electrochemical workstation, an AC signal was applied, and the response signal of the nanopore sensor was acquired within a preset frequency range to obtain a current blockade signal. Cu-EDTA was detected based on the current blockade signals corresponding to the Cd-EDTA sample and standard Cd-EDTA solutions of different concentrations.

[0009] Optionally, the process of fabricating the nanopore using electron beam lithography includes:

[0010] After cleaning the Si3N4 substrate, a layer of photoresist is coated on the surface, where the thickness of the photoresist is determined based on the aperture requirements of the nanopore. After the photoresist is coated, spin coating technology is used to adjust the uniformity of the photoresist coating. An electron beam lithography machine scans the photoresist according to a preset pattern for exposure. After exposure, a development process is performed to remove the unexposed photoresist portion. The pattern is transferred to the Si3N4 substrate using plasma etching technology, and the material in the exposed area is removed by etching to complete the nanopore preparation.

[0011] Optionally, the process of obtaining the nanopore sensor further includes:

[0012] Graphene was grown on the surface of a metal catalyst using chemical vapor deposition. The graphene surface was functionalized using oxygen plasma treatment and chemical activation, and then transferred to a Si3N4 substrate via a solution transfer method.

[0013] Optionally, the electrolyte solution is a mixture of ammonia water and methanol, and the pH range of the electrolyte solution is 9.5-11.5.

[0014] Optionally, the process for preparing a Cd-EDTA sample includes:

[0015] The required mass of the Cd-EDTA complex is calculated according to the required concentration and weighed; it is completely dissolved in deionized water and stirred or ultrasonically treated; the Cd-EDTA solution is added to the tailings and stirred evenly until the complex is fully adsorbed on the tailings surface, and finally a Cd-EDTA tailings sample with the required concentration is obtained.

[0016] Optionally, the standard Cd-EDTA solution includes four concentrations: 1 mol / L, 2 mol / L, 4 mol / L, and 6 mol / L.

[0017] Optionally, the process of obtaining the current blockade signal of the standard Cd-EDTA solution includes: dropping standard Cd-EDTA solutions of different concentrations onto the surface of the nanopore sensor and maintaining a contact time of 30 minutes.

[0018] Compared with the prior art, the present invention has the following advantages and technical effects:

[0019] The present invention provides a novel device for detecting complex pollutants (Cd-EDTA) in mining areas. The device is based on a nanopore sensor and introduces a two-dimensional material graphene layer to enhance the conductivity and selectivity, thereby improving the detection sensitivity of the Cd-EDTA complex. Nanopores are prepared using electron beam lithography technology, and graphene is grown on the surface of a metal catalyst by chemical vapor deposition (CVD) and transferred to a Si3N4 substrate. The graphene surface is functionalized and covalently bound to an antibody. The impedance spectrum is measured by an electrochemical workstation. After applying a small-amplitude AC signal, the change in charge transfer resistance in the Nyquist plot is analyzed to further evaluate the response characteristics of the sensor to the Cd-EDTA complex. Through gradient experiments of Cd-EDTA solutions of different concentrations, combined with Nyquist plot analysis, the sensitivity and detection performance of the sensor can be accurately evaluated, providing effective technical support for the monitoring of heavy metal pollutants in mining environments. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] The accompanying drawings, which constitute part of this application, are intended to provide a further understanding of this application. The exemplary embodiments and descriptions of this application are intended to explain this application and do not constitute an improper limitation on this application. In the accompanying drawings:

[0021] Figure 1 is a schematic diagram of a device according to an embodiment of the present invention;

[0022] Figure 2 Schematic diagram of the sample measurement principle of an embodiment of the present invention, (a) is a schematic diagram of the nanopore detection device; (b) is a schematic diagram of the reference current; (c) and (d) are schematic diagrams of the blockade current generated by analytes of different sizes passing through the nanopore;

[0023] Figure 3 Flowchart of a method according to an embodiment of the present invention. DETAILED DESCRIPTION

[0024] It should be noted that, in the absence of conflict, the embodiments and features of the embodiments in this application can be combined with each other. The present application will be described in detail below with reference to the accompanying drawings and in combination with the embodiments.

[0025] It should be noted that the steps shown in the flowcharts of the accompanying drawings can be executed in a computer system such as a set of computer-executable instructions, and that, although a logical order is shown in the flowcharts, in some cases, the steps shown or described can be executed in an order different from that shown here.

[0026] Example 1

[0027] like Figure 1-3 As shown, this embodiment provides a Cd-EDTA detection method based on solid-state nanopore detection technology, including:

[0028] Obtaining a nanopore sensor, wherein the nanopore in the nanopore sensor is fabricated using electron beam lithography;

[0029] Furthermore, the process of preparing nanopores using electron beam lithography includes:

[0030] After cleaning the Si3N4 substrate, a layer of photoresist is coated on the surface, where the thickness of the photoresist is determined based on the aperture requirements of the nanopore. After the photoresist is coated, spin coating technology is used to adjust the uniformity of the photoresist coating. An electron beam lithography machine scans the photoresist according to a preset pattern for exposure. After exposure, a development process is performed to remove the unexposed photoresist portion. The pattern is transferred to the Si3N4 substrate using plasma etching technology, and the material in the exposed area is removed by etching to complete the nanopore preparation.

[0031] Furthermore, the process of obtaining the nanopore sensor also includes:

[0032] Graphene was grown on the surface of a metal catalyst using chemical vapor deposition. The graphene surface was functionalized using oxygen plasma treatment and chemical activation, and then transferred to a Si3N4 substrate via a solution transfer method.

[0033] Specifically, a two-dimensional graphene layer was added to the traditional nanopore sensor to enhance the conductivity and selectivity of the nanopore and improve the detection sensitivity of the Cd-EDTA complex. The nanopore was manufactured using electron beam lithography to precisely control the pore size (typically 10-15nm), and the pore pattern was transferred to the Si3N4 substrate by plasma etching. The graphene layer was grown on the surface of the metal catalyst by chemical vapor deposition (CVD) and transferred to the Si3N4 substrate surface by solution transfer.

[0034] To improve selectivity, the graphene surface is treated with oxygen plasma to generate oxygen-containing functional groups such as carboxyl and hydroxyl groups. The carboxyl groups are further activated using reagents such as 1-chloro-3-chloromethyl-1,3,5-triazine (CDI), or converted into active esters through EDC / NHS cross-linking reaction, which are then covalently bound to amino groups (such as lysine residues) on the antibody to achieve antibody immobilization.

[0035] Prepare electrolyte solution; prepare Cd-EDTA samples and prepare standard Cd-EDTA solutions of different concentrations;

[0036] Furthermore, the electrolyte solution is a mixture of ammonia water and methanol, and the pH range of the electrolyte solution is 9.5-11.5.

[0037] Furthermore, the process of preparing the Cd-EDTA sample includes:

[0038] The required mass of the Cd-EDTA complex is calculated according to the required concentration and weighed; it is completely dissolved in deionized water and stirred or ultrasonically treated; the Cd-EDTA solution is added to the tailings and stirred evenly until the complex is fully adsorbed on the tailings surface, and finally a Cd-EDTA tailings sample with the required concentration is obtained.

[0039] Furthermore, the standard Cd-EDTA solution includes four concentrations: 1 mol / L, 2 mol / L, 4 mol / L, and 6 mol / L.

[0040] Specifically, precisely prepare a Cd-EDTA complex solution based on the desired concentration (1 mol / L, 2 mol / L, 4 mol / L, or 6 mol / L). Weigh the desired amount of Cd-EDTA complex and dissolve it in an appropriate amount of deionized water, ensuring complete dissolution. Stir or ultrasonicate the solution to ensure uniformity. Finally, add the dissolved Cd-EDTA solution to the tailings, ensuring the complex is fully adsorbed on the tailings surface, to obtain a Cd-EDTA tailings sample of the desired concentration.

[0041] Exemplarily, a Cd-EDTA sample is prepared. First, the mass of the required Cd-EDTA complex is calculated based on the required concentration. Next, the required mass of Cd-EDTA complex is weighed. You can choose to use a Cd-EDTA standard solution or synthesize the Cd-EDTA complex from Cd(II) and EDTA powder. Then, it is dissolved in an appropriate amount of deionized water, ensuring that the Cd-EDTA is completely dissolved, and ensure uniformity by stirring or ultrasonic treatment. Finally, the dissolved Cd-EDTA solution is added to the tailings and stirred evenly to ensure that the complex is fully adsorbed on the tailings surface, and finally a Cd-EDTA tailings sample of the required concentration is obtained.

[0042] After configuring the electrochemical workstation, an AC signal is applied to both sides of the nanopore sensor, and the response signal of the nanopore sensor is obtained within a preset frequency range to obtain a current blockade signal. Cu-EDTA is detected based on the current blockade signals corresponding to the Cd-EDTA sample and standard Cd-EDTA solutions of different concentrations.

[0043] Furthermore, the process of obtaining the current blockade signal of the standard Cd-EDTA solution includes: dropping the standard Cd-EDTA solutions of different concentrations onto the surface of the nanopore sensor and maintaining a contact time of 30 minutes.

[0044] Specifically, the prepared Cd-EDTA solution was added dropwise to the surface of the nanopore sensor for a 30-minute contact reaction. Using an electrochemical workstation, a working electrode, reference electrode, and auxiliary electrode were set up, and a small-amplitude AC signal (typically 10mV or 50mV) was applied. The sensor's response to the signal was measured within a frequency range of 10Hz to 100kHz. The impedance data recorded by the electrochemical workstation was used to analyze the change in charge transfer resistance (Rct) in the impedance spectrum using a Nyquist plot to evaluate the relationship between the Cd-EDTA complex concentration and the sensor response.

[0045] Electrochemical workstations such as Figure 1 As shown, Figure 1 (d) is a Faraday shielding box, and the nanopore sensor is fixed on Figure 1 (e) The nanopore on the nanopore sensor becomes the only channel in the fluid pool, and the fluid pool is divided into two independent fluid pools by the nanopore sensor; Figure 1 (b) The power supply is used to apply electrical signals to both sides of the nanopore sensor;

[0046] Figure 1 (a) shows the data processing module, which is used to Figure 1 Middle (c) The patch clamp amplifier collects the electrical signal of the nanopore sensor and determines the baseline current and time current diagram.

[0047] The impedance data was analyzed using Nyquist plots. The relationship between the real (Z') and imaginary (Z") parts of the impedance was plotted to observe how the charge transfer resistance (Rct) changes with Cd-EDTA concentration. By comparing Nyquist plots at different concentrations, the diameter of the semicircle arc was quantified or the impedance data was fitted, and the detection performance and sensitivity of the sensor were further analyzed.

[0048] Concentration gradient experiments and performance evaluation

[0049] Four concentration gradient experiments were conducted by replacing the Cd-EDTA solution in the flow cell. Cd-EDTA solutions with concentrations of 1 mol / L, 2 mol / L, 4 mol / L, and 6 mol / L were added to the positive electrode side of the flow cell, while 0.5 mol / L Cd-EDTA solution or electrolyte solution was added to the negative electrode side. The Nyquist plots of the different Cd-EDTA concentrations were compared to further evaluate the sensor's detection sensitivity and performance.

[0050] This invention improves the detection sensitivity of Cd-EDTA complexes, enabling accurate detection at low concentrations. It achieves real-time, label-free detection, making it suitable for pollutant monitoring in complex mining environments. Through impedance spectroscopy and Nyquist plot analysis, it provides highly accurate, quantitative detection results with good stability and repeatability. The introduction and functionalization of graphene improves the sensor's selectivity and enhances its ability to capture target pollutants.

[0051] The above are merely preferred embodiments of the present application, but the scope of protection of the present application is not limited thereto. Any changes or substitutions that can be easily conceived by a person skilled in the art within the technical scope disclosed in this application should be included in the scope of protection of the present application. Therefore, the scope of protection of the present application should be based on the scope of protection of the claims.

Claims

1. A Cd-EDTA detection method based on solid-state nanopore detection technology, characterized in that: The following steps are involved: Obtaining a nanopore sensor, wherein the nanopore in the nanopore sensor is fabricated using electron beam lithography; Prepare electrolyte solution; prepare Cd-EDTA samples and prepare standard Cd-EDTA solutions of different concentrations; After configuring the electrochemical workstation, an AC signal is applied and the response signal of the nanopore sensor is acquired within a preset frequency range to obtain a current blockade signal. Cd-EDTA is detected based on the current blockade signals corresponding to the Cd-EDTA sample and standard Cd-EDTA solutions of different concentrations. The process of fabricating nanopores using electron beam lithography involves: After cleaning the Si3N4 substrate, a layer of photoresist is coated on the surface, wherein the thickness of the photoresist is determined based on the required aperture of the nanopore; after the photoresist is coated, a spin coating technique is used to adjust the uniformity of the photoresist coating; an electron beam lithography machine scans the photoresist according to a preset pattern for exposure; after exposure, a development process is performed to remove the unexposed photoresist portion; the pattern is transferred to the Si3N4 substrate using a plasma etching technique, and the material in the exposed area is removed by etching to complete the nanopore preparation; The electrolyte solution is a mixture of ammonia water and methanol, and the pH range of the electrolyte solution is 9.5-11.5; The process of preparing Cd-EDTA samples includes: The required mass of the Cd-EDTA complex is calculated according to the required concentration and weighed; it is completely dissolved in deionized water and stirred or ultrasonically treated; the Cd-EDTA solution is added to the tailings and stirred evenly until the complex is fully adsorbed on the tailings surface, and finally a Cd-EDTA tailings sample with the required concentration is obtained.

2. The Cd-EDTA detection method based on solid-state nanopore detection technology according to claim 1, characterized in that: The standard Cd-EDTA solution includes four concentrations: 1 mol / L, 2 mol / L, 4 mol / L, and 6 mol / L.

3. The Cd-EDTA detection method based on solid-state nanopore detection technology according to claim 1, characterized in that: The process of obtaining the current blockade signal of the standard Cd-EDTA solution includes: dropping the standard Cd-EDTA solutions of different concentrations onto the surface of the nanopore sensor and maintaining a contact time of 30 minutes.

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