Preparation method and application of a glucose sensor based on acupuncture needles

Abstract

The present invention discloses a preparation method and application of a glucose sensor based on acupuncture needles. The sensor is prepared based on acupuncture needles and a specific structural design and molecular recognition mechanism. The sensor can detect glucose concentration with high specificity and high sensitivity through the change of drain current, and has an ultra-wide dynamic detection range (0-40mmol·L ‑1 ), covering the clinical range from hypoglycemia to severe hyperglycemia, and with a detection limit as low as 54µmol·L ‑1 The present invention has excellent gate control capabilities. During the detection process, the device exhibits a significant switching ratio and fast response characteristics, meeting the biosensor's requirements for sensitivity and real-time performance; and has a fast response capability to gate voltage changes, indicating that it has good stability and repeatability under repeated operations, which can provide direction and basis for the development of blood glucose monitoring technology with continuous, real-time and precise characteristics.

Description

Technical Field

[0001] The present invention relates to the technical field of chemical sensors, and in particular to a preparation method and application of an acupuncture needle-based glucose sensor. Background Art

[0002] For diabetic patients, blood glucose monitoring is a core part of disease management, maintaining blood glucose stability, and preventing complications. Although the traditional fingertip blood glucose testing method has long been used in clinical practice, it has many problems that are difficult to ignore. For example, this detection method is susceptible to interference from various external factors, such as changes in ambient temperature and humidity. These factors may cause deviations in test results and fail to accurately reflect the patient's true blood glucose level; and continuous, real-time detection is impossible. Therefore, it is very meaningful to develop a blood glucose monitoring technology with continuous, real-time, and accurate characteristics.

[0003] At the same time, acupuncture needles, as key instruments that directly act on acupuncture points in acupuncture treatment, have received widespread attention for their function expansion research. The organic combination of advanced sensor technology and acupuncture needles to construct an acupuncture needle sensor system with integrated detection function not only provides a new perspective for in-depth research on the mechanism of action of acupuncture, but is also expected to explain the scientific mechanism of traditional acupuncture therapy from a molecular level. It also opens up a technical path with important innovative value in the field of biomedical testing. For example, a study targeted implantable needle-type microsensors and used them for real-time monitoring of acupuncture-mediated adenosine release in vivo. The sensor was modified with Prussian blue nanoparticles, multi-enzyme systems and poly-o-phenylenediamine membranes, and the linear range for in vitro detection of adenosine was 0-50μM. Another study developed an AN / AuPs / G biosensor for the detection of histamine based on bare acupuncture needles through methods such as electrodeposition, brushing and annealing. The in vitro detection limit of this sensor was approximately 4.352 (±3.419)×10 -12 mol·L -1 , the sensitivity is about 6.296 (± 3.873) μA·μM -1 Real-time monitoring in rats revealed minimal changes in histamine concentrations at non-acupoints, while histamine concentrations at acupoint PC6 initially increased and then decreased, peaking at approximately 18 minutes. However, there are currently no studies on acupuncture needle-based sensors for glucose detection. Summary of the Invention

[0004] Therefore, based on the above background, the present invention provides a preparation method and application of a glucose sensor based on acupuncture needles. The present invention uses acupuncture needles as a substrate and fixes PEDOT:PSS hydrogel on the cross section of a stainless steel acupuncture needle to form a conductive channel connecting the gold source and the copper drain, presenting a "sandwich" structure. It can achieve high specificity, stability and high sensitivity detection of glucose, so as to provide direction and basis for the development of continuous, real-time and accurate blood glucose monitoring technology in clinical practice.

[0005] The technical solution provided by the present invention is:

[0006] A method for preparing a glucose sensor based on acupuncture needles comprises the following steps:

[0007] S1: Substrate pretreatment

[0008] Stainless steel acupuncture needles AN were used as substrates, which were roughened and electrochemically deposited with AuNPs.

[0009] S2: Multi-layer structure construction and insulation treatment

[0010] S2.1 Coat the stainless steel acupuncture needle AN pretreated in step S1 with epoxy resin, and after drying, form an insulating layer;

[0011] S2.2 sputter-deposit a copper drain electrode onto the stainless steel acupuncture needle AN with an insulating layer prepared in step S2.1;

[0012] S2.3 The stainless steel acupuncture needle AN prepared in step S2.2 is coated with epoxy resin again. After curing, the needle tip is polished until the Au / Cu bimetallic layer is exposed on the cross section;

[0013] S3: Functional layer integration and gate modification

[0014] S3.1 Constructing a conductive channel

[0015] Fix the PEDOT:PSS hydrogel on the cross section of the stainless steel acupuncture needle AN prepared in step S2.3 to form a conductive channel connecting the gold source electrode and the copper drain electrode;

[0016] S3.2 Gate functionalization modification

[0017] A silver wire was selected as the gate and immersed in a solution containing EDOT-FPBA. A poly(EDOT-FPBA) molecular recognition layer was formed on the gate surface using a constant potential electropolymerization method, thus completing the sensor preparation.

[0018] Furthermore, the operation of electrochemically depositing AuNPs in step S1 is as follows:

[0019] The roughened stainless steel acupuncture needle AN was placed in a tetrachloroauric acid solution, and a reference electrode and a counter electrode were inserted to construct a three-electrode system. The system was connected to an electrochemical workstation, and a CV method was used for electropolymerization cyclic deposition to form an AuNPs layer, which was then cleaned after the reaction was completed.

[0020] Furthermore, the operation of CV electropolymerization is as follows: the CV electropolymerization is carried out in the range of -1.5V to 0.5V at 100mV·s -1 The scan rate was 20 cycles of deposition.

[0021] Furthermore, the preparation method of the PEDOT:PSS hydrogel used in step S3.1 includes the following steps:

[0022] ① Add polyvinyl alcohol to deionized water, heat and stir to dissolve to prepare polyvinyl alcohol solution;

[0023] ② Add PEDOT:PSS containing ethylene glycol to the polyvinyl alcohol solution and stir evenly to obtain a mixed solution;

[0024] ③ Place the mixed solution in a mold, freeze it at -10°C to -25°C, and then thaw it at room temperature. Repeat this freeze-thaw cycle at least 3 times.

[0025] Furthermore, in step ②, 1-10 mL of PEDOT:PSS containing 10% ethylene glycol is added to every 1 g of polyvinyl alcohol.

[0026] Furthermore, in step ③, the freeze-thaw cycle is freezing for 8 hours and thawing for 3 hours.

[0027] Furthermore, the operation of sputtering and depositing the copper drain electrode in step S2.2 is as follows:

[0028] A stainless steel acupuncture needle AN was placed in a high vacuum sputtering chamber, the bias power supply was set to 100 V, the sputtering power was set to 200 W, argon gas was introduced and the flow rate was adjusted to 60 sccm, and a copper drain was deposited by coating.

[0029] Based on the same inventive concept, the present invention also provides a glucose sensor prepared by the above-mentioned method for preparing a glucose sensor based on acupuncture needles.

[0030] Based on the same inventive concept, the present invention also provides an application of a glucose sensor prepared by a method for preparing an acupuncture needle-based glucose sensor in preparing a product for detecting a liquid sample or real-time monitoring of glucose in blood of an organism. The organism includes a human body.

[0031] Based on the same inventive concept, the present invention also provides a method for detecting glucose for non-diagnostic purposes, comprising the following steps:

[0032] The glucose sensor is placed on a probe station; the source and drain electrodes of the probe station are respectively led out using electrical wires, and then the electrodes on the probe station are respectively connected to the source and drain electrodes on the glucose sensor using conductive clips. The liquid sample is then used as a liquid gate, and the tip sensing part of the glucose sensor is inserted into the PBS solution. After the probe station applies bias and gate voltage, the glucose content in the liquid sample can be detected.

[0033] The beneficial effects achieved by the present invention are:

[0034] The sensor, which is based on acupuncture needles and a specific structural design and molecular recognition mechanism, can detect glucose concentration with high specificity and sensitivity through changes in drain current, and has an ultra-wide dynamic detection range (0-40mmol·L -1 ), covering clinical hypoglycemia (<3.8mmol·L -1 ) to severe hyperglycemia (>10mmol·L -1 ) range, and the detection limit is as low as 54µmol·L -1 .

[0035] This invention fabricates an organic electrochemical transistor (OECT) by combining acupuncture needles with hydrogels. During sensor fabrication, the needles are roughened and then modified with metallic Au nanoparticles, significantly increasing the number of surface active sites, providing a foundation for subsequent reactions and a stable electrochemical interface. Subsequently, the OECT sensor is fabricated through multilayer construction and insulation treatment. During the functional layer integration stage, PEDOT:PSS hydrogel serves as the core conductive channel material. Its inherent high conductivity and ion-electron coupling properties give the OECT sensor excellent electrical properties, enabling more sensitive glucose detection.

[0036] The present invention rationally adjusts the usage ratio of polyvinyl alcohol and PEDOT:PSS with ethylene glycol. When 4 mL of PEDOT:PSS containing 10% ethylene glycol is added to every 1 g of polyvinyl alcohol, the PVA / PEDOT:PSS hydrogel immobilized on the sensor has the highest redox peak current and the lowest charge transfer impedance. The diffusion coefficient, ion mobility and conductivity are all optimized, and the ion transport and conductivity properties are most outstanding.

[0037] Experiments have shown that the present invention has excellent gate regulation capabilities. During the detection process, the device exhibits a significant switching ratio and fast response characteristics, meeting the biosensor's requirements for sensitivity and real-time performance. It also has a fast response capability to gate voltage changes, indicating that it has good stability and repeatability under repeated operations, and can provide direction and basis for the development of clinical blood glucose monitoring technology with continuous, real-time and accurate characteristics. BRIEF DESCRIPTION OF THE DRAWINGS

[0038] Attachment Figure 1 The preparation process and functionalization diagram of the acupuncture needle OECT sensor of the embodiment are shown in FIG. Figure 1 (a) Step-by-step preparation process of acupuncture needle transistor sensor; Appendix Figure 1 (b) is a schematic diagram of PEDOT:PSS hydrogel fixation; Figure 1 (c) is the gate functionalization process.

[0039] Attachment Figure 2 The preparation and molding of PVA / PEDOT:PSS hydrogel in the embodiment are shown as follows: Figure 2 (a) is a schematic diagram of the preparation; Figure 2 (b) is the actual molding diagram.

[0040] Attachment Figure 3 Electrochemical characterization of PVA / PEDOT:PSS hydrogels: Figure 3 (a) Cyclic voltammetry (CV) test diagram of PVA / PEDOT:PSS hydrogel with different PEDOT:PSS addition amounts (0-4mL); Figure 3 (b) Electrochemical impedance spectroscopy (EIS) test graph corresponding to different ratios of hydrogels; Figure 3 (c) is the CV test graph of PEDOT:PSS-4 hydrogel at a scan rate of 50-300mV / s; Figure 3 (d) Based on the Figure 3 (c) Linear fit plot of the data.

[0041] Attachment Figure 4 The ductility, self-healing and resilience of the PVA / PEDOT:PSS hydrogel in the embodiment.

[0042] Attachment Figure 5 The mechanical properties test curves of hydrogels with different PEDOT:PSS contents in the examples are as follows: Figure 5 (a) is the tensile stress-strain curve; Figure 5 (b) is the compressive stress-strain curve.

[0043] Attachment Figure 6 Surface morphology analysis of acupuncture needle OECT sensor for example: Figure 6 (a) is the overall SEM image of the cross section; Figure 6 (b) is the cross-sectional enlarged SEM image; Figure 6 (c) is the enlarged SEM image of the Cu layer; Figure 6 (d) is the enlarged SEM image of the gold layer; Figure 6 (e) is the EDS diagram of Fe element; Figure 6 (f) is the EDS image of Au element; Figure 6 (g) is the EDS image of Cu element.

[0044] Attachment Figure 7 The electrical characteristics of the OECT device of the embodiment are analyzed: Figure 7 (a) is a schematic diagram of acupuncture needle OECT biosensor detection; Figure 7 (b) is the VD-ID characteristic curve when VG = 0 to 0.6V, with a step size of 0.1V; Figure 7 (c) is the VG-ID characteristic curve, VDS=0.5V; Figure 7 (d) Real-time current measurement with different VG (0.1 V, 0.2 V, 0.3 V, 0.4 V, 0.5 V, 0.6 V) applied in PBS, VDS = 0.5 V; Figure 7 (e) Real-time current measurement repeated 5 times with different VG (0.1 V, 0.5 V) applied in PBS, VDS = 0.5 V.

[0045] Attachment Figure 8 Acupuncture needle OECT sensor for detecting glucose: Attached Figure 8 (a) Sensitivity test of different concentrations of glucose in PBS solution; Figure 8 (b) The linear relationship between peak current and glucose concentration. DETAILED DESCRIPTION

[0046] To make the objectives, technical solutions, and advantages of the embodiments of the present invention more clear, the technical solutions in the embodiments of the present invention will be clearly and completely described below in conjunction with the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. All other embodiments obtained by ordinary technicians in this field based on the embodiments of the present invention without making any creative efforts shall fall within the scope of protection of the present invention.

[0047] Example 1: Preparation of a glucose sensor based on acupuncture needles

[0048] Raw materials used in this embodiment:

[0049] Chloroauric acid, AR, was purchased from Anaiji Chemical Reagent Co., Ltd.;

[0050] Novolac epoxy resin, AR, was purchased from Anaiji Chemical Reagent Co., Ltd.;

[0051] PEDOT:PSS, AR, purchased from Anaiji Chemical Reagent Co., Ltd.;

[0052] Polyvinyl alcohol, AR, was purchased from Anaiji Chemical Reagent Co., Ltd.;

[0053] Glucose, AR, was purchased from Anaiji Chemical Reagent Co., Ltd.;

[0054] Phosphate buffer, AR, was purchased from Solarbio Life Sciences;

[0055] Disposable sterile acupuncture needles (stainless steel needles AN), 0.35 mm × 25 mm, Changchun Aikang Medical Instrument Co., Ltd.;

[0056] Unless otherwise specified, the experimental methods used in the following examples are conventional methods.

[0057] Unless otherwise specified, the materials and reagents used in the following examples can be obtained from commercial sources.

[0058] The following abbreviations are used:

[0059] AuNPs: gold nanoparticles; AN: stainless steel acupuncture needle; CV: cyclic voltammetry; OECT: organic electrochemical transistor; PVA: polyvinyl alcohol;

[0060] like Figure 1 As shown, the preparation of the acupuncture needle-based glucose sensor includes the following steps:

[0061] S1: Substrate pretreatment

[0062] A stainless steel acupuncture needle (AN) was used as a base and roughened: sandpaper was used to roughen the surface. After polishing, the needle was ultrasonically cleaned in ultrapure water and anhydrous ethanol for 15 minutes. After cleaning, the needle was dried and set aside.

[0063] The pretreated AN was placed in 8 mmol·L -1 A three-electrode system was constructed by inserting a reference electrode and a counter electrode into a solution of HAuCl. This was connected to an electrochemical workstation and subjected to 20 cycles of CV deposition in the range of -1.5 V to 0.5 V at a scan rate of 100 mV·s⁻¹, forming a uniform AuNP layer. The AN was then rinsed three times with deionized water to remove any residual HAuCl.

[0064] S2: Multi-layer structure construction and insulation treatment

[0065] S2.1 Coating the surface of the AuNPs-modified stainless steel acupuncture needles pretreated in step S1 with phenolic epoxy resin, placing them in a drying oven, and drying them at 70°C for 2 h to form an insulating layer;

[0066] S2.2 Place the stainless steel acupuncture needle with an insulating layer prepared in step S2.1 into a high vacuum sputtering chamber, set the bias power supply to 100V and the sputtering power to 200W, introduce argon and adjust the flow rate to 60sccm, and deposit the copper drain electrode after coating for 3 hours.

[0067] S2.3 Coat the stainless steel acupuncture needle with phenolic epoxy resin again to cover the copper layer. After curing, use a grinder to polish the needle tip until the cross section exposes the Au / Cu bimetallic layer.

[0068] S3: Functional layer integration and gate modification

[0069] S3.1 Constructing a conductive channel

[0070] Using a 10uL pipette, the prepared PEDOT:PSS hydrogel was fixed on the cross section of the AN sensor to form a conductive channel connecting the gold source and copper drain electrodes;

[0071] The preparation of PEDOT:PSS hydrogel in this step is as follows:

[0072] Weigh 1g of polyvinyl alcohol (PVA) and add 9ml of deionized water. Pour the mixture into a container and place it on a heated stirring device at 95°C. Stir continuously until the PVA is completely dissolved. After the solution cools to room temperature, add PEDOT:PSS containing 10% ethylene glycol and stir until uniform. The mixed solution is then injected into molds and frozen at -20°C for 8 hours. Thaw at room temperature for 3 hours. Repeat this freeze-thaw cycle three times to obtain a PEDOT:PSS hydrogel.

[0073] In this step, five groups of experiments were set up according to the ratio of the added amounts of polyvinyl alcohol and PEDOT:PSS, namely 0 mL, 1 mL, 2 mL, 3 mL, and 4 mL. The hydrogels prepared in each group were named PEDOT:PSS-0, PEDOT:PSS-1, PEDOT:PSS-2, PEDOT:PSS-3, and PEDOT:PSS-4, respectively.

[0074] S3.2 Gate functionalization modification

[0075] Silver wire was selected as the gate and immersed in 10mmol·L -1 In the EDOT-FPBA solution, constant potential method was used for electropolymerization, the voltage was set to 1.5V, the electropolymerization time was 20s, and a poly (EDOT-FPBA) molecular recognition layer was formed on the gate surface, thus completing the sensor preparation.

[0076] Characterize the intermediate products and final products of the acupuncture needle OECT sensor preparation process

[0077] Some of the main instruments used for characterization:

[0078] Electrochemical workstation, model: INTERFACE1010E, Gamry Instruments, USA;

[0079] Scanning electron microscope, model: S4800, Hitachi, Japan;

[0080] X-ray powder diffractometer, model: Smartlab, Hitachi, Japan;

[0081] X-ray photoelectron spectroscopy analysis, model: AXISSUPRA+Shimadzu Corporation, Japan;

[0082] Fourier transform infrared spectrometer, model: NicoletiS5, Thermo Fisher Scientific, USA;

[0083] Infrared thermal imager, model: AmericaT62101, PI400i, Optris Electronic Technology Co., Ltd.

[0084] Electronic balance, model: BS2245 Sartorius Scientific Instruments Co., Ltd.;

[0085] Tube furnace, model: OTF-1200X Hefei Kejing Material Technology Co., Ltd.

[0086] Semiconductor analysis tester, model: 4200A-SCS Keithley Instruments, USA;

[0087] Vacuum drying oven, model: DZX-6020B Shanghai Fuma Experimental Equipment Co., Ltd.

[0088] Heating magnetic stirrer, model: S10-3 Shanghai Silu Instrument Co., Ltd.

[0089] High vacuum doba magnetron sputtering coating system, model: JCP500, Beijing Techno Technology Co., Ltd.;

[0090] H NMR spectrum, model: 500 MHz, Bruker, Germany;

[0091] Ultra-performance liquid chromatography-mass spectrometry, model: XevoG2-XSQTof, Waters Technology Co., Ltd.

[0092] 1) Electrochemical characterization of PVA / PEDOT:PSS hydrogel

[0093] Five groups of PVA / PEDOT:PSS hydrogel samples were processed into consistent circular sheets. The hydrogel sheets were tightly fixed on the surface of a glassy carbon electrode. In the CV test, an electrochemical workstation was used in a three-electrode system (with an Ag / AgCl electrode as the reference electrode and a platinum wire as the counter electrode). The modified electrode was immersed in a solution containing 5 mmol·L -1 [Fe(CN)6] 3- / 4-Cyclic voltammetry was performed in a potential window of -0.2 V to 0.6 V at a scan rate of 100 mV·s-1. The electrochemical activity and reversibility of the hydrogel material were evaluated by analyzing the changes in redox peak current and peak potential.

[0094] Electrochemical impedance spectroscopy (EIS) was used to investigate the interfacial charge transfer characteristics of the material. The test was conducted in the same three-electrode system with an electrolyte containing 5 mmol·L -1 [Fe(CN)6] 3- / 4- The test was performed with a PBS solution containing 50 mV of sine wave voltage applied to a 0.2 V DC bias voltage. The entire test was performed at room temperature, and each sample group was measured three times under the same experimental conditions.

[0095] 2) Reaction kinetics test of PVA / PEDOT:PSS hydrogel

[0096] PEDOT:PSS-4 hydrogel was prepared in the presence of 5 mM [Fe(CN)6] 3- / 4- The scanning rates (10 mV·s -1 , 50mV·s -1 , 100mV·s -1 , 150mV·s -1 , 200mV·s -1 , 250mV·s -1 、300mV·s -1 ) Cyclic voltammetry scans were performed in the potential window of -0.2 V to 0.6 V.

[0097] 3) Mechanical properties test of PVA / PEDOT:PSS hydrogel

[0098] Mechanical properties tests were performed using an AGX-v500N electronic universal testing machine. For the tensile stress-strain curve test, the sample was made into a dumbbell shape with a total length of 50 mm, a width of 4 mm, and a thickness of 2 mm, and the tensile rate was 5 mm / min. For the compressive stress-strain curve test, the sample was made into a cylindrical shape with a height of 20 mm and a diameter of 10 mm, and the compression rate was 5 mm / min. -1 , read the maximum compression strength.

[0099] 4) Electrical signal detection of acupuncture needle OECT sensor

[0100] An acupuncture needle OECT sensor (PEDOT:PSS-4 hydrogel) was placed on a probe station. Wires were used to connect the source and drain electrodes of the probe station to the source and drain electrodes of the OECT sensor using conductive clips. Electrical testing was then performed by inserting the tip sensing portion of the OECT sensor into a PBS solution. After applying bias and gate voltages to the probe station, the output and transfer characteristic curves of the device were obtained using a semiconductor analyzer.

[0101] 5) Quantitative detection of glucose

[0102] An acupuncture needle OECT sensor (PEDOT:PSS-4 hydrogel) was placed on a probe station. Wires were used to connect the source and drain electrodes of the probe station to the source and drain electrodes of the OECT sensor using conductive clips. Electrical testing was then performed. A PBS solution containing 0-40 mmol·L⁻¹ of glucose was used as a liquid gate. The tip sensing portion of the OECT sensor was inserted into the PBS solution. After applying bias and gate voltages from the probe station, a semiconductor analyzer was used to obtain the sensor's output characteristic curves in solutions of varying glucose concentrations. By analyzing the changes in the output characteristic curves, the electrical response of the dual-needle sensor to solutions of varying glucose concentrations was calculated, and a working curve was constructed to assess its sensitivity to glucose in solution.

[0103] The above characterization results are shown below:

[0104] 1) Preparation and molding of PEDOT:PSS hydrogel

[0105] Attachment Figure 2 The preparation mechanism and morphology shaping ability of PVA / PEDOT:PSS hydrogel are demonstrated. Figure 2 As shown in (a), the hydrogel is constructed through the synergistic effect of chemical crosslinking and physical crosslinking: PEDOT:PSS nanofibers are mixed with PVA viscous solution, and PEDOT (blue chain), PSS (green chain) and PVA (purple chain) in the system form chemical crosslinks through chemical bonds (red nodes). At the same time, intermolecular interactions promote physical crosslinking, eventually forming a PVA / PEDOT:PSS hydrogel with a three-dimensional network structure. The illustration further analyzes the interaction mechanism at the molecular level and reveals the microscopic behavior of the synergistic crosslinking between components. Figure 2 (b) shows the physical molding effect of the hydrogel, which can be processed into a variety of regular shapes such as heart, star, cat, and bone, reflecting the excellent processing plasticity and shape stability of PVA / PEDOT:PSS hydrogel.

[0106] 2) Electrochemical characterization of PVA / PEDOT:PSS hydrogel

[0107] Attachment Figure 3 The electrochemical performance characteristics of PVA / PEDOT:PSS hydrogel are presented. Figure 3 (a) CV test shows the difference in electrochemical activity of hydrogels with different PEDOT:PSS addition amounts (0-4 mL). As the PEDOT:PSS content increases, the redox peak current increases significantly, indicating that the PEDOT:PSS content has an enhancing effect on the electrochemical activity of the hydrogel; Figure 3 The EIS spectrum of (b) further reveals the conductive properties of the material. As the PEDOT:PSS content increases, the charge transfer impedance gradually decreases, indicating that this component constructs a more efficient electron transfer path for the system, which is consistent with the CV test results. Figure 3 (c) shows the relationship between the current and potential of the electrode surface of PEDOT:PSS-4 hydrogel at a scan rate of 50-300 mV / s. Each curve represents a specific scan rate. As the scan rate increases, the peak current also increases accordingly, indicating that there is a positive correlation between the reaction rate and the scan rate. Figure 3 (d) The peak current is further analyzed ( The quantitative relationship between the peak current and the scan rate (v) is obtained by linear fitting, and the equation is y=0.0068x-0.0072 with a fitting degree R2=0.9978, which indicates that in the electrochemical reaction studied, the peak current is proportional to the square root of the scan rate, which is consistent with the diffusion-controlled electrochemical reaction characteristics.

[0108] Based on the redox peak currents of PEDOT:PSS-0, PEDOT:PSS-1, PEDOT:PSS-2, PEDOT:PSS-3, and PEDOT:PSS-4 in CV tests, the diffusion coefficient, ion mobility, and conductivity were calculated in combination with electrochemical theoretical formulas. In electrochemical research, the following parameters and their corresponding formulas are often used to analyze in order to gain a deeper understanding of ion transport behavior and the conductive properties of materials:

[0109] Diffusion coefficient (D): used to measure the diffusion rate of ions and can be derived from the Randles-Sevcik formula.

[0110] Its expression is Ip=2.69×10 5 n 3 / 2 D 1 / 2 v 1 / 2 AC, where Ip is the peak current, n is the number of electrons transferred, v is the scan rate, A is the electrode area, and C is the ion concentration. This formula, after obtaining the values ​​of the various parameters in the experiment, allows us to solve for the diffusion coefficient D, thereby clarifying the diffusion rate of ions in the system.

[0111] Ion mobility (μ): This reflects the efficiency of ion directional migration under the influence of an electric field and is calculated using the formula μ = DFz / RT. F is the Faraday constant, z is the ion charge, R is the gas constant, and T is the temperature. This formula relates the diffusion coefficient to ion mobility, taking into account multiple physical constants and the ion's inherent characteristics to accurately describe the ion's ability to migrate in an electric field.

[0112] Conductivity (σ): This reflects a material's ability to conduct electricity and is characterized by the formula σ = μczF. Here, σ represents ion mobility, c represents the total ion concentration in the solution, z represents the ion valence, and F represents the Faraday constant. This formula integrates key factors such as ion mobility and the concentration and valence of ions in the solution, providing a quantitative indicator for evaluating a material's electrical conductivity.

[0113] As shown in Table 1, as the amount of PEDOT:PSS increases from 0 mL to 4 mL, the three indicators are significantly improved. When the amount of PEDOT:PSS is 4 mL, the diffusion coefficient reaches 4.1×10 -6 cm 2 / s, and the ion mobility is 1.6×10 -4 cm 2 / (V·s), the conductivity is 3.1×10 -3 S / cm, both reached the optimal value, indicating that the ion transport and conductivity properties of PVA / PEDOT:PSS hydrogel were most outstanding at this dosage. Therefore, PEDOT:PSS-4 was selected for subsequent experiments.

[0114] Table 1: Diffusion coefficient, ion mobility and conductivity of PEDOT:PSS hydrogels with different dosages

[0115]

[0116] 3) Mechanical properties testing of PVA / PEDOT:PSS hydrogel

[0117] Attachment Figure 4 Demonstrating the ductility, self-healing, and resilience of the PVA / PEDOT:PSS hydrogel. The upper left image shows the hydrogel in its initial, nearly spherical state, approximately 1 cm long. The lower image shows the hydrogel stretched into a long, thin strip, reaching 6.5 cm in length, demonstrating its excellent ductility and ability to withstand significant tensile deformation without breaking.

[0118] In the upper right image, the hydrogel was cut into two parts, showing its damaged state. After the two broken parts were reconnected and left for 1 minute, the hydrogel regained its original shape and properties.

[0119] The three images on the lower right show the hydrogel's resilience testing process. The first image shows the hydrogel's initial shape; the second image shows continued compression by a finger, while the hydrogel maintains its deformed state; and the third image shows the hydrogel returning to a near-initial shape after the finger is removed, demonstrating the hydrogel's excellent resilience and its ability to quickly return to its original shape after being deformed by external forces.

[0120] Attachment Figure 5 Mechanical property test curves for hydrogels with varying PEDOT:PSS content are shown. The tensile and compressive stress-strain curves show significant differences in the fracture stress, fracture strain, and material modulus of the different hydrogel groups. The endpoint stress value of the tensile stress curve for PEDOT:PSS-1 is significantly higher than that of the other groups, indicating optimal tensile strength. However, PEDOT:PSS-3 and PEDOT:PSS-4, due to excessive addition, have lower endpoint stress values, indicating insufficient material strength. Regarding fracture strain, PEDOT:PSS-1 can withstand greater deformation without breaking, demonstrating excellent toughness. However, groups like PEDOT:PSS-3 exhibit brittleness due to low strain values. The material modulus is reflected in the initial slope of the curve. For example, the steep slope of PEDOT:PSS-1 indicates strong resistance to elastic deformation, while the shallow slope of PEDOT:PSS-4 indicates a softer material. The lack of monotonic changes in mechanical properties with PEDOT:PSS addition is due to the nonlinear structural evolution of the composite system with PVA. When the addition amount is appropriate, such as PEDOT:PSS-1, PEDOT:PSS and PVA optimize the cross-linking network through intermolecular forces, forming a uniform and dense structure and improving the mechanical properties; while excessive addition, such as PEDOT:PSS-3 and PEDOT:PSS-4, will destroy the original cross-linking stability of PVA, induce phase separation and form stress concentration points, resulting in performance degradation.

[0121] However, comprehensive electrochemical performance analysis shows that PEDOT:PSS-4 exhibits the highest redox peak current, the lowest charge transfer impedance, and the best diffusion coefficient, ion mobility, and conductivity. Although its mechanical properties are not the best among the groups, its electrical properties fully meet the core requirement of high conductivity for OECT channels, and its mechanical properties still provide the required processing adaptability.

[0122] 4) Morphological characterization

[0123] With the help of SEM and energy-dispersive X-ray spectroscopy (EDS) technology, the acupuncture needle transistor was observed and analyzed at the microscopic level after a series of preparation processes, clearly showing that it has a distinct sandwich structure inside.

[0124] Attachment Figure 6The microstructure and element distribution of the acupuncture needle transistor are shown, revealing the formation process and characteristics of its sandwich structure. Figure 6 (a) shows the overall SEM image of the cross section of the acupuncture needle transistor, in which the cavity structure inside the acupuncture needle and the modified layer on the surface can be seen, indicating that the acupuncture needle has undergone fine pretreatment and modification. Figure 6 (b) is an enlarged SEM image of the cross section, which further reveals the details of the AuNPs layer and the copper layer on the surface of the acupuncture needle, as well as the interface between them and the substrate. It can be seen that the AuNPs layer is evenly covered on the surface of the acupuncture needle, while the copper layer is tightly attached to the epoxy resin layer, forming a stable multilayer structure. Figure 6 (c) and attached Figure 6 (d) shows the magnified SEM images of the Cu layer and the Au layer, respectively. Figure 6 In (c), the uniform deposition of the copper layer can be seen, and the surface is smooth and dense, indicating the successful implementation of the sputtering deposition process. Figure 6 (d) shows the nanoparticle structure of the Au layer, and it can be seen that a stable nanoparticle layer is formed, which is due to the precise control of the electrochemical deposition process. Figure 6 (e) Attachment Figure 6 (f) and Figure 6 (g) Energy dispersive spectroscopic (EDS) images of Fe, Au, and Cu, respectively. The distribution of fluorescence colors allows for a visual visualization of the location and distribution of the Fe substrate, AuNPs layer, and Cu layer within the acupuncture needle transistor. Fe is primarily concentrated within the needle's interior and substrate, while Au is concentrated in the AuNPs layer on the surface. Cu is distributed in the copper layer, further confirming the formation of a sandwich structure.

[0125] The preparation process of the present invention includes substrate roughening treatment, AuNPs modification, preparation of epoxy resin insulation layer, sputtering deposition of copper drain, and then to the overall preparation process of secondary insulation and electrode exposure. Each step is indispensable for the final formation of an acupuncture needle transistor with a clear sandwich structure.

[0126] 5) Electrical characterization

[0127] The acupuncture needle OECT of the present invention uses PEDOT:PSS conductive polymer as the active channel layer, and its structure and test circuit are shown in the attached figure. Figure 7 (a) shows the device. The device is inserted into a PBS solution using acupuncture needles as a conductive channel. The source-drain current (ID) is measured under the conditions of applying source-drain voltage (VDS) and gate voltage (VG) to evaluate its electrical performance. The experiment first scans VDS and gradually increases VG (0-0.6V) to observe the change pattern of ID. The results show (see Appendix) Figure 7As shown in (b), ID decreases with the increase of VG. This shows that positive gate voltage can effectively regulate the channel hole density. This phenomenon is due to the depletion mode working mechanism of OECT: positive VG drives the Na + Ions migrate into the PEDOT:PSS layer, neutralizing the PSS- groups it doped, leading to semiconductor dedoping and a reduction in hole concentration, which in turn inhibits current conduction. Furthermore, good ohmic contact between the PEDOT:PSS layer and the source and drain electrodes was confirmed, as was the typical depletion-mode transistor behavior of the device.

[0128] To further verify the gate regulation capability, the experiment measured I D With V G Changes in Figure 7 (c)). The results show that I D With V G The increase shows a monotonically decreasing trend, for example, V G When it increases from 0V to 0.5V, I D The current drops from approximately 23μA to a near-off state. This response characteristic stems from the ion-electron coupling effect of the OECT: the gate voltage dynamically modulates the channel conductivity by regulating the distribution of ions at the electrolyte / semiconductor interface, ultimately achieving continuous modulation of the drain current. During testing, the device demonstrated a significant on-off ratio and fast response characteristics, meeting the sensitivity and real-time requirements of biosensors.

[0129] Attachment Figure 7 (d) shows the source-drain voltage V DS Real-time current measurements are shown in Figure 1. The drain current ID exhibits a step-wise decrease as the gate voltage increases from 0.1V to 0.6V in a PBS solution. This intuitively demonstrates the real-time current variation of the device under different gate voltages, further demonstrating that gate voltage can effectively regulate drain current, with the current suppression increasing as the gate voltage increases.

[0130] Attachment Figure 7(e) shows the real-time current measurement of two specific gate voltages of 0.1V and 0.5V repeated 5 times in a PBS solution with a source-drain voltage VDS = 0.5V. When the gate voltage is switched between 0.1V and 0.5V, the drain current switches rapidly and stably between two different levels, and the current quickly reaches a stable value after each switch. This not only demonstrates that the OECT device has a fast response capability to changes in gate voltage, but also shows that it has good stability and repeatability under repeated operations, and can stably and effectively control the flow of current by changing the gate voltage in practical applications, meeting the requirements of precise control and reliable performance as a biosensor.

[0131] It can be seen from the above characterization results that the acupuncture needle OECT prepared in the present invention has good electrical properties and switching characteristics, and can effectively control the flow of current under different gate voltages.

[0132] 6) Glucose detection

[0133] The principle of glucose detection using the acupuncture needle OECT prepared in this invention is based on its unique structural design and molecular recognition mechanism. First, the acupuncture needles are pretreated and modified with bimetallic Au nanoparticles, significantly increasing the surface active sites, providing high catalytic efficiency and a stable electrochemical interface for subsequent reactions. Subsequently, the OECT is prepared through multilayer structure construction and insulation treatment. During the functional layer integration stage, PEDOT:PSS hydrogel serves as the core conductive channel material. Its inherent high conductivity and ion-electron coupling properties give the OECT excellent electrical properties.

[0134] In the process of glucose detection in the present invention, the poly (EDOT-FPBA) molecular recognition layer on the gate surface plays a key role. The FPBA (3-fluorophenylboronic acid) group has a specific binding ability to the cis-diol structure of glucose. When the glucose molecule forms a reversible borate ester bond with the borate group of FPBA, it will trigger a conformational change in the recognition layer, thereby changing its electron cloud distribution or surface charge state. This change acts on the gate-electrolyte interface through ion migration in the electrolyte, regulating the gate potential. Since the OECT is a P-type transistor, the hole carrier concentration in its conductive channel is regulated by the gate voltage: when the gate potential increases, the cations (such as Na + ) is embedded into the PEDOT:PSS channel from the electrolyte, neutralizing the negative charge of PSS-, resulting in the PEDOT + The dedoping and conversion to a neutral state reduce the channel conductivity. This conductivity change is directly reflected in the attenuation of the drain current, the amplitude of which is positively correlated with the glucose concentration.

[0135] To improve detection sensitivity, the system uses a constant bias voltage to maintain the transconductance close to its maximum range while avoiding hydrolysis reactions. This amplifies small gate voltage changes into significant drain current signals. A concentration gradient drives molecular migration toward the sensing interface, ensuring a rapid response. This integrated design not only overcomes the stability bottleneck of traditional enzyme-based sensors but also significantly reduces application complexity, providing innovative direction and basis for continuous glucose monitoring solutions.

[0136] 7) Quantitative detection of glucose

[0137] Attachment Figure 8 The results of sensitivity test for different concentrations of glucose in PBS solution are shown, as well as the linear relationship between peak current and glucose concentration. Figure 8 (a) shows the results of different glucose concentrations (from 0 mmol·L -1 to 40 mmol·L -1 ) under the condition of , the drain current (IDS) changes with the drain voltage (VDS). As the glucose concentration increases, the drain current gradually decreases, indicating that the sensor has a good response to the change of glucose concentration. Figure 8 (b) further demonstrates the linear relationship between peak current and glucose concentration. The equation obtained by linear regression analysis is y = -0.3875x + 23.5071, and the correlation coefficient R 2 is 0.9985, indicating a highly linear relationship between the two. In addition, the detection limit (LOD) is 54 µmol·L -1 , indicating that the sensor has high sensitivity and low detection limit, and is suitable for accurate measurement of glucose concentration.

[0138] The acupuncture needle OECT sensor of the present invention has an ultra-wide dynamic detection range (0-40 mmol·L -1 ), can cover hypoglycemia (<3.8 mmol·L -1 ) to severe hyperglycemia (>10 mmol·L -1 ) clinical testing needs, and the detection limit is as low as 54µmol·L -1 .

[0139] The acupuncture needle OECT sensor for glucose detection fabricated in this invention exhibits a "sandwich" structure and exhibits typical P-type transistor characteristics. It effectively regulates drain current using gate voltage, resulting in a significant on / off ratio, rapid response, and excellent stability and repeatability. Glucose detection is achieved through changes in drain current, based on a specific structural design and molecular recognition mechanism. Its ultra-wide dynamic detection range and high sensitivity make it highly promising for high-sensitivity, continuous, and real-time glucose detection.

[0140] The present invention and its embodiments are described above. Such description is not restrictive. The embodiment shown in the embodiment is only one embodiment of the present invention, and the actual structure is not limited thereto. In short, if a person skilled in the art is inspired by the above, and does not deviate from the purpose of the invention, without creatively designing a structure and embodiment similar to the technical solution, they shall fall within the scope of protection of the present invention.

Claims

1. A method for preparing a glucose sensor based on acupuncture needles, characterized in that: It includes the following steps: S1: Substrate pretreatment Stainless steel acupuncture needles AN were used as substrates, which were roughened and electrochemically deposited with AuNPs. S2: Multi-layer structure construction and insulation treatment S2.1 Coat the stainless steel acupuncture needle AN pretreated in step S1 with epoxy resin, and after drying, form an insulating layer; S2.2 sputter-deposit a copper drain electrode onto the stainless steel acupuncture needle AN with an insulating layer prepared in step S2.1; S2.3 The stainless steel acupuncture needle AN prepared in step S2.2 is coated with epoxy resin again. After curing, the needle tip is polished until the Au / Cu bimetallic layer is exposed in the cross section; S3: Functional layer integration and gate modification S3.1 Constructing a conductive channel Fix the PEDOT:PSS hydrogel on the cross section of the stainless steel acupuncture needle AN prepared in step S2.3 to form a conductive channel connecting the gold source electrode and the copper drain electrode; S3.2 Gate functionalization modification A silver wire was selected as the gate and immersed in a solution containing EDOT-FPBA. Constant potential electropolymerization was used to form a poly (EDOT-FPBA) molecular recognition layer on the gate surface, thus completing the sensor preparation.

2. The method for preparing a glucose sensor based on acupuncture needles according to claim 1, characterized in that: The operation of electrochemical deposition of AuNPs in step S1 is as follows: The roughened stainless steel acupuncture needle AN was placed in a tetrachloroauric acid solution, and a reference electrode and a counter electrode were inserted to construct a three-electrode system. The system was connected to an electrochemical workstation, and a CV method was used for electropolymerization cyclic deposition to form an AuNPs layer, which was then cleaned after the reaction was completed.

3. The method for preparing a glucose sensor based on acupuncture needles according to claim 2, characterized in that: The operation of CV electropolymerization is as follows: the CV isotope is induced in the range of -1.5 V to 0.5 V at a rate of 100 mV·s -1 The scan rate was 20 cycles of deposition.

4. The method for preparing a glucose sensor based on acupuncture needles according to claim 1, characterized in that: The preparation method of the PEDOT:PSS hydrogel used in step S3.1 includes the following steps: ① Add polyvinyl alcohol to deionized water, heat and stir to dissolve to prepare polyvinyl alcohol solution; ② Add PEDOT:PSS containing ethylene glycol to the polyvinyl alcohol solution and stir evenly to obtain a mixed solution; ③ Place the mixed liquid in a mold, freeze it at -10℃ to -25℃, and then thaw it at room temperature. Repeat this freeze-thaw cycle at least 3 times.

5. The method for preparing a glucose sensor based on acupuncture needles according to claim 4, characterized in that: In step ②, add 1-10 mL of PEDOT:PSS containing 10% ethylene glycol to every 1 g of polyvinyl alcohol.

6. The method for preparing a glucose sensor based on acupuncture needles according to claim 4, characterized in that: The freeze-thaw cycle in step ③ is 8 h of freezing and 3 h of thawing.

7. The method for preparing a glucose sensor based on acupuncture needles according to claim 1, characterized in that: The operation of sputtering and depositing the copper drain electrode in step S2.2 is as follows: A stainless steel acupuncture needle AN was placed in a high vacuum sputtering chamber, the bias power was set to 100 V, the sputtering power was set to 200 W, argon was introduced and the flow rate was adjusted to 60 sccm, and a copper drain was deposited by coating.

8. A glucose sensor prepared by the method for preparing a glucose sensor based on acupuncture needles according to any one of claims 1 to 7.

9. Use of the glucose sensor prepared by the method for preparing a glucose sensor based on acupuncture needles according to any one of claims 1 to 7 in preparing a product for detecting a liquid sample or for real-time monitoring of glucose in blood.

10. A method for detecting glucose for non-diagnostic purposes, characterized in that: The steps include: The glucose sensor according to claim 8 is placed on a probe station; the source and drain electrodes of the probe station are respectively led out using electrical wires, and then the electrodes on the probe station are respectively connected to the source and drain electrodes on the glucose sensor using conductive clips. Then, the liquid sample is used as a liquid gate, and the tip sensing part of the glucose sensor is inserted into a PBS solution. After the probe station applies bias voltage and gate voltage, the glucose content in the liquid sample can be detected.

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