Preparation method of high-sensitivity organic electrochemical transistor cortisol sensor

Abstract

The invention provides a preparation method of a high-sensitivity organic electrochemical transistor (OECT) cortisol sensor, and belongs to the crossing field of a biosensing technology and organic electronics. A grid electrode of the organic electrochemical transistor cortisol sensor is a carbon electrode modified by Prussian blue (PB) and molecularly imprinted polymer (MIP) in sequence, an evaporated gold electrode is used as a source electrode and a drain electrode, and a PEDOT: PSS film printed by aerosol is used as a channel material. PB is used as a built-in redox probe, polypyrrole and poly-3-APBA are used as MIP substrates, a bare carbon electrode is modified through electrochemical deposition and electrochemical polymerization, preparation of the cortisol electrochemical sensor is achieved, the electrochemical sensor is used as a grid electrode, OECT is prepared, and high-sensitivity cortisol reagent-free detection is achieved. The preparation method of the cortisol sensor is easy to operate, chemical synthesis is not needed, repeatability is high, the detection method is simple, the sensitivity of the prepared sensor can reach 5.6 microamperes / dec, and the cortisol sensor has good reproducibility.

Description

Technical Field

[0001] This invention belongs to the interdisciplinary field of biosensing technology and organic electronics, and relates to a method for preparing an organic electrochemical transistor (OECT) sensor using a molecularly imprinted polymer (MIP) as the sensing unit. Background Technology

[0002] Sweat, as an easily accessible biofluid, exhibits a significant correlation between its cortisol concentration and the level of free cortisol in the blood, providing an ideal window for non-invasive, continuous monitoring of human stress. However, achieving accurate detection of sweat cortisol faces two major challenges: First, its concentration is extremely low, typically at the nanomolar or even picomolar level (1-400 nM), placing extremely high demands on the sensitivity of the detection method; second, the complex sweat matrix, with various metabolites (such as glucose, urea, uric acid, etc.) easily generating signal interference, necessitates sensors with excellent selectivity.

[0003] To address these challenges, researchers have explored various pathways, from traditional immunoassays to emerging electrochemical sensing. For example, Moreno et al.'s work ("Aptamer-based electrochemical biosensor for cortisol detection in human sweat", Biosensors and Bioelectronics, 2020, 156, 112029) developed an aptamer-based electrochemical sensor. This sensor utilizes gold nanoparticles to modify the electrode to increase the specific surface area and detects cortisol by detecting changes in interfacial electron transfer resistance caused by the binding of cortisol to the immobilized aptamer. This method demonstrates good sensitivity and the long-term stability of the aptamer. However, aptamers are susceptible to protease degradation or non-specific adsorption in real sweat environments, and their complex chemical modification and immobilization processes are not conducive to large-scale, low-cost device fabrication. Another mainstream strategy relies on the high specificity of antibodies. As demonstrated in the research of Zhou's team ("Field-effect transistor biosensor for ultrasensitive cortisol detection with carbon nanofibers as a floating gate", ACS Applied Materials & Interfaces, 2021, 13, 25, 29987-29994), they immobilized cortisol antibodies on the gate of a carbon nanofiber-modified field-effect transistor, amplifying the signal through gate potential drift caused by antigen-antibody binding. The advantage of this method lies in its extremely high molecular recognition specificity, but the inherent drawbacks of antibodies—including high cost, demanding storage conditions, and susceptibility to inactivation during repeated use—limit their application prospects in wearable and regenerable sensors. While the aforementioned bio-based sensors have made significant progress in laboratory settings, their stability and practicality bottlenecks have prompted researchers to turn their attention to more robust "artificial antibodies"—molecularly imprinted polymers. To overcome the limitations of biological elements, Kumar et al. ("Electrochemical sensor for cortisol based on molecularly imprinted polypyrrole synthesized from corticosteroid template", Sensors and Actuators B: Chemical, 2022, 358, 131489) prepared a polypyrrole MIP film with cortisol as the template molecule on the surface of a screen-printed carbon electrode by electropolymerization.After eluting template molecules, the sensor's holes can specifically recombine cortisol, causing a change in the detection signal (such as the differential pulse voltammetric peak current), demonstrating excellent chemical stability and low-cost potential. Specific recognition is converted into a measurable electrical signal. However, the detection sensitivity of such traditional two-electrode or three-electrode sensors is fundamentally limited by background interference from non-Radidatic current (double-layer charging current). For ultra-low concentrations of cortisol in sweat, this background noise often obscures the effective signal, resulting in insufficient signal-to-noise ratio and detection limits that fail to meet practical requirements. Furthermore, most such MIP sensors still require the addition of exogenous redox probes (such as [Fe(CN)6)) to the solution during detection. 3- / 4- This not only increases the number of operational steps and undermines the integration and simplicity of detection, but the potential biocompatibility of the probe also hinders its integrated application in wearable devices. Tang et al. ("Touch-Based Stressless Cortisol Sensing", Advanced. Materials. 2021, 33, 2008465) developed a cortisol MIP electrochemical sensor with Prussian blue as the built-in probe. They used a co-electrodeposition method of polypyrrole and Prussian blue, utilizing a monolayer modified structure of MIP and Prussian blue to modify a screen-printed carbon electrode. However, in actual preparation, due to the reducing properties of pyrrole, Fe can be removed in the deposition solution. 3+ Reduced to Fe 2+ Fe 2+ and [Fe(CN)6] 3- Subsequently, Prussian blue precipitates. As time goes on, the Prussian blue grains grow, and pyrrole is gradually oxidized, which seriously affects the quality of the MIP film, the consistency of device performance, and the reproducibility of the preparation method. Furthermore, since the template removal method is electrochemical peroxidation, the device cannot be regenerated and can only be used once.

[0004] Based on the above background technical analysis, the existing technologies still have the following prominent problems: the sensitivity of existing cortisol electrochemical sensors is insufficient, making it difficult to detect extremely low concentrations of cortisol in sweat; they rely on unstable biometric elements or non-renewable MIP sensors, resulting in high costs and short lifespans; most sensors require external redox probes, which is not conducive to wearable integration applications; and the existing OECT preparation technology has poor repeatability, which restricts its reliability and commercialization prospects. Summary of the Invention

[0005] The purpose of this invention is to overcome the shortcomings of the prior art and provide a high-performance, regenerable OECT sensor for cortisol detection.

[0006] To achieve the above objectives, this invention proposes a method for fabricating a high-sensitivity cortisol sensor based on molecularly imprinted polymers (MIPs) and OECT (Organic Electrochemical Transistor). The method uses a Prussian blue (PB) and MIP-modified carbon electrode as the gate, an aerosol-printed (AJP) PEDOT:PSS film as the channel, and gold electrodes as the source and drain. An organic electrochemical transistor (OECT) cortisol sensor is fabricated using a stepwise deposition method. Due to the regenerability of MIPs and the high signal amplification capability of OECT, device regeneration and high-sensitivity cortisol detection can be achieved.

[0007] The present invention also relates to an organic electrochemical transistor cortisol sensor prepared by the above method, and a method for high-sensitivity detection using the organic electrochemical transistor cortisol sensor.

[0008] This invention creatively combines the high signal amplification characteristics of OECT with a PB built-in probe strategy. A PB / MIP dual-layer modification is constructed on the gate, with each layer modified independently, ensuring device repeatability. The binding of cortisol inhibits the redox reaction of PB, causing a change in gate potential. This minute change is amplified by the OECT channel, thereby achieving high detection sensitivity without the need for any external reagents. Specific advantages are reflected in the following aspects:

[0009] Significantly improved sensitivity: While MIP-based electrochemical sensors already possess a certain level of sensitivity, this invention utilizes the working electrode obtained through electrochemical deposition of PB / MIP as the gate electrode to fabricate an OECT cortisol sensor. By leveraging the high signal amplification characteristics of OECT, the sensitivity is significantly improved.

[0010] Breakthrough in regenerability and stability: This invention fully utilizes the solvent-eluting properties of MIPs to achieve multiple regeneration and reuse of the sensor. By optimizing elution conditions, the hole activity of the MIPs can be maintained after multiple cycles, significantly extending device life and reducing the cost per detection, which is unmatched by antibody-based sensors.

[0011] A revolutionary advancement in device manufacturing repeatability: This invention employs aerosol printing technology to fabricate PEDOT:PSS channels. This digital, mask-free direct-write process enables precise control over the channel geometry and thickness, fundamentally solving the problem of low channel repeatability caused by traditional spin coating and other processes. This lays the foundation for consistent device performance and large-scale production.

[0012] In summary, this invention provides a cortisol detection solution that exhibits significant advantages in sensitivity, selectivity, repeatability, and reusability through synergistic innovation in materials, structure, and process, offering a new technical path for advancing non-invasive and continuous health monitoring technologies. Attached Figure Description

[0013] Those skilled in the art will understand that the accompanying drawings are provided to better understand the invention and do not constitute any limitation on the scope of the invention.

[0014] Figure 1 A schematic diagram of the structure and working principle of a cortisol electrochemical sensor modified with a Prussian blue and molecularly imprinted polymer bilayer.

[0015] Figure 2 The cyclic voltammograms are shown in a table (a, b) after PB deposition, after polymer deposition, and after template elution during the synthesis process, and in a table (c, d) during polymer deposition.

[0016] Figure 3 Scanning electron microscope (SEM) images of the electrode surface at four stages: bare carbon electrode (a), after PB deposition (b), after polymer deposition (c), and after template molecule elution (d).

[0017] Figure 4 Selectivity of the electrochemical sensor for glucose, uric acid, and urea (a) and regenerability after solvent elution (5 regenerations) (b).

[0018] Figure 5 Linear fitting of the response (a) and sensitivity of the electrochemical sensor to cortisol solutions of different concentrations (b).

[0019] Figure 6 The response (a) and linear fitting of the sensitivity of the OECT sensor with gate-modified MIP to different concentrations of cortisol solution (b). Detailed Implementation

[0020] To make the technical solutions, advantages, and objectives of the present invention clearer, various embodiments are described in detail below with reference to the accompanying drawings. It should be understood that the specific embodiments described herein are merely illustrative of the invention and are not intended to limit the invention. Furthermore, the technical features involved in the various embodiments of the present invention described below can be combined with each other as long as they do not conflict with each other.

[0021] Example 1

[0022] This embodiment provides a method for fabricating a high-sensitivity cortisol sensor based on molecularly imprinted polymers and OECT (Optical Electron Conversion). The method comprises two steps: fabrication of a cortisol sensor without the need for an external redox probe using electrochemical deposition and electrochemical polymerization, and OECT fabrication. Step one yields a cortisol electrochemical sensor with certain sensitivity and selectivity. Step two further enhances the sensor's sensitivity by leveraging the signal amplification capability of OECT, and the AJP (Advanced Persistent Electron Conversion) process is used to fabricate the channel, ensuring convenient, rapid, and reproducible device fabrication.

[0023] The first step involves the preparation of a cortisol sensor without the need for an external redox probe through electrochemical deposition and electrochemical polymerization.

[0024] Figure 1 The device structure and sensing mechanism were demonstrated, showing that when the target molecule fills the specific cavity on the MIP, K... + The diffusion channel was blocked, hindering the redox reaction of the redox probe PB, which in turn reduced the peak current signal of its cyclic voltammetry (CV) or differential pulse voltammetry (DPV). Several deposition processes were performed during the fabrication of the electrochemical cortisol sensor. First, PB deposition: Before PB deposition, the electrode was cleaned and activated with 0.5 M dilute sulfuric acid. 100 μL of 0.5 M dilute sulfuric acid was added to the electrode surface, and the electrode was scanned 10 times within the range of -1.5 V to 1.5 V. After activation, the electrode surface was rinsed with deionized water and dried. Using a 0.1 M acetate buffer solution at pH=2 as the starting solution, a deposition solution containing 5 mM FeCl3, 5 mM K3Fe(CN)6, and 0.1 MKCl was prepared. 100 μL of the deposition solution was added to the surface of the screen-printed carbon electrode, and the electrode was scanned once at a scan rate of 0.05 V / s within the range of -0.2 V to 0.4 V. After deposition, gently rinse the electrode surface with deionized water to remove excess ions and loose Prussian blue nanoparticles, and then air dry the electrode. Prepare the MIP polymerization solution: Weigh 68.47 mg 3-APBA, 72.49 mg cortisol, and 74.55 mg KCl, add 9 mL of anhydrous ethanol to aid dissolution, sonicate to dissolve the cortisol and 3-APBA, then add 104 μL of pyrrole and mix well. This solution serves as a stock solution for MIP deposition; to maintain its long-term physicochemical stability, store the solution in a light-proof container at 4°C. MIP deposition: With the PB layer deposited, place the electrode flat with the adapter connected, add 100 μL of the MIP deposition solution to cover the three-electrode system, and use cyclic voltammetry to scan 10 times in the -0.2 V to 0.9 V region at a scan rate of 0.025 V / s. Figure 2(b) illustrates the MIP deposition process. After the MIP layer is deposited, the template molecules should be eluted immediately. In this scheme, an 8% acetic acid aqueous solution is used as the elution solution. The screen-printed carbon electrode is suspended in a beaker containing the 8% acetic acid solution and stirred for 3 hours for elution. Figure 2 (a) and Figure 3 The CV curves and SEM images for each stage fully demonstrate the successful modification of the electrode by the PB layer and MIP.

[0025] Due to the low sensitivity of CV (CV) due to background interference from charging current, DPV (Digital Voltammetry) was chosen as the electrochemical method for detecting cortisol concentration. To improve the accuracy of DPV testing, the electrode was activated by cyclic voltammetry scanning before the test. The scanning range was -0.4 V to 0.4 V, the scanning speed was 0.1 V / s, and the number of cycles was 10, repeated twice until the CV curves completely overlapped. Then, the DPV test was performed with a test range of -0.4 V to 0.4 V, an amplitude of 0.05 V, a pulse period of 0.5 s, a pulse width of 0.25 s, and a sampling width of 0.1 s. Three DPV tests were performed. The mean and standard deviation were calculated. Based on the cortisol concentration in sweat (1 nM to 400 nM), 100 pM, 1 nM, 10 nM, 100 nM, 1 μM, 10 μM, and 100 μM solutions were selected for testing. Each solution was incubated for 10 min before testing.

[0026] like Figure 5 As shown in (a), the sensor has a clear response to cortisol; the DPV peak decreases with increasing cortisol concentration, consistent with the testing principle. Figure 5 In (b), the sensitivity is 0.067 μA / dec, and the correlation coefficient is 0.994. For example... Figure 4 As shown in (a) and (b), the electrode exhibits good selectivity for metabolic disruptors glucose, urea and uric acid, and the sensor can be regenerated by washing with an 8% acetic acid aqueous solution for reuse.

[0027] The second step is OECT preparation, as shown below.

[0028] Cr / Au source and drain electrodes were obtained on a substrate using a thermal evaporation method. Cr was deposited to improve electrode adhesion to the substrate; the Cr layer thickness was 10 nm, and the Au layer thickness was 400 nm. Channels were formed using a rapid, simple, and patternable AJP printing method. The ink was a mixed solution of PEDOT:PSS, deionized water, and GOPS ((3-glycidoxypropyl)trimethoxysilane), with a volume ratio of PEDOT:PSS:deionized water:GOPS = 1:1:1%. Deionized water reduced ink viscosity, and GOPS acted as a crosslinking agent to enhance the mechanical properties of PEDOT:PSS and improve the adhesion of the PEDOT:PSS film to the substrate. A pneumatic atomizer was used with the following parameters: exhaust gas: 50 SCCM, sheath gas: 700 SCCM, and atomizing gas: 600 SCCM. A 200 or 300 μm nozzle was used, and the printing speed was set accordingly. After printing, the electrodes were annealed at 120°C for 30 min, and then the resistance was tested.

[0029] The working electrode portion of the screen-printed carbon electrode prepared in step one is removed and used as the gate in step two. The gate, source, drain, and channel are then precisely bonded to a substrate, with the distance between the gate and channel controlled at 200 μm. After fixing with silicone rubber, the gate-modified cortisol OECT sensor is obtained. Sensor performance is then tested using this sensor. Figure 6 As shown in (a), V is set s =0, V d =-700~0 mV, V g =550~0 mV, the output characteristic curve as a function of gate voltage was scanned in a 0.1M KCl solution with cortisol blank. The test showed that the source-drain current gradually decreased with changing gate voltage, demonstrating good gate control performance. Then, V... d =-400 mV, V g =550~0 mV, scan I in a series of cortisol solutions. d -V g Curve. For example... Figure 6 As shown in (b), as the concentration of cortisol increases, I d Gradually decreasing, for I d Fitting with cortisol concentration, such as Figure 6 As shown in (c), a linear regression curve was obtained, with a sensitivity of 5.6 μA / dec and a correlation coefficient of 0.992. Comparing the sensitivities of the electrochemical cortisol sensor and the OECT sensor, the OECT sensor significantly improved the sensitivity of the original sensor due to its gate control effect and excellent signal amplification capability.

[0030] Those skilled in the art will readily understand that the above description is merely a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, and improvements made within the spirit and principles of the present invention should be included within the scope of protection of the present invention.

Claims

1. A method for preparing an organic electrochemical transistor cortisol sensor, characterized in that, An organic electrochemical transistor cortisol sensor was prepared by using a carbon electrode modified with Prussian blue and molecularly imprinted polymer as the gate, a PEDOT:PSS thin film as the channel, and gold electrodes as the source and drain, through a stepwise deposition method.

2. The method for preparing the organic electrochemical transistor cortisol sensor as described in claim 1, characterized in that, The gate electrode is a carbon electrode modified layer by layer with Prussian blue and molecularly imprinted polymer.

3. The method for preparing the organic electrochemical transistor cortisol sensor as described in claim 2, characterized in that, The deposition solution used for Prussian blue was a mixed solution of FeCl3, K3Fe(CN)6 and KCl prepared with acetic acid buffer solution as the base solution.

4. The method for preparing the organic electrochemical transistor cortisol sensor as described in claim 2, characterized in that, The formation of the molecularly imprinted polymer includes: first preparing a polymer film by electrochemical polymerization, and then removing the template molecules by solvent elution.

5. The method for preparing the organic electrochemical transistor cortisol sensor as described in claim 1, characterized in that, The channel was prepared using aerosol printing technology.

6. An organic electrochemical transistor cortisol sensor prepared by the method described in any one of claims 1 to 5.

7. The organic electrochemical transistor cortisol sensor as described in claim 6, characterized in that, The regenerative properties of the molecularly imprinted polymer are used to regenerate the organic electrochemical transistor cortisol sensor.

8. A method for high-sensitivity detection using an organic electrochemical transistor cortisol sensor as described in claim 6 or 7.

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