Preparation method of field effect transistor sensor based on nanosheet self-assembly morphology transition metal carbide and detection application of cTnI of field effect transistor sensor

Abstract

The invention discloses a preparation method of a field effect transistor sensor based on nanosheet self-assembly morphology transition metal carbide and detection application of cTnI of the field effect transistor sensor. The preparation method comprises the following steps: step 1, preparing nanosheet self-assembly morphology MXene: step a, etching by taking a TiAlCMAX phase as a raw material; step b, dispersing the etched MXene nanosheets in a CTAB (Cetyltrimethyl Ammonium Bromide) template agent solution, and enabling the etched MXene nanosheets to form a structure in which the nanosheets are vertically arranged on an oil-water interface; c, fixing the three-dimensional network structure on the MXene nanosheet by adopting a freeze drying-confinement annealing method; step d, modifying the MXene material prepared in the step c by using a glutaraldehyde GA cross-linking agent; and 2, preparing the field effect transistor sensor of the nanosheet self-assembly morphology transition metal carbide based on the material prepared in the step 1. The sensor prepared by the invention can realize ultra-sensitive detection of cTnI.

Description

Technical Field

[0001] The present invention belongs to the field of biosensors, and in particular relates to a preparation method of a field effect transistor sensor based on a nanosheet self-assembled morphology transition metal carbide and its cTnI detection application. Background Art

[0002] Myocardial infarction (MI) is a serious cardiovascular disease, typically caused by myocardial ischemia. It is one of the leading causes of death from cardiovascular disease, resulting in millions of deaths annually. Therefore, early diagnosis of MI is crucial, allowing timely intervention to prevent further progression and save lives. In the diagnosis of MI, cTnI is a crucial biomarker because it is one of the main components released by myocardial cell damage. cTnI appears in the blood within hours of the onset of MI and persists for a period of time. Therefore, measuring blood cTnI levels can effectively diagnose MI, especially early-stage MI, allowing for timely and effective treatment. Furthermore, monitoring cTnI levels can be used to assess the prognosis of MI, understanding its severity and treatment effectiveness. Therefore, cTnI plays a crucial role in the diagnosis of MI, helping physicians to promptly diagnose MI and implement effective treatment, thereby reducing mortality and disability.

[0003] Therefore, developing a practical biosensor for diagnosing myocardial infarction is crucial. However, due to the complex composition and extremely low concentration (cutoff value of 0.1 ng / mL) of clinical biological samples containing cardiac troponin (cTnI), the detection of cardiac troponin cTnI poses a severe challenge.

[0004] Existing methods typically use field-effect transistor (FET) sensors to detect cardiac troponin (cTnI). However, traditional FET sensors rely on silicon or two-dimensional materials (such as graphene), and suffer from issues such as interfacial charge scattering and poor biocompatibility, resulting in insufficient sensitivity and failing to meet higher requirements.

[0005] MXene, a novel two-dimensional material, offers advantages such as large surface area, excellent conductivity, and ease of modification, making it a promising candidate for electrochemical sensors. MXene's high surface area increases the contact area between the material and the molecule being measured, thereby enhancing sensor sensitivity. Furthermore, MXene's excellent conductivity can enhance the intensity of the electrochemical signal, further improving sensor sensitivity. However, MXene materials do have some drawbacks. While highly conductive, their conventional layered structure has a limited specific surface area, and the surface functional groups (-O, -F) are unevenly distributed, affecting the efficiency of biomolecule immobilization. Consequently, directly using MXene materials to improve existing FET sensors will still present numerous challenges.

[0006] In order to obtain more efficient and accurate cTnI detection results, people have been committed to making better detection sensors. Summary of the Invention

[0007] The purpose of the present invention is to provide a method for preparing a field-effect transistor sensor based on a nanosheet self-assembled morphology of transition metal carbides. The method first prepares a nanosheet self-assembled morphology MXene, specifically by directional stacking to form a three-dimensional porous network structure, significantly improving the specific surface area and active site density. The nanosheet self-assembled MXene is then combined with a FET to prepare an improved FET sensor. The FET sensor can achieve ultra-sensitive detection of cTnI with a detection limit of ≤0.05 pg / mL.

[0008] The present invention is achieved through the following technical solutions: A method for preparing a field effect transistor sensor based on a nanosheet self-assembled morphology of transition metal carbides comprises the following steps: Step 1: Prepare MXene with a self-assembled nanosheet morphology. The preparation method of MXene with a self-assembled nanosheet morphology comprises the following steps: Step a, Precursor Selection: Using Ti3AlC2 MAX phase as the raw material, a hydrofluoric acid (HF)-lithium fluoride (LiF) gradient etching method is used. Etching is performed at a controlled temperature of 45-50°C for 24-36 hours to remove the Al layer while preserving the integrity of the Ti3C2Tx MXene nanosheets. This involves removing the aluminum layer while preserving the complete multilayer structure. Preferably, the hydrofluoric acid (HF)-lithium fluoride (LiF) gradient etching method uses a molar ratio of HF:LiF of 1:4.

[0009] Step b, morphology control: The etched MXene nanosheets are dispersed in a cetyltrimethylammonium bromide (CTAB) template solution, and the etched MXene nanosheets are formed into a vertically aligned structure at the oil-water interface by interfacial self-assembly. Preferably, the concentration of cetyltrimethylammonium bromide (CTAB) in the cetyltrimethylammonium bromide (CTAB) template solution is 10 wt%. Preferably, after the etched MXene nanosheets are formed into a vertically aligned structure at the oil-water interface by interfacial self-assembly, they are rinsed three times with deionized water to remove the CTAB.

[0010] The interfacial self-assembly method refers to dissolving a polymer in an oil phase and dispersing nanomaterials in an aqueous phase. If there are electrostatic, hydrogen bonding, and other interactions between the polymer and the nanomaterial, polymer nanocomposites with electrical, optical, and magnetic functions can be prepared through self-assembly at the water-oil interface.

[0011] Step c, Morphology Fixation: The three-dimensional network structure on the MXene nanosheets obtained in step b is fixed using a freeze-drying-confined annealing method. Preferably, the freeze-drying-confined annealing process includes freezing with liquid nitrogen at -196°C followed by annealing in argon at 400-450°C for 2-5 hours. Preferably, the interlamellar spacing of the fixed three-dimensional network structure is 5-20 nm.

[0012] The principle of this annealing method is to use temperature to regulate surface tension, promote the release of residual stress, and thus construct a unique morphology. This principle and parameters are closely related to the parameters for regulating surface tension.

[0013] The freeze-drying-confined annealing method (-196°C liquid nitrogen freezing, 400-450°C argon annealing for 2-5 hours) is used because temperature changes are conducive to the formation of a layered network. The specific details are the result of optimization. Incomplete freezing (such as temperature below -196°C) will cause the ice crystals to coarsen and destroy the nanopore structure; insufficient annealing temperature / time will lead to insufficient atomic diffusion and uneven interlayer spacing, while excessively high temperature will induce particle sintering.

[0014] Step d, surface functionalization: The MXene material prepared in step c is modified using a glutaraldehyde (GA) crosslinker or EDC@NHS method to construct a dual-functional interface of -CHO and -NH2 covalently bonded on the MXene material, ultimately obtaining the desired nanosheet self-assembled morphology MXene.

[0015] In the process of preparing MXene with a self-assembled nanosheet morphology, the present invention can achieve morphological changes such as delamination and curling of the etched MXene nanosheets into wrinkles because the surfactant properties of CTAB are cleverly utilized. Through the surfactant properties, the effective effect of the van der Waals force between the MXene nanosheet layers is weakened, thereby promoting the delamination of the MXene nanosheets. CTAB is then used as a soft template to promote the curling of the MXene into wrinkles, thereby realizing the construction of a unique semiconductor material. Because this material has a higher specific surface area and active site density than general MXene materials with a conventional layered structure, the sensor material prepared using this material can have higher detection efficiency and better detection accuracy than existing sensors prepared with conventional layered MXene materials.

[0016] The idea behind this approach stems from a serendipitous discovery by the inventors. This discovery stems from molecular dynamics simulations, which revealed a non-classical interaction between the hydrophobic chains of CTAB and the residual -F groups on the surface of MXene. Specifically, they discovered that CTAB's surfactant properties can weaken the effective van der Waals forces between MXene layers, thereby promoting delamination. Specifically, CTAB, as a surfactant, inserts into the interlayers of the MXene material, generating steric hindrance and electrostatic repulsion, effectively weakening the interlayer van der Waals forces of the MXene layered material. This inhibits interlayer aggregation and promotes delamination. This method breaks the widely held academic prejudice that surfactants are only useful for dispersion. Therefore, the present invention first utilizes CTAB's surfactant properties to weaken the effective van der Waals forces between MXene layers and promote delamination. Then, using CTAB as a soft template, it promotes the curling of the MXene into wrinkles. This method achieves the construction of a unique semiconductor material, providing favorable conditions for the preparation of efficient sensors.

[0017] In addition, during the preparation of the nanosheet self-assembled MXene, the present invention uses a clever annealing method to achieve the stable fixation of the morphology of the unique semiconductor material obtained above.

[0018] Step 2: Based on the self-assembled MXene nanosheets prepared in Step 1, a field-effect transistor (FET) sensor with a transition metal carbide nanosheet structure was fabricated. Specifically, a microstructured electrode was fabricated by photolithography, modified with an antibody, and then blocked to create a FET sensor with a gate composed of interdigitated electrodes with a 10 μm interdigital spacing and cTnI antibody-loaded. The sensor's detection mechanism is that cTnI binding to the aptamer causes a change in the electronic transconductance between the MXene nanosheets, resulting in an increase in the transconductance, enabling detection.

[0019] Preferably, the method for preparing a field effect transistor sensor of a transition metal carbide with a nanosheet self-assembled morphology in the step 2 specifically includes the following steps: first, dispersing the nanosheet self-assembled morphology MXene material prepared in step 1 in deionized water to ensure that the concentration of the nanosheet self-assembled morphology MXene material is 2-5 mg / mL, and then ultrasonicating for 30 minutes to form it into a uniform colloidal solution; then, a photolithography process with a mask accuracy of 1 μm and an ultraviolet exposure dose of 100-150 mJ / cm² is used to prepare a gold layer with a thickness of 100 nm and a line width of 10 μm on a silicon substrate, and then spin-coating the MXene solution that has been formed into a uniform colloidal solution on the interdigital electrode to form a conductive film with a thickness of 50-100 nm on the interdigital electrode; then preparing a glutaraldehyde solution, and using the glutaraldehyde solution to covalently modify the cTnI antibody on the interdigital electrode with a conductive film prepared above by glutaraldehyde, and the concentration of the added cTnI antibody is 2 μg / mL, incubated for 2 hours, and the modification reaction time was 30 minutes; finally, the nonspecific sites were blocked with 1% BSA solution for 1 hour, thus making a functional electrochemical sensor for real-time detection of cTnI target molecules.

[0020] More preferably, when the MXene solution that has formed into a uniform colloidal solution is spin-coated on the interdigitated electrodes, the rotation speed is controlled to be 3000 rpm and the time is 30 seconds.

[0021] More preferably, the glutaraldehyde solution is a 0.5%-2.5% (v / v) glutaraldehyde solution buffered in PBS at pH 7.4.

[0022] The invention also discloses a field effect transistor sensor of transition metal carbide with a nanosheet self-assembly morphology prepared based on the method.

[0023] The present invention also discloses an application of a field effect transistor sensor of the transition metal carbide with the nanosheet self-assembled morphology, namely, for detecting cardiac troponin I (cTnI).

[0024] The beneficial effects of the present invention are: The preparation method of the field effect transistor sensor based on the transition metal carbide with nanosheet self-assembly morphology of the present invention effectively weakens the van der Waals force between the layers of the MXene material by utilizing the surfactant properties of CTAB (intercalation between layers and generating steric hindrance and electrostatic repulsion), thereby promoting its delamination. CTAB is then used as a soft template to promote the curling of MXene into wrinkles, thereby realizing the construction of a unique semiconductor material, thereby providing favorable conditions for the preparation of an efficient sensor. The semiconductor material is then used as the basis to make a FET sensor with a gate modified by a nanosheet self-assembly morphology MXene interdigitated electrode and loaded with cTnI antibody. The sensor can achieve 10-10 6 The effective detection of cTnI at a low level of pg / mL is very accurate, and can achieve ultra-sensitive detection with a detection limit of ≤0.05 pg / mL. It can help doctors diagnose whether a patient has myocardial infarction more promptly and accurately and clearly understand its severity. This can facilitate doctors to take accurate and effective treatment measures, thereby reducing the patient's mortality rate. The economic and social benefits are huge, the practicality is strong, and it is worthy of promotion. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] The present invention will now be described by way of example with reference to the accompanying drawings, in which: Figure 1 This is the XRD analysis diagram of the nanosheet self-assembled MXene prepared in an embodiment of the present invention; Figure 2 Figure 2 is an SEM image of a multi-level MXene structure, wherein Figure a is an SEM image of a traditional multi-layered MXene material, and Figure b is an SEM image of a MXene with a self-assembled nanosheet morphology prepared in an embodiment of the present invention; Figure 3 This is the standard curve obtained by detecting cTnI using the sensor prepared in an embodiment of the present invention. DETAILED DESCRIPTION

[0026] All features disclosed in this specification, or all steps in the disclosed methods or processes, except mutually exclusive features and / or steps, can be combined in any manner.

[0027] Any feature disclosed in this specification (including any appended claims, abstract, and drawings), unless otherwise stated, may be replaced by other equivalent or similar features. In other words, unless otherwise stated, each feature is only an example of a series of equivalent or similar features.

[0028] A method for preparing a field effect transistor sensor based on a nanosheet self-assembled morphology of transition metal carbides comprises the following steps: Step 1: Prepare MXene with a self-assembled nanosheet morphology. The preparation method of MXene with a self-assembled nanosheet morphology comprises the following steps: Step a, Precursor Selection: Using Ti3AlC2 MAX phase as the raw material, a hydrofluoric acid (HF)-lithium fluoride (LiF) gradient etching method is used. Etching is performed at a controlled temperature of 45-50°C for 24-36 hours to remove the Al layer while preserving the integrity of the Ti3C2Tx MXene nanosheets. This involves removing the aluminum layer while preserving the complete multilayer structure. Preferably, the hydrofluoric acid (HF)-lithium fluoride (LiF) gradient etching method uses a molar ratio of HF:LiF of 1:4.

[0029] Step b, morphology control: The etched MXene nanosheets were dispersed in a cetyltrimethylammonium bromide (CTAB) template solution. Interfacial self-assembly allowed the etched MXene nanosheets to form a vertically aligned structure at the oil-water interface. The CTAB concentration in the cetyltrimethylammonium bromide (CTAB) template solution was 10 wt%. After the etched MXene nanosheets formed a vertically aligned structure at the oil-water interface through interfacial self-assembly, the CTAB was rinsed three times with deionized water to remove the CTAB.

[0030] Step c, Fixing the Morphology: The three-dimensional network structure on the MXene nanosheets obtained in step b is fixed using a freeze-drying-confined annealing method. This freeze-drying-confined annealing process involves freezing the nanosheets at -196°C in liquid nitrogen followed by annealing at 400-450°C in argon atmosphere for 2-5 hours. The fixed three-dimensional network structure has an interlamellar spacing of 5-20 nm.

[0031] Step d, surface functionalization: The MXene material prepared in step c is modified using a glutaraldehyde (GA) crosslinker or EDC@NHS method to construct a dual-functional interface of -CHO and -NH2 covalently bonded on the MXene material, ultimately obtaining the desired nanosheet self-assembled morphology MXene.

[0032] Step 2: Based on the self-assembled MXene nanosheets prepared in Step 1, a field-effect transistor (FET) sensor with a transition metal carbide nanosheet structure was fabricated. Specifically, a microstructured electrode was fabricated by photolithography, modified with an antibody, and then blocked to create a FET sensor with a gate composed of interdigitated electrodes with a 10 μm interdigital spacing and cTnI antibody-loaded. The sensor's detection mechanism is that cTnI binding to the aptamer causes a change in the electronic transconductance between the MXene nanosheets, resulting in an increase in the transconductance, enabling detection.

[0033] The method for preparing a field effect transistor sensor of a transition metal carbide with a nanosheet self-assembled morphology in the step 2 specifically includes the following steps: first, dispersing the nanosheet self-assembled morphology MXene material prepared in the step 1 in deionized water to ensure that the concentration of the nanosheet self-assembled morphology MXene material is 2-5 mg / mL, and then ultrasonicating for 30 minutes to form a uniform colloidal solution; then, using a photolithography process with a mask accuracy of 1 μm and an ultraviolet exposure dose of 100-150 mJ / cm², a gold layer with a thickness of 100 nm and a line width of 10 μm is prepared on a silicon substrate, and then the MXene solution that has been formed into a uniform colloidal solution is spin-coated on the interdigital electrode to form a conductive film with a thickness of 50-100 nm on the interdigital electrode, and the rotation speed is controlled to be 3000 rpm during spin coating, and the time is 30 seconds; then, a pH of 0.5%-2.5% (v / v) is prepared. 7.4. Prepare a PBS-buffered glutaraldehyde solution, and use this glutaraldehyde solution to covalently modify the interdigitated electrode with a conductive film prepared above with cTnI antibody at a concentration of 2 μg / mL. Incubate for 2 hours, and the modification reaction time is 30 minutes. Finally, block nonspecific sites with 1% BSA solution for 1 hour to produce a functional electrochemical sensor capable of real-time detection of the cTnI target molecule.

[0034] Figure 1 This is the XRD analysis diagram of the nanosheet self-assembled MXene prepared in the embodiment of the present invention; Figure 1 It can be seen that although the morphology of the MXene prepared in the present invention has changed, the angle of 39.2° in its XRD analysis is consistent with that recorded in various literatures, indicating that the material prepared is still a MXene material.

[0035] Figure 2 Figure 2 is an SEM image of a MXene multi-level structure, wherein Figure a is an SEM image of a conventional multi-layered MXene material, and Figure b is an SEM image of a MXene with a self-assembled nanosheet morphology prepared in an embodiment of the present invention. Through this comparison, it can be seen that the method of the present invention is effective and efficient, and can transform a conventional MXene material with a multi-layer structure into a MXene material with a larger interlayer spacing, multiple wrinkles, and a greatly increased specific surface area.

[0036] Effect verification: The sensor prepared above is used to detect cardiac troponin I (cTnI), and the detection steps include: 1) Prepare a sensor modified with cTnI antibody; 2) Dilute cTnI antigen to various concentrations from low to high, thereby preparing 10-10 6pg / mL cTnI standard; 3) The sensor was sequentially tested with different concentrations of cTnI standard, each reaction lasting 15 minutes, and the corresponding current change was recorded; 4) After the test, the response current of each concentration was analyzed. The standard curve was analyzed based on the concentration and response current values ​​to analyze the correlation and detection limit.

[0037] Figure 3 The standard curve of cTnI detection was obtained by using the sensor made of nanosheet self-assembled MXene prepared in the embodiment of the present invention. It can be seen from the standard curve that the sensor can achieve 10-10 6 The effective detection of cTnI at pg / mL is very accurate and can achieve ultra-sensitive detection with a detection limit of ≤0.05 pg / mL.

[0038] The present invention is not limited to the aforementioned specific embodiments, but extends to any new features or any new combination disclosed in this specification, as well as any new method or process steps or any new combination disclosed.

Claims

1. A method for preparing a field effect transistor sensor based on a nanosheet self-assembled morphology of transition metal carbides, characterized in that: The steps include: Step 1: Prepare MXene with a self-assembled nanosheet morphology. The preparation method of MXene with a self-assembled nanosheet morphology comprises the following steps: Step a: Precursor selection: Using Ti3AlC2 MAX phase as the raw material, a hydrofluoric acid (HF)-lithium fluoride (LiF) gradient etching method is used, with temperature-controlled etching at 45-50°C for 24-36 hours to peel off the Al layer while preserving the integrity of the Ti3C2Tx MXene nanosheets. Step b, morphology control: the etched MXene nanosheets are dispersed in a cetyltrimethylammonium bromide (CTAB) template solution, and the etched MXene nanosheets are formed into a vertically aligned structure at the oil-water interface through interfacial self-assembly. Step c, fixing the morphology: using freeze drying-confined annealing method to fix the three-dimensional network structure on the MXene nanosheet obtained in step b; Step d, surface functionalization: The MXene material prepared in step c is modified using a glutaraldehyde (GA) crosslinker or an EDC@NHS method to construct a bifunctional interface of -CHO and -NH2 covalently bonded on the MXene material, ultimately obtaining the desired nanosheet self-assembled morphology MXene; Step 2: Based on the nanosheet self-assembled morphology MXene prepared in step 1, a field effect transistor sensor of a nanosheet self-assembled morphology transition metal carbide is prepared, that is, a microstructure electrode is prepared by photolithography, an antibody is modified, and a blocking method is used to prepare a FET sensor in which the gate is an interdigitated electrode with an interdigital spacing of 10 μm modified with a nanosheet self-assembled morphology MXene and loaded with cTnI antibodies.

2. The method for preparing a field effect transistor sensor based on a nanosheet self-assembled morphology transition metal carbide according to claim 1, characterized in that: In the step 1 and step a, in the hydrofluoric acid HF-lithium fluoride LiF gradient etching method, the molar ratio of hydrofluoric acid HF and lithium fluoride LiF used is HF:LiF = 1:

4.

3. The method for preparing a field effect transistor sensor based on a nanosheet self-assembled transition metal carbide according to claim 1, characterized in that: In the step 1 and step b, the concentration of the cetyltrimethylammonium bromide (CTAB) template solution is 10 wt %.

4. The method for preparing a field effect transistor sensor based on a nanosheet self-assembled morphology transition metal carbide according to claim 1, characterized in that: In the step 1 and step c, the freeze drying-confined annealing method includes freezing with liquid nitrogen at -196°C and then annealing with argon at 400-450°C for 2-5 hours.

5. The method for preparing a field effect transistor sensor based on a nanosheet self-assembled transition metal carbide according to claim 1, characterized in that: In the step 1 and step c, the interlamellar spacing of the fixed three-dimensional network structure is 5-20 nm.

6. The method for preparing a field effect transistor sensor based on a nanosheet self-assembled morphology transition metal carbide according to claim 1, characterized in that: In the step 2, the method for preparing a field-effect transistor sensor of a transition metal carbide with a nanosheet self-assembled morphology specifically includes the following steps: first, dispersing the nanosheet self-assembled morphology MXene material prepared in the step 1 in deionized water to ensure that the concentration of the nanosheet self-assembled morphology MXene material is 2-5 mg / mL, and then ultrasonicating for 30 minutes to form a uniform colloidal solution; then, using a photolithography process with a mask accuracy of 1 μm and a UV exposure dose of 100-150 mJ / cm², preparing a gold layer with a thickness of 100 nm and a line width of 10 μm on a silicon substrate, and then spin-coating the MXene solution that has been formed into a uniform colloidal solution on the interdigitated electrodes to form a conductive film with a thickness of 50-100 nm on the interdigitated electrodes; Glutaraldehyde solution was then prepared and used to covalently modify cTnI antibodies on the interdigitated electrodes with conductive thin films prepared above. The concentration of the added cTnI antibodies was 2 μg / mL, and the cells were incubated for 2 hours, with the modification reaction time being 30 minutes. Finally, nonspecific sites were blocked with 1% BSA solution for 1 hour, thereby producing a functional electrochemical sensor capable of real-time detection of cTnI target molecules.

7. The method for preparing a field effect transistor sensor based on a nanosheet self-assembled transition metal carbide according to claim 6, characterized in that: In the step 2, when the MXene solution that has formed a uniform colloidal solution is spin-coated on the interdigitated electrodes, the rotation speed is controlled to be 3000 rpm and the time is 30 seconds.

8. The method for preparing a field effect transistor sensor based on a nanosheet self-assembled transition metal carbide according to claim 6, characterized in that: In the step 2, the glutaraldehyde solution prepared is a 0.5%-2.5% (v / v) glutaraldehyde solution with a pH of 7.4 and buffered in PBS.

9. A field effect transistor sensor based on a nanosheet self-assembled morphology of transition metal carbides, characterized in that: Prepared according to the preparation method according to any one of claims 1 to 8.

10. Application of a field effect transistor sensor based on a nanosheet self-assembled transition metal carbide prepared by the preparation method according to any one of claims 1 to 8, characterized in that: Used for the detection of cardiac troponin I cTnI.

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