An organic field effect transistor biosensor based on interface topological engineering and a preparation method and application thereof

Abstract

This invention proposes an organic field-effect transistor (OFET) biosensor based on interface topology engineering, its fabrication method, and its application, belonging to the field of biosensor technology. The OFET biosensor comprises a substrate, electrodes, an organic semiconductor layer, an interface topology modification layer, and a biorecognition probe. Through a kinetically controlled vacuum deposition process, an interface topology modification layer with a nanoscale uniform rough morphology is constructed on the semiconductor surface. This topology induces double-layer overlap at the solid-liquid interface, significantly reducing interface capacitance and physically stretching the effective Debye length, thereby overcoming the Debye shielding effect in the high ionic strength environment of physiological fluids. Simultaneously, the aldehyde groups on the surface of the modification layer achieve covalent fixation of the probe without connecting arms and provide encapsulation protection. This sensor achieves ultrasensitive detection of allergen-specific IgE and exhibits excellent stability and signal-to-noise ratio in undiluted human serum, providing a universal physical sensitization strategy for the point-of-care testing of complex clinical samples.

Description

Technical Field

[0001] This invention belongs to the field of biosensor technology, and particularly relates to an organic field-effect transistor biosensor based on interface topology engineering, its fabrication method, and its application. Background Technology

[0002] In the field of bioelectronics, constructing high-performance interfaces capable of precisely transducing biological signals and solid-state electronic signals is crucial for realizing next-generation personalized medicine and point-of-care testing (POCT). Organic field-effect transistors (OFETs), with their intrinsic signal amplification capabilities, label-free detection, low-cost fabrication, excellent biocompatibility, and solution-processable properties, overcome the dependence of traditional immunoassay techniques on operators and large-scale testing equipment, and are considered ideal biosensing platforms. However, OFET biosensors face a severe "Debye shielding effect" when detecting physiological fluids with high ionic strength (~150 mM), such as blood and serum. Counterions in the solution form a dense electrical double layer at the sensing interface, with a Debye length (approximately 0.7 nm) much smaller than the size of most biomolecules (>5-10 nm), causing the charge of the target molecule to be shielded, preventing any charge change in the semiconductor channel.

[0003] In existing technologies, to circumvent the limitations of Debye shielding, existing strategies are mainly divided into three categories: (1) interface and environment regulation, such as dilution buffer or introducing polymer / graphene interface modification to physically extend the Debye length; (2) probe miniaturization, using small-sized recognition elements (such as antibody fragments, molecular imprints) or directional fixation strategies to shorten the sensing distance; and (3) dynamic field perturbation, using high-frequency electric fields to disrupt the double-layer stability. However, the above methods often face challenges such as impaired biological activity, complex device fabrication, or cumbersome detection procedures, making it difficult to meet the requirements of physiological authenticity and convenience for clinical applications, thus limiting their actual clinical application.

[0004] Therefore, developing a general strategy for novel biosensor structures that are simple to manufacture and can effectively overcome the Debye shielding effect to achieve high-sensitivity detection without damaging the physiological environment or increasing equipment complexity is of great practical significance and urgent need for realizing biosensors suitable for practical applications. Summary of the Invention

[0005] To address the technical problems of low sensitivity and poor stability in liquid environments caused by the Debye shielding effect in existing OFETs biosensors for physiological fluid detection, this invention proposes an organic field-effect transistor biosensor based on interface topology engineering, its fabrication method, and its application.

[0006] To achieve the above objectives, the present invention provides the following technical solution: This invention provides an organic field-effect transistor biosensor based on interface topology engineering, comprising a substrate, a gate electrode, an insulating layer, source and drain electrodes, and an organic semiconductor layer. An interface topology modification layer is deposited on the surface of the organic semiconductor layer, and a biometric probe is fixed on the surface of the interface topology modification layer. The interface topology modification layer has a nanoscale uniform rough morphology and is composed of a small molecule material containing aldehyde functional groups. The biometric probe is fixed to the surface of the interface topology modification layer by covalent bonding with the aldehyde group.

[0007] Furthermore, the material of the organic semiconductor layer is poly{2,5-bis(2-octyldodecyl)-2,3,5,6-tetrahydro-3,6-dioxopyrrolo[3,4-c]pyrrolo-1,4-diyl-alt-[2,2'-(2,5-thiophene)bisthiophene[3,2-b]thiophene]-5,5'-diyl} (abbreviated as PDBT-co-TT polymer).

[0008] Furthermore, the PDBT- co The preparation method of -TT polymer is as follows: Under a nitrogen atmosphere, 200 mg of DPP38 (3,6-bis-(5-bromothiophene-2-yl)-2,5-bis-(2-octyldodecyl)pyrrolo[3,4-c]pyrrolo-1,4-dione), 100 mg of TH289 (2,5-bis-(trimethyltinyl)thieno[3,2-b]thiophene), 5 mg of (CH3C6H4)3P, and 4 mg of C were added. 51 H 42 O3Pd2 and ultra-dry chlorobenzene (20 mL) were added to a Shrek flask and stirred at 130 °C for 72 h. One hour before the reaction, ultra-dry bromobenzene (0.2 mL) was added to the reaction system for end-capping. After the reaction was completed and cooled to room temperature, the product was poured into 100 mL of methanol and 8 mL of concentrated hydrochloric acid for precipitation. The product was then purified by Soxhlet extraction using 200 mL of methanol, 200 mL of ethyl acetate, and 200 mL of chloroform to remove impurities and low molecular weight oligomers. The chloroform extraction solution was then rotary evaporated. Finally, 100 mL of methanol was added to precipitate the polymer, which was then filtered through a 0.45 μm organic microporous membrane and dried in a vacuum oven at 60 °C for 12 h to collect the polymer, which is the PDBT- co -TT polymer.

[0009] Furthermore, the material of the interface topology modification layer is a benzaldehyde derivative containing an aldehyde group (-CHO); even further, the material of the interface topology modification layer is 9,10-bis(4-formylphenyl)anthracene (BFPA).

[0010] Furthermore, the root mean square roughness of the nanoscale uniform rough morphology of the interface topology modification layer is 15.0 nm-17.0 nm; the thickness of the interface topology modification layer is 6 nm-7 nm.

[0011] Furthermore, the biometric probe is ovalbumin (OVA), used to detect specific immunoglobulin E (sIgE).

[0012] This invention also provides a method for fabricating the above-mentioned organic field-effect transistor biosensor based on interface topology engineering, comprising the following steps: (1) Spin-coating an organic semiconductor solution onto the modified substrate and performing thermal annealing to obtain an organic semiconductor layer; (2) An interface topology modification layer material is deposited on the surface of the organic semiconductor layer by vacuum deposition to obtain an interface topology modification layer (BFPA modification layer). (3) The solution containing the bio-identification probe is incubated on the surface of the interface topology modification layer, and non-specific sites are blocked using bovine serum albumin to obtain the organic field-effect transistor biosensor based on interface topology engineering.

[0013] Furthermore, provide heavy n + A doped silicon wafer serves as both the substrate and the gate electrode, with a thermally grown silicon dioxide layer on its surface acting as an insulating layer. In step (1), the specific preparation steps for the modified substrate are as follows: (1) Substrate cleaning: Clean the substrate with the thermally grown SiO2 layer using deionized water, acetone and isopropanol. + For doped silicon wafers, the process is as follows: First, use deionized water to sonicate for 5-15 minutes to remove easily cleanable dust from the substrate surface. Then, use acetone to sonicate for 5-15 minutes to remove difficult-to-clean dust from the substrate surface. Finally, use isopropanol to sonicate for 5-15 minutes for final cleaning. (2) Substrate modification: After treating the substrate with plasma in an oxygen environment, the substrate is modified with octadecyltrichlorosilane (OTS). Specifically, the substrate is first treated with plasma in an oxygen environment for 5-25 min at a power of 100W. Then, the silicon wafer is placed in the OTS modification solution (n-heptane:OTS=1000:1, volume ratio) and left to stand at room temperature for 2-5 h. Then, the silicon wafer is taken out and placed in beakers containing n-hexane solution, chloroform solution and isopropanol solution for ultrasonic treatment for 10 min. (3) Fabrication of source and drain electrodes: Using vacuum thermal evaporation technology, a 30-50 nm thick (preferably 50 nm) gold layer is deposited on a clean substrate through a mask as the source and drain electrodes.

[0014] Furthermore, in step (1), the process of spin-coating an organic semiconductor solution onto the modified substrate and then performing a thermal annealing treatment specifically includes the following steps: PDBT- co -TT polymer was dissolved in o-dichlorobenzene (or chlorobenzene) to prepare a semiconductor solution with a concentration of 5 mg / mL. The solution was heated and stirred overnight on a hot stage to ensure complete dissolution. After the solution cooled to room temperature, it was filtered through a 0.22 μm PTFE filter and spin-coated onto the modified substrate at a speed of 2500-4500 rpm (preferably 4000 rpm). The device was then placed on a glove box hot stage and annealed at 150℃-180℃ for 10-30 min (preferably 180℃ for 15 min) to remove solvent and improve film morphology and flatness, forming a uniform organic semiconductor layer.

[0015] Further, in step (2), the deposition rate of the vacuum deposition method is 2.5-3.5 Å / s (preferably 3.0 Å / s), and the temperature is 45-55℃ (preferably 50℃), which induces the formation of a nanoscale uniform rough morphology.

[0016] Further, in step (3), the amount of solution containing the biometric probe is 20 μL per device; the concentration of bovine serum albumin is 1 wt%, and the amount is 20-50 μL per device.

[0017] The present invention also provides an application of the above-mentioned organic field-effect transistor biosensor based on interface topology engineering in the detection of disease biomarkers in physiological fluids.

[0018] Furthermore, the physiological fluid is whole blood or serum; the disease marker is allergen-specific immunoglobulin E.

[0019] This invention provides an organic field-effect transistor (OFET) biosensor based on interface topology engineering, comprising a substrate, a gate electrode, an insulating layer, an organic semiconductor layer, an interface modification layer, and a biorecognition probe layer. The interface modification layer is deposited on the surface of the organic semiconductor layer, forming a nanoscale uniform rough interface. The nanoscale rough structure of the interface modification layer induces double-layer overlap at the solid-liquid interface, thereby stretching the effective Debye length. The interface modification layer is composed of small molecule materials with aldehyde functional groups, which directly immobilize the biorecognition probe through covalent bonds. Based on this strategy, a functionalized OFET platform successfully achieved ultrasensitive detection of allergen-specific IgE (sIgE), with a detection limit (LOD) as low as 25.77 pg / mL, far below the clinically routine detection threshold (approximately 96%). The final results confirm that, thanks to the effective penetration of ion shielding by the topological structure, this sensor maintains excellent signal-to-noise ratio and specificity even in undiluted human serum samples, providing a simple and universal physical strategy for solving precise biosensing in complex clinical environments.

[0020] Compared with the prior art, the present invention has the following advantages and technical effects: (1) Physical enhancement (overcoming Debye shielding): This invention constructs a nanoscale rough interface, uses the topological structure to restrict the spatial distribution of ions, induces double layer overlap, reduces the interface capacitance, and physically stretches the effective Debye length, enabling the sensor to detect biomolecular charges beyond the Debye length.

[0021] (2) Chemical anchoring (high activity): The interface modification layer has aldehyde groups, which can achieve "zero distance" covalent fixation of the probe without the need for complex connecting arms, shortening the sensing distance while maintaining the bioactivity of the probe.

[0022] (3) Environmental stability: The dense interface modification layer acts as a physical barrier, effectively blocking the erosion of organic semiconductors by water molecules and oxygen, significantly reducing the hysteresis effect (from 14.23mV to 1.57mV), and extending the service life of the device in the liquid phase.

[0023] (4) Ultra-high sensitivity: The theoretical detection limit of this sensor for sIgE is as low as 25.77 pg / mL, and it still maintains excellent performance in undiluted human serum. Attached Figure Description

[0024] The accompanying drawings, which form part of this invention, are used to provide a further understanding of the invention. The illustrative embodiments of the invention and their descriptions are used to explain the invention and do not constitute an undue limitation of the invention. In the drawings: Figure 1 In example (a), PDBT- is prepared by Stille coupling reaction in step (1) of Example 1. coSynthetic route of -TT polymer, (b) is for PDBT- co - Gel permeation chromatography analysis results of TT polymer; Figure 2 The organic semiconductor layer (PDBT-) prepared in step (3) of Example 1 co Atomic force microscopy (AFM) images of the TT layer, where (a) represents thickness, (b) represents surface roughness, and (c) represents three-dimensional surface morphology. Figure 3 The BFPA-modified layer (BFPA / PDBT-) obtained in step (4) of Example 1 co Atomic force microscopy images of the TT layer, where (a) represents the thickness, (b) represents the surface roughness, and (c) represents the three-dimensional surface morphology. Figure 4 (a) is the contact angle between the BFPA modified layer and the PDBT-co-TT layer obtained in step (4) of Example 1, and (b) is the Fourier transform infrared spectrum of the BFPA modified layer and the PDBT-co-TT layer. Figure 5 The electrical performance characterization results of the device with BFPA modification layer obtained in step (4) of Example 1 are as follows: (a) is the transfer curve of NRI-OFETs, (b) is the output curve of VG in the range of 0 to -0.8V (step size of -0.1V), (c) is the mobility of 30 NRI-OFETs, (d) is VTH, (e) is the turn-off current ratio, (f) is the mobility change of unmodified OFET and NRI-OFETs after 20 repeated measurements, (g) is the hysteresis phenomenon of NRI-OFETs and (h) unmodified OFET, and (i) is the performance change of randomly selected NRI-OFETs devices during long-term storage in an atmospheric environment. Figure 6 In the middle (a), the probe OVA is covalently grafted onto the BFPA-modified layer (BFPA / PDBT-). co (a) Schematic diagram of the TT layer, (b) left side is OVA / BFPA / PDBT- co - Atomic force microscopy (AFM) thickness characterization results of the TT layer, with the right side showing the AFM thickness characterization results of the biosensor modified with the target sIgE. (c) shows the OVA / BFPA / PDBT- co -TT layer, BFPA / PDBT- co -TT layer and PDBT- co - High-resolution N 1s XPS spectrum of the TT layer, (d) is the PDBT- modified with fluorescent label OVA. co - Confocal laser scanning microscopy image of TT (left) and BFPA-modified layer (right), (e) is OVA / BFPA / PDBT- co -TT layer, BFPA / PDBT- co -TT layer and PDBT- co - High-resolution C 1s XPS spectrum of the TT layer; Figure 7 The VTH electrical signal of the organic field-effect transistor biosensor based on interface topology engineering obtained in Example 1 is a good linear fit curve when exposed to different concentrations of sIgE allergen biomarkers. Figure 8 This is a schematic diagram of the fabrication process of the organic field-effect transistor biosensor (NRI-OFETs) based on interface topology engineering in Example 1, specifically including substrate and electrode fabrication (i), PDBT- co - Spin coating of TT polymer semiconductor layer (ii), vacuum deposition of BFPA modified layer (iii), covalent grafting of ovalbumin (OVA) probe (iv), specific recognition of target sIgE (v), and PBS buffer addition sensing test (vi). Figure 9 This is a schematic diagram illustrating the mechanism by which the interface topology of the organic field-effect transistor biosensor based on interface topology engineering, obtained in Example 1, induces double electric layer (EDL) overlap to overcome the Debye shielding effect. Figure 10 The selective test results of the device with BFPA modified layer obtained in step (4) of Example 1 for sIgE allergen biomarker sensing; Figure 11 The results of the anti-interference test of the device with BFPA modified layer obtained in step (4) of Example 1 in real physiological fluid. Detailed Implementation

[0025] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as a limitation of the present invention, but rather as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0026] It should be understood that the terminology used in this invention is merely for describing particular embodiments and is not intended to limit the invention. Furthermore, with respect to numerical ranges in this invention, it should be understood that each intermediate value between the upper and lower limits of the range is also specifically disclosed. Every smaller range between any stated value or intermediate value within a stated range, and any other stated value or intermediate value within said range, is also included in this invention. The upper and lower limits of these smaller ranges may be independently included or excluded from the range.

[0027] Unless otherwise stated, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. While only preferred methods and materials have been described herein, any methods and materials similar or equivalent to those described herein may be used in the implementation or testing of this invention. All references to this specification are incorporated by way of citation to disclose and describe methods and / or materials associated with those references. In the event of any conflict with any incorporated reference, the content of this specification shall prevail.

[0028] Various modifications and variations can be made to the specific embodiments described in this specification without departing from the scope or spirit of the invention, as will be apparent to those skilled in the art. Other embodiments derived from this specification will also be obvious to those skilled in the art. This specification and embodiments are merely exemplary.

[0029] The terms “include,” “including,” “have,” “contain,” etc., used in this article are all open-ended terms, meaning that they include but are not limited to.

[0030] This invention provides an organic field-effect transistor biosensor based on interface topology engineering, comprising a substrate, a gate electrode, an insulating layer, source and drain electrodes, and an organic semiconductor layer. An interface topology modification layer is deposited on the surface of the organic semiconductor layer, and a biometric probe is fixed on the surface of the interface topology modification layer. The interface topology modification layer has a nanoscale uniform rough morphology and is composed of small molecule materials containing aldehyde functional groups. The biometric probe is fixed to the surface of the interface topology modification layer by covalent bonding with the aldehyde group.

[0031] In a preferred embodiment of the present invention, the material of the organic semiconductor layer is poly{2,5-bis(2-octyldodecyl)-2,3,5,6-tetrahydro-3,6-dioxopyrrolo[3,4-c]pyrrolo-1,4-diyl-alt-[2,2'-(2,5-thiophene)bisthiophene[3,2-b]thiophene]-5,5'-diyl} (abbreviated as PDBT-co-TT polymer), PDBT- co The preparation method of -TT polymer is as follows: Under a nitrogen atmosphere, 200 mg of DPP38 (3,6-bis-(5-bromothiophene-2-yl)-2,5-bis-(2-octyldodecyl)pyrrolo[3,4-c]pyrrolo-1,4-dione), 100 mg of TH289 (2,5-bis-(trimethyltinyl)thieno[3,2-b]thiophene), 5 mg of (CH3C6H4)3P, and 4 mg of C were added. 51 H 42O3Pd2 and ultra-dry chlorobenzene (20 mL) were added to a Shrek flask and stirred at 130 °C for 72 h. One hour before the reaction, ultra-dry bromobenzene (0.2 mL) was added to the reaction system for end-capping. After the reaction was completed and cooled to room temperature, the product was poured into 100 mL of stirred methanol and 8 mL of concentrated hydrochloric acid for precipitation. The product was then purified by Soxhlet extraction using 200 mL of methanol, 200 mL of ethyl acetate, and 200 mL of chloroform to remove impurities and low molecular weight oligomers. The chloroform extraction solution was then rotary evaporated. Finally, 100 mL of methanol was added to precipitate the polymer, which was then filtered through a 0.45 μm organic microporous membrane and dried in a vacuum oven at 60 °C for 12 h to collect the polymer, which is PDBT-. co -TT polymer; Among them, PDBT- co The structural formula of the -TT polymer is: ; where n takes the value of an integer between 10 and 100.

[0032] The structural formula of DPP38 is: ; The structural formula of TH289 is .

[0033] In a preferred embodiment of the present invention, the material of the interface topology modification layer is a benzaldehyde derivative containing an aldehyde group (-CHO); more preferably, it is a benzaldehyde derivative 9,10-bis(4-formylphenyl)anthracene (BFPA) containing an aldehyde group (-CHO).

[0034] In a preferred embodiment of the present invention, the root mean square roughness of the nanoscale uniform rough morphology of the interface topology modification layer is 15.0 nm-17.0 nm; the thickness of the interface topology modification layer is 6 nm-7 nm.

[0035] In a preferred embodiment of the present invention, the biometric probe is ovalbumin (OVA) used to detect specific immunoglobulin E (sIgE).

[0036] This invention also provides a method for fabricating the above-mentioned organic field-effect transistor biosensor based on interface topology engineering, comprising the following steps: (1) Spin-coating an organic semiconductor solution onto the modified substrate and performing thermal annealing to obtain an organic semiconductor layer; (2) An interface topology modification layer material is deposited on the surface of an organic semiconductor layer by vacuum deposition to obtain an interface topology modification layer (BFPA modification layer). (3) The solution containing the biometric probe was incubated on the surface of the interface topology modification layer, and non-specific sites were blocked by bovine serum albumin to obtain an organic field-effect transistor biosensor based on interface topology engineering.

[0037] In a preferred embodiment of the present invention, a heavy n is provided + A doped silicon wafer is used as both the substrate and the gate electrode. Its surface has a thermally grown silicon dioxide layer as an insulating layer (commercially available, purchased from the 46th Research Institute of China Electronics Technology Group Corporation). In step (1), the specific preparation steps for the modified substrate are as follows: (1) Substrate cleaning: Clean the substrate with the thermally grown SiO2 layer using deionized water, acetone and isopropanol. + For doped silicon wafers, the process is as follows: First, use deionized water to sonicate for 5-15 minutes to remove easily cleanable dust from the substrate surface. Then, use acetone to sonicate for 5-15 minutes to remove difficult-to-clean dust from the substrate surface. Finally, use isopropanol to sonicate for 5-15 minutes for final cleaning. (2) Substrate modification: After treating the substrate with plasma in an oxygen environment, the substrate is modified with octadecyltrichlorosilane (OTS). Specifically, the substrate is first treated with plasma in an oxygen environment for 5-25 min at a power of 100W. Then, the silicon wafer is placed in the OTS modification solution (n-heptane:OTS=1000:1, volume ratio) and left to stand at room temperature for 2-5 h. Then, the silicon wafer is taken out and placed in beakers containing n-hexane solution, chloroform solution and isopropanol solution for ultrasonic treatment for 10 min. (3) Fabrication of source and drain electrodes: Using vacuum thermal evaporation technology, a 30-50 nm thick (preferably 50 nm) gold layer is deposited on a clean substrate through a mask as the source and drain electrodes.

[0038] In a preferred embodiment of the present invention, step (1), spin-coating an organic semiconductor solution onto the modified substrate and performing thermal annealing, specifically includes the following steps: PDBT- co -TT polymer was dissolved in o-dichlorobenzene (or chlorobenzene) to prepare a semiconductor solution with a concentration of 5 mg / mL. The solution was heated and stirred overnight on a hot stage to ensure complete dissolution. After the solution cooled to room temperature, it was filtered through a 0.22 μm PTFE filter and spin-coated onto the modified substrate at a speed of 2500-4500 rpm (preferably 4000 rpm). The device was then placed on a glove box hot stage and annealed at 150℃-180℃ for 10-30 min (preferably 180℃ for 15 min) to remove solvent and improve film morphology and flatness, forming a uniform organic semiconductor layer.

[0039] In a preferred embodiment of the present invention, in step (2), the deposition rate of the vacuum deposition method is 2.5-3.5 Å / s (preferably 3.0 Å / s), and the temperature is 45-55℃ (preferably 50℃), inducing the formation of a nanoscale uniform rough morphology. By controlling the deposition rate and the substrate temperature, a nanoscale uniform rough morphology is induced. This deposition condition is optimized to ensure that the formation not only has a certain roughness to induce double layer overlap, but also maintains the best morphology with uniformity.

[0040] In a preferred embodiment of the present invention, in step (3), the amount of solution containing biometric probes is 20 μL per device; the concentration of bovine serum albumin is 1 wt%, and the amount is 20-50 μL per device.

[0041] The organic field-effect transistor biosensor based on interface topology engineering prepared in this embodiment of the invention can be used to detect disease biomarkers in physiological fluids.

[0042] All raw materials used in this invention were commercially available. Specifically, 3,6-bis-(5-bromothiophene-2-yl)-2,5-bis-(2-octyldodecyl)pyrrolo[3,4-c]pyrrolo-1,4-dione (DPP38) and 2,5-bis-(trimethyltinyl)thieno[3,2-b]thiophene (TH289) were purchased from Detong OPV Co., Ltd.; tris(o-tolyl)phosphine ((CH3C6H4)3P) and bis(benzylmethylacetone)dipalladium (C 51 H 42 O3Pd2 was purchased from Tianjin Haines Optoelectronic Technology Co., Ltd.; phosphate-buffered saline (PBS) (0.1M, pH=7.40) was prepared using NaCl (137mM), Na2HPO4·12H2O (8.10mM), KCl (2.70mM), and KH2PO4 (1.50mM). 9,10-bis(4-formylphenyl)anthracene (BFPA) was purchased from Shanghai Bodi Biotechnology Co., Ltd.

[0043] The substrate used in the embodiments of the present invention is of weight n. + A doped silicon wafer is used as both a substrate and a gate electrode, with a thermally grown silicon dioxide layer on its surface serving as an insulating layer.

[0044] The technical solution of the present invention will be further illustrated by the following embodiments.

[0045] Example 1 A method for fabricating organic field-effect transistor biosensors (NRI-OFETs) based on interface topology engineering includes the following steps: (1) PDBT- co Preparation of TT polymers Under a nitrogen atmosphere, 200 mg of DPP38, 100 mg of TH289, 5 mg of (CH3C6H4)3P, and 4 mg of C were... 51 H 42 O3Pd2 and ultra-dry chlorobenzene (20 mL) were added to a Shrek flask and stirred at 130 °C for 72 h. One hour before the reaction, ultra-dry bromobenzene (0.2 mL) was added to the reaction system for end-capping. After the reaction was completed and cooled to room temperature, the product was poured into 100 mL of methanol and 8 mL of concentrated hydrochloric acid for precipitation. The product was then purified by Soxhlet extraction using 200 mL of methanol, 200 mL of ethyl acetate, and 200 mL of chloroform to remove impurities and low molecular weight oligomers. The chloroform extraction solution was then rotary evaporated. Finally, 100 mL of methanol was added to precipitate the polymer, which was then filtered through a 0.45 μm organic microporous membrane and dried in a vacuum oven at 60 °C for 12 h to collect the polymer, which is PDBT-. co -TT polymer.

[0046] PDBT- co The yield of the -TT polymer was 70%, and the reaction equation was as follows: Figure 1 As shown in (a) (the number-average molecular weight (Mn) of the PDBT-co-TT polymer was determined to be 14.3 kDa by gel permeation chromatography (GPC). Based on the repeating unit structure of PDBT-co-TT, the value of n in the structural formula of the PDBT-co-TT polymer is specifically 14); the product PDBT- co The -TT polymer was characterized, and the results are shown below. Figure 1 In (b), gel permeation chromatography showed that PDBT- co The number-average molecular weight (Mn) of the -TT polymer is 14.3 kDa, indicating that the polymerization reaction was successful and a polymer material with a suitable molecular weight was obtained, which is beneficial for subsequent film formation processing.

[0047] (2) Substrate and electrode fabrication a. Substrate cleaning: Clean the substrate containing the thermally grown SiO2 layer with deionized water, acetone, and isopropanol. + For doped silicon wafers, the process is as follows: First, use deionized water to sonicate for 10 minutes to remove easily cleanable dust from the substrate surface. Then, use acetone to sonicate for 10 minutes to remove difficult-to-clean dust from the substrate surface. Finally, use isopropanol to sonicate for 10 minutes for final cleaning. b. Substrate modification: The substrate was treated with plasma for 10 min in an oxygen environment at a power of 100 W. Then the silicon wafer was placed in an OTS modification solution (n-heptane:OTS=1000:1, volume ratio) and left to stand at room temperature for 3 h. Then the silicon wafer was taken out and placed in beakers containing n-hexane solution, chloroform solution and isopropanol solution in sequence for ultrasonic treatment for 10 min. c. Fabrication of source and drain electrodes: Using vacuum thermal evaporation technology, a 50nm thick layer of gold is deposited on a clean substrate through a mask to serve as the source and drain electrodes, thus obtaining the modified substrate.

[0048] (3) Fabrication of organic semiconductor layer The PDBT- obtained in step (1) co -TT polymer was dissolved in chlorobenzene to prepare a semiconductor solution with a concentration of 5 mg / mL. The solution was heated and stirred overnight on a hot plate to ensure complete dissolution. After the solution cooled to room temperature, it was filtered through a 0.22 μm PTFE filter and spin-coated onto the modified substrate obtained in step (2) at a speed of 4000 rpm. The device was then placed on a glove box hot plate and annealed at 180 °C for 15 min to remove solvent and improve film morphology and flatness, forming a uniform organic semiconductor layer (PDBT- co -TT layer).

[0049] The prepared organic semiconductor layer (PDBT-) co Atomic force microscopy (AFM) images of the -TT layer are shown below. Figure 2 In the figure, (a) represents the thickness, (b) represents the surface roughness, and (c) represents the three-dimensional surface morphology. It can be seen that the thickness of the organic semiconductor layer is 25.38 nm and the surface roughness is 2.05 nm, indicating that the film has good flatness and uniformity.

[0050] (4) Preparation of BFPA modified layer 9,10-bis(4-formylphenyl)anthracene was deposited onto the surface of the organic semiconductor layer obtained in step (3) using vacuum deposition to obtain a BFPA-modified layer (BFPA / PDBT-). co -TT layer), wherein the deposition rate of vacuum deposition is 3.0 Å / s and the substrate temperature is 50 °C.

[0051] The resulting BFPA-modified layer (BFPA / PDBT- co Atomic force microscopy images of the -TT layer are shown below. Figure 3 Where (a) represents thickness, (b) represents surface roughness, and (c) represents three-dimensional surface morphology, it can be seen that, compared with the organic semiconductor layer (PDBT-) obtained in step (3), coCompared to the TT layer, the BFPA modified layer has a dense rough morphology at the nanoscale (root mean square roughness = 16.0 nm), and the total film thickness increases from 25.38 nm in the initial semiconductor layer to 31.99 nm. The calculated thickness of the BFPA modified layer is 6.61 nm.

[0052] The obtained BFPA / PDBT- co The TT layer was characterized by Fourier transform infrared spectroscopy, and the results are shown in [the table below]. Figure 4 In Figure (b), it can be seen that the BFPA-modified layer did not introduce any new functional groups, and the intensity of the -C=O peak increased significantly, indicating the successful deposition of BFPA on the semiconductor layer; the organic semiconductor layer (PDBT- co -TT layer) and BFPA modified layer (BFPA / PDBT- co The contact angle test results between the -TT layers are shown in the figure. Figure 4 As can be seen in (a), the contact angle test shows that the surface angle decreased from 107.6° to 99.9°.

[0053] The electrical performance of the devices (NRI-OFETs) with BFPA-modified layers obtained in step (4) was characterized using a Keithley 4200SCS semiconductor parameter analyzer. The results are shown in [Figure number missing]. Figure 5 Wherein, (a) is the transfer curve of NRI-OFETs, (b) is the output curve of VG in the range of 0 to -0.8V (with a step size of -0.1V), (c) is the mobility of 30 NRI-OFETs, and (d) is the V TH (e) represents the turn-off current ratio; (f) represents the mobility change of the unmodified OFET (Comparative Example 1, the same as Example 1, except that step (4) of preparing the BFPA modification layer is omitted) and NRI-OFETs after 20 repeated measurements; (g) represents the hysteresis phenomenon of NRI-OFETs and (h) the unmodified OFET; and (i) represents the performance change of randomly selected NRI-OFETs devices during long-term storage in an atmospheric environment. The test results show that the electrical performance of the NRI-OFETs devices is well maintained after the deposition of the modification layer, with an average mobility of 0.25 cm⁻¹. 2 V -1 s -1 The average threshold voltage is -224.4mV, and the average on / off ratio is 4.6×10⁻⁶. 3 Furthermore, the introduction of the modification layer significantly improved the device's stability and hysteresis characteristics. The hysteresis voltage for both forward and reverse scanning was reduced from 14.23 mV for unmodified OFETs to 1.57 mV for NRI-OFETs, and the device maintained good performance even after 120 days of storage in an atmospheric environment. These characteristics confirm the electrical basis for high-sensitivity detection in liquid environments using devices with BFPA-modified layers.

[0054] (5) Immobilization of biological probe OVA 20 μL of ovalbumin (OVA) solution was directly added dropwise to the BFPA / PDBT solution obtained in step (4). co -TT layer surface, utilizing BFPA / PDBT- co -The aldehyde groups of BFPA in the TT layer undergo a Schiff base reaction with the amino groups on the OVA surface to achieve spontaneous covalent coupling, as shown in the schematic diagram. Figure 6 (a) After incubation, the surface was rinsed three times with phosphate buffered saline solution to remove unbound protein molecules, yielding a modified layer (OVA / BFPA / PDBT-) immobilized with the biological probe OVA. co -TT).

[0055] Each measurement was performed after the sensing surface was thoroughly rinsed to improve the accuracy and reproducibility of the characterization. Atomic force microscopy (AFM) was used to study the effects of OVA fixation on (OVA / BFPA / PDBT-) co -TT) was highly characterized (see results) Figure 6 (middle (b) left side). With Figure 3 Compared with the BFPA-modified layer without OVA grafting in (a), the relative thickness of the sensing surface increased from 31.99 nm to 35.92 nm after OVA fixation. The increased thickness is consistent with the globular protein size of OVA (4-7 nm), which proves the successful binding of probe molecules on rough interfaces. To more intuitively observe the fixation of OVA on the functional layer BFPA surface, confocal laser scanning microscopy (CLSM) was used for characterization. As a control, fluorescently labeled OVA was fixed on BFPA / PDBT according to the method in step (5). co -TT layer (obtained in step (4)) surface or PDBT- co -TT layer (obtained in step (3)) surface, results are shown in Figure 6 (d) As can be seen from this figure, after the fluorescently labeled OVA is immobilized on the surface of the polymer layer before BFPA molecule modification, the fluorescence signal is sparse and unevenly distributed (physical adsorption, weak binding force); while on the surface of the film modified with BFPA molecules, due to the specific covalent binding between OVA and the aldehyde group of BFPA, dense and uniform green fluorescence is observed. Therefore, modifying BFPA molecules with nano-rough morphology is beneficial for the efficient and high-density immobilization of probes; Before OVA fixation, the sensing surface modified with BFPA was in the N1s spectrum ( Figure 6In (c) only a weak signal (from the polymer) is shown; however, after OVA fixation, a distinct N element characteristic peak was observed at 399.3 eV, and the C1s spectrum ( Figure 6 In (e) the C=O bond binding energy shifts positively to 288.17 eV (corresponding to the peptide bond). The above characterization results fully demonstrate that the nano-roughened sensing surface modified with BFPA molecules can successfully covalently immobilize the biological probe OVA.

[0056] (6) Blocking nonspecific sites The product immobilized with the biological probe OVA in step (5) was used to block non-specific sites with 1 wt% bovine serum albumin (BSA, 20 μL), and then the surface was rinsed three times with phosphate buffered saline solution (PBS, pH = 7.4) and dried at room temperature to obtain an organic field-effect transistor biosensor (NRI-OFETs) based on interface topology engineering.

[0057] The transfer curve of the sensor prepared in Example 1 was tested, and the value of the output signal threshold voltage was obtained. V 0 ; 20 μL of different concentration gradients of sIgE solution (concentrations of 0.1, 0.5, 1, 5, 10, and 100 ng / mL, respectively) were added dropwise to the surface of the sensor prepared in Example 1. The solution was incubated at room temperature and then dried at room temperature to obtain the antibody-modified biosensor. The thickness results of the atomic force microscopy image are shown below. Figure 6 The image on the right of (b) shows the test transfer curve and the value of the output signal threshold voltage. V ; calculate △ V TH = V - V 0 As the sensor output signal, the test results are shown below. Figure 7 By extracting different concentrations △ V TH To obtain the calibration curve. Due to the effective suppression of ion shielding effect by the interface topology and the shortened sensing distance by covalent fixation of the OVA probe through zero-connector arms, the sensor exhibits good linearity in the range of 0.1 ng / mL to 100 ng / mL (R0). 2 =0.991), and the theoretical limit of detection (LOD) is as low as 25.77 pg / mL, which is far below the threshold for routine clinical testing, indicating that the sensor has extremely high sensitivity.

[0058] Based on the foregoing, a schematic diagram of the fabrication process of the organic field-effect transistor biosensor (NRI-OFETs) based on interface topology engineering in Example 1 is shown below. Figure 8 Specifically, this includes substrate and electrode fabrication (i), PDBT- co - The device structure utilizes the BFPA-modified layer to achieve physical encapsulation of the semiconductor channel through spin coating of the TT polymer semiconductor layer (ii), vacuum deposition of the BFPA-modified layer (iii), covalent grafting of the ovalbumin (OVA) probe (iv), specific recognition of the target sIgE (v), and PBS buffer addition sensing test (vi).

[0059] A schematic diagram illustrating the mechanism by which the interface topology of the organic field-effect transistor biosensor based on interface topology engineering, obtained in Example 1, induces double-layer (EDL) overlap to overcome the Debye shielding effect is shown below. Figure 9 In traditional planar interfaces, counterions are tightly packed to form a dense shielding layer, making it impossible to sense the charge of biomolecules. However, in the nanoscale roughened interface (NRI) constructed in this invention, the confined space induces the overlap and diffusion of the electric double layer, physically stretching the effective Debye length, allowing electric field lines to penetrate the ion shielding layer and enabling effective detection of sIgE.

[0060] Interfering protein (IgG, HSA, AFP) solutions and their mixtures with concentrations of 100 ng / mL, 1 μg / mL, and 10 μg / mL were prepared and dropped onto the sensor surface for testing. For comparison, a 100 ng / mL sIgE target solution and its mixture with the aforementioned high-concentration interfering proteins were dropped onto the surface for testing. The selectivity of the BFPA-modified device (NRI-OFETs) obtained in step (4) of Example 1 for sIgE allergen biomarker sensing was tested, and the results are shown in […]. Figure 10 It can be seen that, under the same conditions, interfering proteins, including immunoglobulin G (IgG), human serum albumin (HSA), alpha-fetoprotein (AFP), and mixtures thereof, were tested. The results showed that the VTH changes induced by the interfering proteins were weak, while the sample containing sIgE elicited a significant electrical signal response. This indicates that the binding of the OVA probe to the BFPA-modified layer maintained high biological specificity, and the nano-roughened interface did not cause significant non-specific adsorption. High concentrations of interfering proteins induced minimal electrical signals, while extremely low concentrations of sIgE could elicit a significant response.

[0061] sIgE was dissolved in PBS buffer and undiluted human serum samples to prepare test solutions with concentration gradients of 0.1, 0.5, 1, 5, 10, and 100 ng / mL. These solutions were then added to the sensor surface to measure the threshold voltage change. The interference resistance of the BFPA-modified devices (NRI-OFETs) obtained in step (4) of Example 1 was tested in real physiological fluids. Different concentrations of sIgE were dissolved in PBS buffer and undiluted human serum samples for detection. The results are shown in […]. Figure 11 The results showed that the VTH electrical signal response was almost indistinguishable between the two environments. This result fully demonstrates that the interface topology engineering strategy of this invention successfully overcomes the Debye shielding limitation in complex physiological environments (high ionic strength), giving the sensor the ability to perform direct detection in real serum samples.

[0062] The above are merely preferred embodiments of the present invention, but the scope of protection of the present invention is not limited thereto. Any variations or substitutions that can be easily conceived by those skilled in the art within the scope of the technology disclosed in the present invention should be included within the scope of protection of the present invention. Therefore, the scope of protection of the present invention should be determined by the scope of the claims.

Claims

1. An organic field-effect transistor biosensor based on interface topology engineering, comprising a substrate, a gate electrode, an insulating layer, source / drain electrodes, and an organic semiconductor layer, characterized in that, An interface topology modification layer is deposited on the surface of the organic semiconductor layer, and a biometric probe is fixed on the surface of the interface topology modification layer. The interface topology modification layer has a nanoscale uniform rough morphology and is composed of a small molecule material containing aldehyde functional groups. The biometric probe is fixed to the surface of the interface topology modification layer by covalent bonding with the aldehyde group.

2. The organic field-effect transistor biosensor based on interface topology engineering according to claim 1, characterized in that, The raw material for preparing the organic semiconductor layer is poly{2,5-bis(2-octyldodecyl)-2,3,5,6-tetrahydro-3,6-dioxopyrrolo[3,4-c]pyrrole-1,4-diyl-alt-[2,2'-(2,5-thiophene)bisthiophene[3,2-b]thiophene]-5,5'-diyl}.

3. The organic field-effect transistor biosensor based on interface topology engineering according to claim 1, characterized in that, The raw material for preparing the interface topology modification layer is a benzaldehyde derivative containing an aldehyde group.

4. The organic field-effect transistor biosensor based on interface topology engineering according to claim 3, characterized in that, The benzaldehyde derivative containing an aldehyde group is 9,10-bis(4-formylphenyl)anthracene.

5. The organic field-effect transistor biosensor based on interface topology engineering according to claim 1, characterized in that, The root mean square roughness of the nanoscale uniform rough morphology of the interface topology modification layer is 15.0 nm-17.0 nm; the thickness of the interface topology modification layer is 6 nm-7 nm.

6. The organic field-effect transistor biosensor based on interface topology engineering according to claim 1, characterized in that, The biometric probe is ovalbumin.

7. A method for fabricating an organic field-effect transistor biosensor based on interface topology engineering as described in any one of claims 1-6, characterized in that, Includes the following steps: (1) Spin-coating an organic semiconductor solution onto the modified substrate and performing thermal annealing to obtain an organic semiconductor layer; (2) An interface topology modification layer material is deposited on the surface of the organic semiconductor layer by vacuum deposition to obtain an interface topology modification layer; (3) The solution containing the bio-identification probe is incubated on the surface of the interface topology modification layer, and non-specific sites are blocked using bovine serum albumin to obtain the organic field-effect transistor biosensor based on interface topology engineering.

8. The method for fabricating an organic field-effect transistor biosensor based on interface topology engineering according to claim 7, characterized in that, In step (2), the deposition rate of the vacuum deposition method is 2.5-3.5 Å / s and the temperature is 45-55℃.

9. The method for fabricating an organic field-effect transistor biosensor based on interface topology engineering according to claim 7, characterized in that, In step (3), the amount of solution containing the biometric probe is 20 μL per device; the concentration of bovine serum albumin is 1 wt%, and the amount is 20-50 μL per device.

10. An application of an organic field-effect transistor biosensor based on interface topology engineering as described in any one of claims 1-6 in detecting disease biomarkers in physiological fluids, characterized in that, The physiological fluid is whole blood or serum; the disease marker is allergen-specific immunoglobulin E.

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