Rare metal doped bi-layer quantum dot modified biosensor and preparation method thereof

Abstract

This disclosure relates to a rare metal-doped double quantum layer modified biosensor and its fabrication method, belonging to the field of photoanode thin film technology. The biosensor includes: a conductive substrate; and a photoanode thin film layer and an aptamer probe layer stacked on one side of the conductive substrate; the photoanode thin film layer includes a BiVO4 thin film layer, a Co-doped TiO2 quantum layer thin film layer, and a Rh-doped SrTiO3 quantum layer thin film layer sequentially disposed thereon. The method includes: 1. depositing a BiVO4 thin film layer on the conductive substrate; 2. alternately depositing TiO2 thin films and CoO2 thin films on the BiVO4 thin film layer. X 1. Form a Co-doped TiO2 quantum layer thin film; 2. Form a Rh-doped SrTiO3 quantum layer thin film; 3. Coat an aptamer probe solution to obtain a biosensor.

Description

Technical Field

[0001] This disclosure relates to the field of photoanode thin film technology for biosensors, and more specifically, to a rare metal-doped double quantum layer modified biosensor and its preparation method. Background Technology

[0002] Semiconductor optoelectronic biosensors are a new type of sensor that combines the advantages of electrochemical and optical biosensors. Their high sensitivity, low cost, and miniaturization meet the current needs of community-based disease screening and have great application prospects in the field of accurate and rapid screening for sudden diseases.

[0003] However, many key technologies still need to be overcome for the application of photoelectrochemical biosensors. PEC biosensing systems involve three major disciplinary issues: "light absorption and utilization," "separation and transport of photogenerated carriers," and "surface reactions of the photoelectrode." In addition, three key technical challenges remain for the application of PEC sensors: how to achieve affinity between the biosensor semiconductor interface and the probe, how to ensure that the introduction of biological probes does not affect the carrier transport process of the photoanode, and how to stably prepare photoanode thin films in large quantities.

[0004] It should be noted that the information disclosed in the background section above is only used to enhance the understanding of the background of this disclosure, and therefore may include information that does not constitute prior art known to those skilled in the art. Summary of the Invention

[0005] The purpose of this disclosure is to overcome the shortcomings of the prior art and provide a rare metal-doped double quantum layer modified biosensor and its preparation method, thereby improving the affinity with the probe and the sensitivity of the sensor.

[0006] According to one aspect of this disclosure, a rare metal-doped double quantum layer modified biosensor includes a conductive substrate, a photoanode thin film layer, and an aptamer probe layer stacked sequentially; the photoanode thin film layer includes a BiVO4 thin film layer, a Co-doped TiO2 quantum layer thin film layer, and a Rh-doped SrTiO3 quantum layer thin film layer stacked sequentially; the BiVO4 thin film layer is disposed on the conductive substrate, and the Rh-doped SrTiO3 quantum layer thin film layer is disposed on the side of the Co-doped TiO2 quantum layer thin film layer away from the conductive substrate.

[0007] In one exemplary embodiment of this disclosure, the thickness of the Co-doped TiO2 quantum layer thin film is 2–5 nm;

[0008] In the Co-doped TiO2 quantum layer thin film, the doping mass ratio of Co to Ti is 2-8%.

[0009] In one exemplary embodiment of this disclosure, the thickness of the Rh-doped SrTiO3 quantum layer thin film is 10 nm to 15 nm;

[0010] In the Rh-doped SrTiO3 quantum layer thin film, the doping mass ratio of Rh to Ti is 0.5–2%.

[0011] In one exemplary embodiment of this disclosure, the thickness of the BiVO4 thin film layer is 150 nm to 200 nm.

[0012] In one exemplary embodiment of this disclosure, the aptamer probe layer includes the base sequence of a DNA aptamer: 5′-CAGCACCGACCTTGTGCTTTGGGAGTGCTGGTCCAAGGGCGTTAATGGACA-3′, for detecting the COVID-19S protein.

[0013] In one exemplary embodiment of this disclosure, the aptamer probe layer includes a DNA aptamer with the base sequence: 5′-GGGGGGCCAAGGCCCAGCCCTCACACA-3′, used for detecting the HIV-21 gene.

[0014] In one exemplary embodiment of this disclosure, the aptamer probe layer includes the base sequence of a DNA aptamer: 5′-GATACTGCGTGCTTGTTCCATA-3′, for detecting the H1N1 gene of influenza A virus.

[0015] According to another aspect of this disclosure, a method for fabricating a rare metal-doped double quantum layer modified biosensor includes the following steps:

[0016] S1. A BiVO4 thin film layer is deposited on a conductive substrate;

[0017] S2. Alternately deposit TiO2 and CoO films on the side of the BiVO4 thin film layer away from the conductive substrate. X A thin film is formed to form a Co-doped TiO2 quantum layer, wherein x≤3 and the doping mass ratio of Co to Ti is 2-8%;

[0018] S3. Obtain an Rh-doped SrTiO3 precursor solution. Coat the side of the Co-doped TiO2 quantum layer film away from the conductive substrate with the Rh-doped SrTiO3 precursor solution. Hold the solution at 450–500°C under inert gas protection for 30–60 min to form an Rh-doped SrTiO3 quantum layer film on the Co-doped TiO2 quantum layer film. The doping mass ratio of Rh to Ti is 0.5–2%.

[0019] S4. Apply aptamer probe solution to the side of the Rh-doped SrTiO3 quantum layer film away from the conductive substrate, and seal the non-specific binding sites with a protein-free blocking solution to form an aptamer probe layer.

[0020] In an exemplary embodiment of this disclosure, step S3, obtaining the Rh-doped SrTiO3 precursor solution includes:

[0021] Tetrabutyl titanate was dissolved in glacial acetic acid solution and stirred for 10-20 min. Rhodium nitrate solution was added according to the required Rh to Ti doping mass ratio to obtain the first mixed solution.

[0022] Add Sr(NO3)2 solution to the first mixed solution and stir to obtain a second mixed solution; after standing, add citric acid solution to the second mixed solution to obtain Rh-doped SrTiO3 precursor solution.

[0023] In one exemplary embodiment of this disclosure, in step S2, TiO2 thin films and CoO are deposited alternately. X The thin film was deposited using atomic layer deposition.

[0024] The photoanode thin film of the biosensor disclosed herein adds a Co-doped TiO2 quantum layer and an Rh-doped SrTiO3 quantum layer to the BiVO4 thin film. The Co and Rh doping constructs an ohmic contact hole transfer path, which, compared to the undoped quantum layer structure, reduces the quantum tunneling barrier during photogenerated hole transport, reduces carrier recombination, and increases the reaction opportunity between carriers and the analyte during detection, thereby improving the sensor's sensitivity. Simultaneously, the TiO2 in the Co-doped TiO2 quantum layer acts as a passivation layer to enhance the stability of the BiVO4 thin film.

[0025] It should be understood that the above general description and the following detailed description are exemplary and explanatory only, and are not intended to limit this disclosure. Attached Figure Description

[0026] The accompanying drawings, which are incorporated in and form part of this specification, illustrate embodiments consistent with this disclosure and, together with the description, serve to explain the principles of this disclosure. It is obvious that the drawings described below are merely some embodiments of this disclosure, and those skilled in the art can obtain other drawings based on these drawings without any inventive effort.

[0027] Figure 1 This is a schematic diagram of a rare metal-doped double quantum layer modified biosensor structure in one embodiment of the present disclosure.

[0028] Figure 2 This is a cross-sectional TEM image of the Rh:STO / Co:TiO2 / BVO photoanode thin film layer in one embodiment of this disclosure.

[0029] Figure 3 This is a cross-sectional HRTEM image of the Rh:STO / Co:TiO2 / BVO photoanode thin film layer in one embodiment of this disclosure.

[0030] Figure 4 This disclosure provides a cross-sectional STEM image and elemental mapping image of a Rh:STO / Co:TiO2 / BVO photoanode thin film layer in one embodiment.

[0031] Figure 5 This is a comparison chart of JV curves for photoanode films with different Rh doping ratios in one embodiment of this disclosure.

[0032] Figure 6 This is a comparison chart of JV curves of photoanode films with different quantum layers in one embodiment of this disclosure.

[0033] Figure 7 This figure shows a comparison of the adsorption capacity of BVO photoanode thin film, Co:TiO2 / BVO photoanode thin film, and Rh:STO / Co:TiO2 / BVO photoanode thin film for COVID-19S protein aptamer probe in one embodiment of this disclosure.

[0034] Figure 8 This is a schematic diagram illustrating the detection results of the COVID-19S protein by a sensor with an Rh:STO / BVO photoanode thin film layer in one embodiment of this disclosure.

[0035] Figure 9 This is a schematic diagram illustrating the detection results of the COVID-19S protein by a sensor with an Rh:STO / BVO photoanode thin film layer in one embodiment of this disclosure.

[0036] Figure 10 This is a schematic diagram illustrating the detection results of the COVID-19S protein by a sensor with an Rh:STO / Co:TiO2 / BVO photoanode thin film layer in one embodiment of this disclosure.

[0037] Figure 11 This is a schematic diagram illustrating the detection results of the COVID-19S protein by a sensor with an Rh:STO / Co:TiO2 / BVO photoanode thin film layer in one embodiment of this disclosure.

[0038] Figure 12 This is a schematic diagram illustrating the detection results of the HIV-21 gene by a sensor with an Rh:STO / Co:TiO2 / BVO photoanode thin film layer in one embodiment of this disclosure.

[0039] Figure 13 This is a schematic diagram illustrating the detection results of the Rh:STO / Co:TiO2 / BVO photoanode thin film layer sensor for the H1N1 gene of influenza A virus, according to one embodiment of this disclosure.

[0040] The attached figures are labeled as follows:

[0041] 1. Conductive substrate; 2. Photoanode thin film layer; 21. BVO thin film layer; 22. Co-doped TiO2 quantum layer thin film layer; 23. Rh-doped STO quantum layer thin film layer; 3. Aptamer probe layer. Detailed Implementation

[0042] Exemplary embodiments will now be described more fully with reference to the accompanying drawings. However, these exemplary embodiments can be implemented in many forms and should not be construed as limited to the embodiments set forth herein; rather, they are provided so that this disclosure will be thorough and complete, and will fully convey the concept of the exemplary embodiments to those skilled in the art. The same reference numerals in the drawings denote the same or similar structures, and therefore detailed descriptions of them will be omitted. Furthermore, the drawings are merely illustrative of this disclosure and are not necessarily drawn to scale.

[0043] Although relative terms, such as "above," are used in this specification to describe the relative relationship of one component of an icon to another, these terms are used only for convenience, such as according to the orientation of the examples shown in the accompanying drawings. It is understood that if the device of the icon is flipped so that it is upside down, the component described as "above" will become the component below. When a structure is "above" another structure, it may mean that the structure is integrally formed on the other structure, or that the structure is "directly" mounted on the other structure, or that the structure is "indirectly" mounted on the other structure through another structure.

[0044] The terms “a,” “one,” “the,” and “the” are used to indicate the existence of one or more elements / components / etc.; the terms “including” and “having” are used to indicate an open-ended inclusion and to mean that there may be other elements / components / etc. in addition to the listed elements / components / etc.; the terms “first,” “second,” etc. are used only as markers and are not a limitation on the number of objects.

[0045] This disclosure provides a rare metal-doped double quantum layer modified biosensor, such as... Figure 1As shown, the system includes a conductive substrate 1, and a photoanode thin film layer 2 and an aptamer probe layer 3 stacked on one side of the conductive substrate 1. The photoanode thin film layer 2 includes a BiVO4 (BiVO4 can be abbreviated as BVO) thin film layer 21, a Co-doped TiO2 quantum layer thin film layer 22 (i.e., a Co:TiO2 quantum layer thin film layer), and a Rh-doped SrTiO3 quantum layer thin film layer 23 (i.e., a Rh:STO quantum layer thin film layer, SrTiO3 can be abbreviated as STO) sequentially disposed on the conductive substrate 1. The BVO thin film layer 21 is disposed on the conductive substrate 1, and the Rh-doped STO quantum layer thin film layer 23 is disposed on the side of the Co-doped TiO2 quantum layer thin film layer 22 away from the conductive substrate 1.

[0046] The photoanode thin film 2 of the biosensor disclosed herein adds a Co-doped TiO2 quantum layer thin film 22 and an Rh-doped STO quantum layer thin film 23 to the single BVO thin film layer 21. Ohmic contact hole transfer pathways are constructed through Co and Rh doping. Compared with the undoped quantum layer structure, this reduces the quantum tunneling barrier during photogenerated hole transport, reduces carrier recombination, and increases the reaction opportunity between carriers and analytes during the detection process, thereby improving the sensor's sensitivity. Simultaneously, the TiO2 in the Co-doped TiO2 quantum layer thin film 22 can act as a passivation layer to improve the stability of the BiVO4 thin film layer.

[0047] In one embodiment of this disclosure, the thickness of the Co-doped TiO2 quantum layer thin film 22 is 2–5 nm. For example, the thickness of the Co-doped TiO2 quantum layer thin film 22 can be 2 nm, 2.5 nm, 3 nm, 3.5 nm, 4 nm, 4.5 nm, or 5 nm.

[0048] In one example, in the Co-doped TiO2 quantum layer thin film 22, the Co-to-Ti doping mass ratio is 2-8%. For example, the Co-to-Ti doping mass ratio can be 2%, 3%, 4%, 5%, 6%, 7%, or 8%.

[0049] In one embodiment of this disclosure, the thickness of the Rh-doped STO quantum layer thin film 23 is 10 nm to 15 nm. For example, the thickness of the Rh-doped STO quantum layer thin film 23 can be 10 nm, 11 nm, 12 nm, 13 nm, 14 nm, or 15 nm.

[0050] In one example, in the Rh-doped STO quantum layer thin film 23, the doping mass ratio of Rh to Ti is 0.5% to 2%. For example, the doping mass ratio of Rh to Ti can be 0.5%, 0.8%, 1%, 1.2%, 1.5%, or 2%.

[0051] In one embodiment of this disclosure, the thickness of the BVO thin film layer 21 is 150 nm to 200 nm. For example, the thickness of the BVO thin film layer 21 can be 150 nm, 160 nm, 170 nm, 180 nm, 190 nm, or 200 nm.

[0052] In one embodiment of this disclosure, the aptamer probe layer 3 includes the base sequence of a DNA aptamer: 5′-CAGCACCGACCTTGTGCTTTGGGAGTGCTGGTCCAAGGGCGTTAATGGACA-3′ (SEQ ID NO:1), which can be used for the specific detection of the COVID-19S protein.

[0053] In one embodiment of this disclosure, the aptamer probe layer 3 includes the base sequence of a DNA aptamer: 5′-GGGGGGCCAAGG CCCAGCCCTCACACA-3′ (SEQ ID NO:2), which can be used for the specific detection of the HIV-21 gene.

[0054] In one embodiment of this disclosure, the aptamer probe layer 3 includes the base sequence of a DNA aptamer: 5′-GATACTGCGTGCTTGTTCCATA-3′ (SEQ ID NO:3), which can be used to specifically detect the H1N1 influenza A virus gene.

[0055] This disclosure also provides a method for fabricating a rare metal-doped double quantum layer modified biosensor, the method comprising the following steps:

[0056] S1. Deposit a BVO thin film layer 21 on a conductive substrate 1; the conductive substrate 1 can be FTO conductive glass or ITO conductive glass.

[0057] In one embodiment of this disclosure, a BVO thin film layer 21 is prepared by electrodeposition of BiOI followed by high-temperature vanadium ion exchange. The method includes the following steps:

[0058] S1.1 Dissolve solid Bi(NO3)3·5H2O in ultrapure water, then add KI in portions and stir for 15-40 minutes each time to ensure that Bi is dissolved in ultrapure water during the KI addition process. 3+ The amount of substance and I - The molar ratio of the substances is ≤1. The pH of the solution is then adjusted to 1.2–1.7 using HNO3, and the resulting solution is the first solution.

[0059] Obtain a p-benzoquinone solution with a concentration of 0.02–0.03 mol / L. For example, p-benzoquinone powder can be dissolved in ethanol and ultrasonically vibrated until completely dissolved to obtain a p-benzoquinone solution. The molar concentration of the p-benzoquinone solution can be 0.02 mol / L, 0.024 mol / L, 0.028 mol / L, or 0.03 mol / L.

[0060] S1.2. Mix the first solution with the p-benzoquinone solution and stir for 30-60 minutes to obtain the second solution.

[0061] S1.3. A three-electrode system consisting of a working electrode, a counter electrode, and a differential electrode is used. The working electrode, counter electrode, and differential electrode are placed in a second solution. The deposition voltage is set to -0.1 to -0.2 V, and the deposition time is set to 30 to 40 s. A BiOI thin film is deposited on one side of the working electrode. For example, the working electrode can be FTO conductive glass (e.g., a 2cm × 1cm × 0.15cm FTO conductive glass substrate), the counter electrode can be a platinum electrode, and the differential electrode can be an Ag / AgCl electrode. A four-channel potentiostat can be used for deposition, and four BiOI thin film electrode plates can be obtained simultaneously after deposition.

[0062] S1.4 Weigh 0.1–0.3 g of vanadium acetylacetonate and dissolve it in 5–10 mL of dimethyl sulfoxide solution, shaking until completely dissolved to obtain a vanadium acetylacetonate solution. For example, 0.1 g of vanadium acetylacetonate is dissolved in 8 mL of dimethyl sulfoxide solution, or 0.2 g of vanadium acetylacetonate is dissolved in 10 mL of dimethyl sulfoxide solution, or 0.3 g of vanadium acetylacetonate is dissolved in 7 mL of dimethyl sulfoxide solution, or 0.25 g of vanadium acetylacetonate is dissolved in 5 mL of dimethyl sulfoxide solution.

[0063] Take 20–50 μL of vanadium acetylacetonate solution and evenly coat it onto the BiOI film (e.g., 20 μL, 25 μL, 30 μL, 40 μL, 45 μL, or 50 μL of vanadium acetylacetonate solution). Place it in a muffle furnace and hold at 400–500 °C for 30–60 min. Set the heating rate of the muffle furnace to 1–5 °C / min. For example, the heating rate of the muffle furnace can be 1 °C / min, 2 °C / min, 3 °C / min, 3.5 °C / min, 4 °C / min, 4.5 °C / min, or 5 °C / min.

[0064] S1.5 After heat preservation in a muffle furnace, the surface of the conductive substrate 1 is cleaned with a 1-2M NaOH solution to obtain a conductive substrate with a BVO thin film layer 21.

[0065] S2. Alternately deposit TiO2 thin films and CoO2 thin films on the side of BVO thin film layer 21 away from conductive substrate 1. XA thin film is formed, comprising a Co-doped TiO2 quantum layer thin film 22. Where x ≤ 3, the Co-to-Ti doping mass ratio is 2–8%. For example, the Co-to-Ti doping mass ratio of 2–8% can be 2%, 3%, 4%, 5%, 6%, 7%, or 8%.

[0066] In one embodiment of this disclosure, an atomic layer deposition (ALD) technique is used to deposit a Co-doped TiO2 (Co:TiO2) quantum layer thin film on the surface of the BVO thin film layer 21. Exemplarily, firstly, the ALD material growth chamber is evacuated to a vacuum state, ensuring the pressure inside the chamber is ≤20 Pa. The temperature of the bottom tray inside the chamber is set to 250–300 °C, and the temperature inside the chamber is set to 200–250 °C. After the bottom tray reaches the set temperature, the chamber is allowed to cool for at least 1–2 hours to ensure uniform temperature distribution. For example, the pressure inside the material growth chamber can be ≤20 Pa, ≤15 Pa, ≤10 Pa, or ≤5 Pa; the temperature inside the material growth chamber can be set to 200 °C, 210 °C, 220 °C, 225 °C, 230 °C, 240 °C, or 250 °C.

[0067] Secondly, set appropriate parameters to alternately deposit TiO2 and CoO films. X Thin film, by controlling TiO2 thin film and CoO X The thickness of the TiO2 film and the Co doping ratio are adjusted by the number of film cycles. Furthermore, the deposition thickness of the Co:TiO2 film is 5 nm. Furthermore, the Co-to-Ti doping mass ratio is 5%.

[0068] S3. Obtain an Rh-doped STO precursor solution. Coat the Co-doped TiO2 quantum layer 22 with the Rh-doped STO precursor solution on the side away from the conductive substrate 1. Hold the solution at 450–500°C under inert gas protection for 30–60 min to form an Rh-doped STO quantum layer 23 on the Co-doped TiO2 quantum layer 22. In other words, after coating the Co-doped TiO2 quantum layer 22 with the Rh-doped STO precursor solution, it needs to be calcined in an inert atmosphere to obtain the Rh-doped STO quantum layer 23. For example, the calcination temperature and time can be 450°C for 40 min, 500°C for 30 min, 480°C for 50 min, 460°C for 60 min, 470°C for 45 min, or 490°C for 55 min. In one example, the Rh doping mass ratio to Ti is 0.5–2%. For example, the Rh doping mass ratio relative to Ti can be 0.5%, 0.8%, 1%, 1.2%, 1.5%, or 2%. Further, the Rh doping mass ratio relative to Ti is 1%.

[0069] Rh-doped STO quantum layer thin film 23 was obtained by sintering under an inert atmosphere. Due to the incomplete combustion of organic matter, a network-like organic carbon layer is formed on the surface of the semiconductor material. The organic carbon layer has good adsorption properties for probes and can effectively fix biological probes. Thus, probe fixation can be achieved without the need for additional modification of two-dimensional materials or probe adsorption layers, thereby reducing the complexity of sensor fabrication and avoiding the introduction of additional uncontrollable factors.

[0070] S4. On the side of the Rh-doped STO quantum layer 23 away from the conductive substrate 1, the aptamer probe solution is uniformly sprayed using a microelectronic sprayer. After spraying the probe, the layer is placed in a constant temperature incubator at 37°C for 20 minutes, followed by rinsing with phosphate buffer. After rinsing, non-specific binding sites are blocked with a protein-free blocking solution, and then the layer is washed and dried again to obtain the biosensor.

[0071] In one example, Rh-doped STO quantum layer thin films 23 with Rh doping mass ratios of 0%, 0.5%, 1.0%, and 1.5% relative to Ti were prepared by the sol-gel method.

[0072] Preparation of the precursor solution of undoped Rh STO (Rh doping ratio of 0): Add 0.85g of tetrabutyl titanate (C 16 H 36 O4Ti) was dissolved in 5 mL of 5% glacial acetic acid solution and stirred for 10–20 min. The resulting solution was then sonicated until it became clear and then allowed to stand until no more bubbles were present to obtain a tetrabutyl titanate solution.

[0073] The tetrabutyl titanate solution was transferred to a micro-volume sample vial and stirred with a magnetic stirrer. During stirring, 2.5 mL of 1M Sr(NO3)2 solution was added dropwise to the tetrabutyl titanate solution at a rate of 0.2–1.0 mL / min using a quantitative syringe pump, followed by standing for 5–10 min. Then, 1.5 mL of 1M citric acid solution was added dropwise at a rate of 0.5 mL / min, and the solution was stirred continuously at room temperature for 30 min. The resulting solution was the precursor solution of STO.

[0074] Preparation of STO precursor solutions with Rh doping mass ratios of 0.5%, 1.0%, and 1.5% relative to Ti: 0.85 g of tetrabutyl titanate was dissolved in glacial acetic acid solution and stirred for 10–20 min. Rhodium nitrate solution was added according to the required Rh doping mass ratio relative to Ti to obtain the corresponding first mixed solutions.

[0075] Sr(NO3)2 solution was added to the first mixed solution and stirred to obtain a second mixed solution; after standing, citric acid solution was added to the second mixed solution to obtain Rh-doped STO precursor solutions with mass doping ratios of 0.5%, 1.0%, and 1.5%, respectively.

[0076] Four precursor solutions with different doping ratios were coated onto Co-doped TiO2 quantum layer thin films 22, and calcined at high temperature under an inert gas to obtain STO / Co:TiO2 / BVO photoanode thin films, 0.5% Rh:STO / Co:TiO2 / BVO photoanode thin films, 1.0% Rh:STO / Co:TiO2 / BVO photoanode thin films, and 1.5% Rh:STO / Co:TiO2 / BVO photoanode thin films. Figure 5 As shown, the performance of the photoanode thin film layer 2 varies with different Rh doping ratios. In this embodiment, the 1.0% Rh:STO / Co:TiO2 / BVO photoanode thin film layer exhibits the best photoelectric performance.

[0077] In one example, the photoelectric properties of BVO photoanode thin film, Rh:STO / BVO photoanode thin film, TiO2 / BVO photoanode thin film, Co:TiO2 / BVO photoanode thin film, and Rh:STO / Co:TiO2 / BVO photoanode thin film were measured respectively. Figure 6 The JV curves shown demonstrate that TiO2 / BVO exhibits superior photoelectric performance compared to pure-phase BVO. Furthermore, Co:TiO2 / BVO also demonstrates better photoelectric performance than TiO2 / BVO, further illustrating the advantages of Co doping. Moreover, compared to simply adding a rare-metal-doped quantum layer, such as Rh:STO / BVO and Co:TiO2 / BVO photoanode films, the Rh:STO / Co:TiO2 / BVO photoanode film with both quantum layers exhibits higher photoelectric performance, indicating a synergistic effect between Rh:STO and Co:TiO2, which further optimizes the photoelectric performance of this photoanode film.

[0078] The following specific embodiments further illustrate the biosensors provided in this disclosure in terms of composition and preparation process.

[0079] Example 1

[0080] Step 1: Pretreatment of conductive substrate. For example, ITO conductive glass (2cm×1cm×0.15cm, sheet resistance ≤10Ω) is immersed in a special ITO cleaning agent, the cleaning solution temperature is maintained at 50℃, and ultrasonication is performed for 30 minutes. After cleaning, it is dried with high-purity N2 gas.

[0081] Step 2: Weigh 2.89g of Bi(NO3)35H2O and dissolve it in 200mL of ultrapure water. Then add 13.28g of KI and stir for 30min. Adjust the pH of the solution to 1.5 with HNO3. The resulting solution is the first solution.

[0082] Weigh 2.92 g of p-benzoquinone powder and dissolve it in 100 mL of ethanol. Sonicate the solution until completely dissolved to obtain the p-benzoquinone solution. Then mix the p-benzoquinone solution with the first solution and stir for 30 min until homogeneous to obtain the second solution.

[0083] Step 3: Using the cleaned ITO conductive glass as the working electrode, a platinum electrode as the counter electrode, and an Ag / AgCl electrode as the reference electrode, a BiOI thin film was prepared by electrodeposition in the second solution. Specifically, deposition was carried out at a constant potential of -0.2V for 30 seconds.

[0084] Step 4: Weigh 0.3g of vanadium acetylacetonate and dissolve it in 5mL of dimethyl sulfoxide solution. Shake until completely dissolved to obtain a vanadium acetylacetonate solution. Drop-coat 20μL of the vanadium acetylacetonate solution onto the BiOI film prepared in Step 3. Place the film in a muffle furnace and hold at 450℃ for 30min. Set the heating rate of the muffle furnace to 2℃ / min. After holding at this temperature, wash with 1M NaOH solution to obtain BVO film layer 21.

[0085] Step 5: On the BVO thin film layer 21 obtained in step 4, a Co-doped TiO2 thin film (Co:TiO2) is deposited by atomic layer deposition (ALD) to obtain a Co-doped TiO2 quantum layer thin film.

[0086] The ALD material growth chamber was evacuated to a vacuum, with the pressure inside the chamber below 10 Pa. The temperature inside the chamber was set to 220°C, and the chamber was allowed to cool for at least 1 hour to ensure uniform temperature.

[0087] The TiO2 and CoOx films were deposited alternately with appropriate parameters, where x ≤ 3. The thickness of TiO2 and the Co doping ratio were adjusted by controlling the number of cycles. Table 1 shows the process parameters for preparing the TiO2 film, where the deposition thickness of the TiO2 film in each cycle is 0.1 nm. Table 2 shows the deposition parameters for the CoOx film, where the deposition thickness of the CoOx film in each cycle is 0.1 nm. X Growth of 0.032nm.

[0088] Table 1. Process parameters for ALD growth of TiO2 thin films

[0089] <![CDATA[TiO2 parameters]]> <![CDATA[H2O source]]> Ti source Pulse time 20ms 100ms Cleaning time 25ms 20s Waiting time 0 0

[0090] Table 2. CoO2 growth of ALD X Thin film process parameters

[0091] <![CDATA[CoO X Parameters Co source <![CDATA[N2]]> <![CDATA[O3]]> Pulse time 10ms 1000ms 500ms Cleaning time 1s 20s 20s Waiting time 0s 5s 5s

[0092] Step 6: Add 0.85g of tetrabutyl titanate (C 16 H 36 O4Ti) was dissolved in 5 mL of 5% glacial acetic acid solution and stirred for 10 min. The resulting solution was then sonicated until clear and allowed to stand until no bubbles remained. Rhodium nitrate solution was added according to the desired Rh to Ti doping mass ratio to obtain the first mixed solution. While stirring, 2.5 mL of 1M Sr(NO3)2 solution was added dropwise to the first mixed solution at a rate of 0.5 mL / min using a metered syringe pump, followed by standing for 5 min. Then, 1.5 mL of 1M citric acid solution was added dropwise at a rate of 0.5 mL / min, and the solution was stirred continuously at room temperature for 30 min. The resulting solution was the Rh-doped STO precursor solution.

[0093] Step 7: The Rh-doped STO precursor solution was coated onto the Co-doped TiO2 quantum layer film 22, and then held at 450℃ under argon protection for 30 min to form the Rh-doped STO quantum layer film, thus obtaining the Rh:STO / Co:TiO2 / BVO photoanode film. The Rh:STO / Co:TiO2 / BVO photoanode film was scanned by transmission electron microscopy (TEM), as shown below. Figure 2 The image shown is a cross-sectional TEM image of the photoanode thin film layer. Figure 3 High-resolution transmission electron microscopy (HRTEM) image of the photoanode thin film layer; Figure 4 This image shows a scanning transmission electron microscope (STEM) image and elemental analysis of the photoanodized thin film layer. The image reveals that quantum layers are uniformly distributed on the upper surface of the BVO thin film layer 21, with a Co:TiO2 thickness of approximately 5 nm and a Rh:STO thickness of approximately 10 nm. Elemental analysis reveals the characteristic elements of the C layer and each thin film layer on the surface.

[0094] Step 8: Uniformly spray 20 μL of a 20 nmol / L DNA aptamer probe solution for detecting COVID-19S protein onto the Rh-doped STO quantum layer film and incubate at 37°C for 20 min. Then, wash the surface of the photoanode film layer 2 with phosphate buffer (pH 7.4) to remove any DNA aptamer probes that are not firmly adsorbed. Next, block the non-specific binding sites with a protein-free blocking solution to form the aptamer probe layer. The base sequence of the DNA aptamer used for detecting COVID-19S protein is: 5′-CAG CAC CGA CCT TGT GCT TTG GGA GTG CTG GTC CAA GGG CGT TAA TGG ACA-3′ (SEQ ID NO:1).

[0095] Taking the specific detection of the COVID-19S protein as an example, analysis showed that Rh:STO quantum layer modification significantly improved the sensor's probe capture capacity. For example... Figure 7 The figures show the measured adsorption capacity of the COVID-19S protein aptamer probe using BVO photoanode film layers, Co:TiO2 / BVO photoanode film layers, and Rh:STO / Co:TiO2 / BVO photoanode film layers, respectively. Figure 7 It can be seen that the addition of the Rh:STO quantum layer significantly improves the adsorption of the probe on the sensor surface.

[0096] like Figure 8 and Figure 9 The sensor with Rh:STO / BVO photoanode thin film layer provides results for the detection of COVID-19S protein. Figure 10 and Figure 11 The results of the sensor with the Rh:STO / Co:TiO2 / BVO photoanode thin film layer for the detection of COVID-19S protein are shown in the figure. As can be seen from the figure, the detection range of the Rh:STO / BVO sensor is 10 ag / mL to 1 ng / mL, and the detection limit is 4.23 ag / mL. The detection range of the Rh:STO / Co:TiO2 / BVO sensor is 10 ag / mL to 1 ng / mL, and the detection limit is 1.99 ag / mL. This indicates that the Co:TiO2 quantum layer can improve the detection sensitivity of the sensor.

[0097] Therefore, this embodiment illustrates that the sensitivity of a sensor can be greatly improved through the synergistic effect of rare metal-doped dual quantum layers (Co-doped TiO2 quantum layer and Rh-doped STO quantum layer).

[0098] Example 2

[0099] Steps 1-7 are the same as in Example 1.

[0100] Step 8: Uniformly spray 20 μL of a 20 nmol / L DNA aptamer probe solution for detecting the HIV-21 gene onto the Rh-doped STO quantum layer film and incubate at 37°C for 20 min. Then, wash the surface of the photoanode film layer 2 with phosphate buffer (pH 7.4) to remove any DNA aptamer probes that are not firmly adsorbed. Next, block the non-specific binding sites with a protein-free blocking solution to form the aptamer probe layer. The HIV-21 gene sequence is: 5′-TGT GTG AGG GCT GGG CCTTGG-3′ (SEQ ID NO:4). The complementary base sequence of the aptamer used for detecting HIV-21 is: 5′-GGG GGG CCAAGG CCC AGC CCT CAC ACA-3′ (SEQ ID NO:2).

[0101] like Figure 12 The image shows the detection results of the biosensor provided in this embodiment for HIV-21. It can be seen that the detection range of the biosensor for HIV-21 is 10 aM to 1 nM, and the detection limit is:

[0102] 3.33aM.

[0103] Example 3

[0104] Steps 1-7 are the same as in Example 1.

[0105] Step 8: Uniformly spray 20 μL of a 20 nmol / L DNA aptamer probe solution for detecting H1N1 influenza A virus onto the Rh-doped STO quantum layer film and incubate at 37°C for 20 min. Then, wash the surface of the photoanode film layer 2 with phosphate buffer (pH 7.4) to remove any loosely adsorbed DNA aptamer probes. Next, block the non-specific binding sites with a protein-free blocking solution to form the aptamer probe layer. The base sequence of the aptamer used to detect the H1N1 influenza A virus gene is: 5′-GATACTGCGTGCTTG TTCCATA-3′ (SEQ ID NO: 3).

[0106] like Figure 13 The results shown are the detection results of the biosensor provided in this embodiment for the H1N1 gene of influenza A virus. It can be seen that the detection range of the biosensor for the H1N1 gene of influenza A virus is 0.1 fg / mL to 10 ng / mL, and the detection limit is 33.3 ag / mL.

[0107] By changing the type of biological probe, the rare metal-doped double quantum layer modified semiconductor photoelectrochemical biosensor disclosed in this disclosure has extremely high detection sensitivity for the novel coronavirus surface spike protein (S protein), HIV-21 gene, and influenza A virus H1N1 gene. It has excellent application breadth and is a universal ultra-sensitive semiconductor photoelectrochemical biosensor.

[0108] It should be noted that although the steps of the fabrication method of the rare metal-doped double quantum layer modified biosensor in this disclosure are described in a specific order in the accompanying drawings, this does not require or imply that these steps must be performed in that specific order, or that all the steps shown must be performed to achieve the desired result. Additional or alternative steps may be omitted, multiple steps may be combined into one step, and / or one step may be broken down into multiple steps.

[0109] Other embodiments of this disclosure will readily occur to those skilled in the art upon consideration of the specification and practice of the invention disclosed herein. This application is intended to cover any variations, uses, or adaptations of this disclosure that follow the general principles of this disclosure and include common knowledge or customary techniques in the art not disclosed herein. The specification and examples are to be considered exemplary only, and the true scope and spirit of this disclosure are indicated by the appended claims.

Claims

1. A method for fabricating a rare metal-doped double quantum layer modified biosensor, characterized in that, Includes the following steps: S1. Deposit a BiVO4 thin film layer on a conductive substrate; S2. Alternately deposit TiO2 and CoO films on the side of the BiVO4 thin film layer away from the conductive substrate. X A thin film is formed to create a Co-doped TiO2 quantum layer, wherein x ≤ 3, and the Co-to-Ti doping mass ratio is 2-8%; wherein the alternating deposition of TiO2 and CoO2 thin films constitutes a Co-doped TiO2 quantum layer. X The thin film was deposited using atomic layer deposition. S3. Obtain an Rh-doped SrTiO3 precursor solution. Coat the side of the Co-doped TiO2 quantum layer film away from the conductive substrate with the Rh-doped SrTiO3 precursor solution. Hold at 450-500°C under inert gas protection for 30-60 minutes to form an Rh-doped SrTiO3 quantum layer film on the Co-doped TiO2 quantum layer film. The doping mass ratio of Rh to Ti is 0.5-2%. S4. Apply aptamer probe solution to the side of the Rh-doped SrTiO3 quantum layer film away from the conductive substrate, and seal the non-specific binding sites with a protein-free blocking solution to form an aptamer probe layer. The rare metal-doped double quantum layer modified biosensor includes a conductive substrate, a photoanode thin film layer, and an aptamer probe layer stacked sequentially. The photoanode thin film layer includes a BiVO4 thin film layer, a Co-doped TiO2 quantum layer thin film layer, and a Rh-doped SrTiO3 quantum layer thin film layer arranged sequentially; the BiVO4 thin film layer is disposed on a conductive substrate, and the Rh-doped SrTiO3 quantum layer thin film layer is disposed on the side of the Co-doped TiO2 quantum layer thin film layer away from the conductive substrate.

2. The method for fabricating a rare metal-doped double quantum layer modified biosensor according to claim 1, characterized in that, The thickness of the Co-doped TiO2 quantum layer thin film is 2~5 nm.

3. The method for fabricating a rare metal-doped double quantum layer modified biosensor according to claim 1, characterized in that, The thickness of the Rh-doped SrTiO3 quantum layer thin film is 10 nm to 15 nm.

4. The method for fabricating a rare metal-doped double quantum layer modified biosensor according to claim 1, characterized in that, The thickness of the BiVO4 thin film layer is 150nm to 200nm.

5. The method for fabricating a rare metal-doped double quantum layer modified biosensor according to any one of claims 1-4, characterized in that, The aptamer probe layer includes the base sequence of a DNA aptamer: 5′-CAGCACCGACCTTGTGCTTTGGGAGTGCTGGTCCAAGGG CGTTAATGGACA-3′, used to detect the COVID-19S protein.

6. The method for fabricating a rare metal-doped double quantum layer modified biosensor according to any one of claims 1-4, characterized in that, The aptamer probe layer includes the base sequence of a DNA aptamer: 5′-GGGGGGCCAAGGCCCAGCCCTCACACA-3′ is used to detect the HIV-21 gene.

7. The method for fabricating a rare metal-doped double quantum layer modified biosensor according to any one of claims 1-4, characterized in that, The aptamer probe layer includes the base sequence of a DNA aptamer: 5′-GATACTGCGTGCTTGTTCCATA-3′, used to detect the H1N1 influenza A virus gene.

8. The method for fabricating a rare metal-doped double quantum layer modified biosensor according to claim 1, characterized in that, In step S3, obtaining the Rh-doped SrTiO3 precursor solution includes: Tetrabutyl titanate was dissolved in glacial acetic acid solution and stirred for 10-20 min. Rhodium nitrate solution was added according to the required Rh to Ti doping mass ratio to obtain the first mixed solution. Add Sr(NO3)2 solution to the first mixed solution and stir to obtain a second mixed solution; after standing, add citric acid solution to the second mixed solution to obtain Rh-doped SrTiO3 precursor solution.

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