Dual-mode aptamer sensor for detecting AFB1, preparation method of dual-mode aptamer sensor and detection method of AFB1

Abstract

The invention relates to the technical field of aptamer sensors, in particular to a dual-mode aptamer sensor for detecting AFB1, a preparation method of the dual-mode aptamer sensor and a detection method of the AFB1. An MXene (at) Thi / PtAuNPs composite material, hDNA, MCH and Fc-Apt are sequentially assembled on a glassy carbon electrode, so that the dual-mode aptamer sensor disclosed by the invention is constructed; in the ratio electrochemical detection mode, the concentration of AFB1 is detected by using the value of a ratio signal ITHi / IFc, the linear detection range is 10 pg / mL to 30 ng / mL, and the detection limit is 6.2 pg / mL; in the impedance detection mode, the concentration of AFB1 is detected by using the value of Ret, the linear detection range is 3 pg / mL-30 ng / mL, and the detection limit is 0.8 pg / mL; detection results under the two modes can be mutually verified, and the method has the advantages of high sensitivity, good selectivity, high precision, high reliability and the like.

Description

Technical Field

[0001] The present invention relates to the technical field of aptamer sensors, and in particular to a dual-mode aptamer sensor for detecting AFB1 and a preparation method thereof, and a method for detecting AFB1. Background Art

[0002] Aflatoxin B1 (AFB1) is a toxic metabolite produced by fungi under high temperature and high humidity conditions. It is the most widely distributed and most toxic mycotoxin among known agricultural product contaminants. AFB1 is one of the strongest known natural carcinogens and is classified as a Class I carcinogen by the International Agency for Research on Cancer (IARC). AFB1 is widely present in moldy agricultural products, such as corn, soybeans, peanuts, rice, wheat and other grains and nuts. AFB1 may contaminate agricultural products during the growth, harvesting, storage, transportation and processing of crops, leading to the accumulation of AFB1, and may enter milk or dairy products through feed, seriously threatening human health and causing environmental problems. Therefore, the development of sensitive, rapid and reliable AFB1 detection methods for AFB1 is of great significance to public health, food safety and environmental control.

[0003] So far, a variety of analytical methods have been developed for the detection of AFB1, such as colorimetry, fluorescence, electrochemistry, photoelectrochemistry, and electrochemiluminescence, etc. However, these single-mode analytical methods mainly rely on a single-mode output signal to achieve quantitative detection, which is easily affected by interferences in the sample, different operators and instruments, non-standard analytical processes, and other factors.

[0004] In recent years, dual-mode detection technology has been developed and applied to build high-performance sensors. These dual-mode sensors detect targets by combining different response signals generated by different analytical methods. They can not only combine the advantages of the two analytical methods to make the detection methods more flexible, but also verify each other and improve the reliability of the detection results. On the one hand, the ratio electrochemical sensing method can output two electrochemical signals at the same time and use their ratio to achieve quantitative detection. Compared with single-signal sensing, the ratio electrochemical method can effectively eliminate the signal fluctuations caused by external experimental conditions through internal self-reference capability, so that the sensor has stronger anti-interference ability and can provide more accurate detection results. On the other hand, electrochemical impedance spectroscopy (EIS) technology, as an advanced electrochemical analysis method, uses a small-amplitude sinusoidal potential perturbation signal to excite the electrochemical system and measures the resulting impedance response to study and analyze the changes in the interface properties of the electrode surface after a biorecognition event occurs. The impedance analysis method is based on a small-amplitude perturbation of the steady-state system, ensuring the non-destructiveness and high precision of the test. The ratio electrochemical / impedance dual-mode sensor that combines the above two analytical techniques can use ratio electrochemical and impedance methods to detect the target at the same time. It has the advantages of fast response speed, high sensitivity, high accuracy, and high reliability, and has potential application prospects in fungal toxin detection. Summary of the invention

[0005] In order to solve the above technical problems, a dual-mode aptamer sensor for detecting AFB1 and a preparation method thereof, as well as a method for detecting AFB1 are provided. The present invention uses a composite material of MXene@thiophene / platinum and gold nanoparticles (MXene@Thi / PtAuNPs), hairpin DNA (hDNA), mercaptohexanol (MCH) and ferrocene-labeled aflatoxin B1 aptamer (Fc-Apt) to be sequentially assembled on a glassy carbon electrode to construct a ratio electrochemical / impedance dual-mode aptamer sensor to achieve sensitive, accurate and reliable detection of aflatoxin B1.

[0006] In order to achieve the above objectives, the present invention is implemented by the following technical solutions:

[0007] The first aspect of the present invention provides a method for preparing a dual-mode aptamer sensor for detecting AFB1, comprising the following steps:

[0008] Ⅰ. Prepare a MXene@Thi / PtAuNPs composite material solution, drop it onto the pretreated glassy carbon electrode GCE, and dry it to obtain the first intermediate electrode;

[0009] II. Modifying the surface of the first intermediate electrode with a hairpin DNA solution, wherein the hairpin DNA is denoted as hDNA, and fixing the hDNA on the electrode surface by gold-sulfur bonds at room temperature to form a second intermediate electrode;

[0010] III. modifying the surface of the second intermediate electrode with a mercaptohexanol (MCH) solution, and incubating at room temperature to block the non-specific binding sites of gold, thereby forming a third intermediate electrode;

[0011] IV. Modify the surface of the third intermediate electrode with a ferrocene-labeled aflatoxin B1 aptamer solution, wherein the ferrocene-labeled aflatoxin B1 aptamer is denoted as Fc-Apt, and react at 35 to 40°C. Utilize its hybridization complementarity with hDNA to assemble Fc-Apt on the electrode to obtain a dual-mode aptamer sensor for detecting AFB1. The sensor is denoted as GCE / MXene@Thi / PtAuNPs / hDNA / MCH / Fc-Apt according to the material assembly order.

[0012] Furthermore, the method for obtaining the MXene@Thi / PtAuNPs composite material solution is:

[0013] (1) Dispersing 2 to 20 mg of MXene in 2 to 20 mL of water and stirring at room temperature for 0.2 to 1 hour to obtain solution A;

[0014] (2) adding 0.02-0.2 mL of 0.1 M H2PtCl6 solution to solution A, stirring at room temperature for 0.2-1 hour, adding 0.06-0.6 mL of formic acid, continuing to stir at room temperature for 6-12 hours, centrifuging and washing three times, and redispersing the precipitate in 2-20 mL of water to obtain solution B;

[0015] (3) adding 0.01 to 0.1 mg of thionine (Thi) to solution B, stirring at room temperature for 6 to 24 hours, centrifuging and washing three times, and then redispersing the precipitate in 2 to 20 mL of pure water to obtain solution C;

[0016] (4) 0.1 to 1 mL of a 5 to 10 nM Au nanoparticle solution was added to solution C, and the mixture was stirred for 0.2 to 1 hour at room temperature to obtain a MXene@Thi / PtAuNPs composite material solution.

[0017] Furthermore, the concentration of the MXene@Thi / PtAuNPs composite material solution is 0.05-5 mg / mL; the concentration of the hairpin DNA solution is 1-8 μM; the concentration of the mercaptohexanol solution is 0.01-10 mM; and the concentration of the ferrocene-labeled aflatoxin B1 aptamer solution is 1-8 μM.

[0018] Furthermore, the volume ratio of the MXene@Thi / PtAuNPs composite material solution, the hairpin DNA solution, the mercaptohexanol solution, and the ferrocene-labeled aflatoxin B1 aptamer solution is 4-8 μL: 4-8 μL: 4-8 μL: 4-8 μL.

[0019] Furthermore, the room temperature placement time in step II is 4 to 12 hours; the room temperature incubation time in step III is 0.5 to 2 hours; the reaction temperature in step IV is 37° C., and the reaction time is 1 to 2 hours.

[0020] The second aspect of the present invention provides a dual-mode aptamer sensor for detecting AFB1 obtained by the above preparation method.

[0021] The third aspect of the present invention provides a method for detecting AFB1, comprising the following steps:

[0022] S1. Prepare a plurality of dual-mode aptamer sensors for detecting AFB1 obtained by the above preparation method, add a series of concentrations of aflatoxin B1 standard solution to the sensor, each sensor corresponds to one concentration, incubate at room temperature and then rinse, use this as the working electrode, and form a three-electrode system with it, a reference electrode and a counter electrode, and perform ratio electrochemical detection and / or impedance detection in the three-electrode system;

[0023] Ratio electrochemical detection: Using phosphate buffer solution as the electrolyte, scanning the electrochemical alternating current voltammetry (ACV) curve, measuring the ratio signal I Thi / I Fc , establish the ratio signal I corresponding to the concentration of aflatoxin B1 Thi / I Fc The standard linear curve is recorded as standard linear curve A; where I Thi is the oxidation current value of thionine, I Fc is the oxidation current value of ferrocene.

[0024] Impedance detection: Using a mixed solution containing K3Fe(CN)6 and K4Fe(CN)6 as the electrolyte, scan the electrochemical impedance spectroscopy (EIS) curve and measure the impedance signal R et , establish the impedance signal R corresponding to the concentration of aflatoxin B1 et The standard linear curve is recorded as standard linear curve B; where R et is the electron transfer resistance of the electrode;

[0025] S2, dripping the sample solution to be tested onto the dual-mode aptamer sensor, incubating at room temperature and then rinsing, and performing ratio electrochemical detection and / or impedance detection in a three-electrode system according to the steps in S1;

[0026] Ratiometric electrochemical detection: The ratio signal I measured by the electrochemical AC voltammetry curve is Thi / I Fc Substituting into the standard linear curve A, the concentration of aflatoxin B1 in the sample to be tested based on ratio electrochemical detection can be obtained;

[0027] Impedance detection: The impedance signal R measured by the electrochemical impedance spectroscopy curve et Substituting it into the standard linear curve B, the concentration of aflatoxin B1 in the sample to be tested based on impedance detection can be obtained.

[0028] Furthermore, a series of concentrations of aflatoxin B1 standard solution ranged from 10 -12 ~10 -8 g / mL;

[0029] The room temperature incubation time is 0.5 to 2 hours;

[0030] The reference electrode is an Ag / AgCl electrode, and the counter electrode is a platinum wire;

[0031] The concentration of the phosphate buffer solution is 0.01 to 0.1 M, and the pH value is 7.0 to 7.4;

[0032] The total concentration of the mixed solution containing K3Fe(CN)6 and K4Fe(CN)6 is 1-10 mM.

[0033] Furthermore, the standard linear curve A:I Thi / I Fc =4.844+0.353LogC AFB1 , R 2 =0.992, the linear range is 10pg / mL~30ng / mL, and the detection limit is 6.2pg / mL; among which C AFB1 Indicates the concentration of aflatoxin B1, in g / mL; I Thi / I Fc It represents the ratio of the oxidation current value of thionine to the oxidation current value of ferrocene;

[0034] The standard linear curve B: R et =66.6-86.7LogC AFB1 , R 2 =0.996, the linear range is 3pg / mL~30ng / mL, and the detection limit is 0.8pg / mL; among which C AFB1 Indicates the concentration of aflatoxin B1 in g / mL.

[0035] Beneficial technical effects:

[0036] The present invention prepares MXene@Thi / PtAuNPs modified glassy carbon electrode, and sequentially assembles hDNA, MCH and Fc-Apt, to form a double-stranded DNA structure based on hairpin DNA on the electrode surface, thereby forming an aptamer sensor GCE / MXene@Thi / PtAuNPs / hDNA / MCH / Fc-Apt. When detecting aflatoxin B1, the double-stranded DNA structure on the aptamer specifically binds to the aptamer to cause Fc-Apt to be peeled off from the electrode surface, resulting in I Fc At the same time, due to steric hindrance, the stripping of Fc-Apt will lead to I Thi Increases, making the electron transfer resistance R et Therefore, by measuring the ratio signal I Thi / I Fc (ratio electrochemical detection mode) and impedance signal R et (Impedance detection mode) can realize dual-mode detection of the target aflatoxin B1.

[0037] The MXene@Thi / PtAuNPs composite material prepared in the present invention can effectively enhance the electron transfer performance of the sensing interface and generate an internal standard reference signal I Thi ; Compared with single-mode sensing, the dual-mode sensor of the present invention can verify the detection results obtained by the two modes, thereby providing more accurate and reliable detection results; The dual-mode aptamer sensor for detecting aflatoxin B1 prepared by the present invention has the characteristics of high sensitivity, good selectivity, high accuracy, high reliability, etc., and has potential application prospects in the field of agricultural product quality detection. BRIEF DESCRIPTION OF THE DRAWINGS

[0038] Figure 1 Schematic diagram of the preparation process and detection process of the dual-mode aptamer sensor of the present invention; Figure A is a schematic diagram of the preparation principle of the MXene@Thi / PtAuNPs composite material; Figure B is a schematic diagram of the preparation principle of the sensor GCE / MXene@Thi / PtAuNPs / hDNA / MCH / Fc-Apt; Figure C is a schematic diagram of the detection process using the sensor;

[0039] Figure 2 The left figure is the ACV response diagram of the dual-mode aptamer sensor of the present invention for detecting different concentrations of AFB1, where the concentration of AFB1 is 1×10 -11 g / mL,3×10 -11 g / mL,1×10 -10 g / mL,3×10 -10 g / mL,1×10 -9 g / mL,3×10 -9 g / mL,1×10-8 g / mL and 3×10 -8 g / mL;

[0040] Figure 2 The right figure is the ratio electrochemical signal I drawn based on the ACV response diagram Thi / I Fc The standard linear curve between and the logarithm of AFB1 concentration;

[0041] Figure 3 The left figure is the EIS response diagram of the dual-mode aptamer sensor of the present invention for detecting different concentrations of AFB1, where the concentration of AFB1 is 3×10 -12 g / mL, 1×10 -11 g / mL, 3×10 -11 g / mL, 1×10 -10 g / mL, 1×10 -9 g / mL, 3×10 -9 g / mL, 1×10 -8 g / mL and 3×10 -8 g / mL;

[0042] Figure 3 The right figure shows the impedance signal R drawn based on the EIS response diagram. et The standard linear curve between the logarithm of AFB1 concentration. DETAILED DESCRIPTION

[0043] The technical solutions in the embodiments of the present invention will be described clearly and completely below in combination with the embodiments of the present invention and the accompanying drawings. Obviously, the described embodiments are only part of the embodiments of the present invention, rather than all of the embodiments. The following description of at least one exemplary embodiment is actually only illustrative and is by no means intended to limit the present invention and its application or use. All other embodiments obtained by ordinary technicians in this field without creative work based on the embodiments of the present invention are within the scope of protection of the present invention.

[0044] Unless otherwise specifically stated, the numerical value set forth in these embodiments does not limit the scope of the present invention. The technology and method known to those of ordinary skill in the relevant art may not be discussed in detail, but in appropriate cases, the technology and method should be considered as a part of the specification. In all examples shown and discussed here, any specific value should be interpreted as being merely exemplary, rather than as a limitation. Therefore, other examples of exemplary embodiments may have different values.

[0045] In addition, it should be noted that the use of terms such as "first" and "second" to represent electrodes is only to facilitate the distinction between the electrodes obtained in each reaction step. Unless otherwise stated, the above terms have no special meaning and therefore cannot be understood as limiting the scope of protection of the present invention.

[0046] The experimental methods in the following examples without specifying specific conditions are usually measured in accordance with national standards; if there is no corresponding national standard, they are carried out in accordance with general standard requirements or general methods.

[0047] The base pair sequence of the ferrocene-labeled aflatoxin B1 aptamer (Fc-Apt) used below is:

[0048] 5′-GTTGGGCACGTGTTGTCTCTCTGTGTCTCGTGCCCTTCGCTAGGCC C-Fc-3′;

[0049] The base pair sequence of the hairpin DNA (hDNA) used below is:

[0050] 5′-SH-(CH2)6-CTGCCTAGCGAAGGGCACGAGACACAGAGAGACAACA CCGCTAGGCA-3′.

[0051] Fc-Apt and hDNA were provided by Sangon Biotechnology Co., Ltd.

[0052] The following electrochemical measurements, alternating current voltammetry (ACV) and electrochemical impedance spectroscopy (EIS), were performed on a CHI660E electrochemical workstation (Shanghai Chenhua).

[0053] Pretreated glassy carbon electrode GCE: A glassy carbon electrode GCE with a diameter of 3 mm was polished with 0.3 μm and 0.05 μm aluminum oxide powders, and ultrasonicated in ethanol and water to remove surface residues, dried at room temperature, and set aside.

[0054] Example 1

[0055] The preparation method of MXene@Thi / PtAuNPs composite material solution comprises the following steps:

[0056] (1) Disperse 5 mg of MXene in 5 mL of water and stir at room temperature for 0.5 h to obtain solution A;

[0057] (2) Add 0.1 mL of 0.1 M H2PtCl6 solution to solution A, stir for 0.5 hour at room temperature, add 0.3 mL of formic acid, continue stirring for 12 hours at room temperature, centrifuge and wash three times, and then redisperse the precipitate in 5 mL of water to obtain solution B;

[0058] (3) 0.05 mg of thionine (Thi) was added to solution B, stirred at room temperature for 18 hours, centrifuged and washed three times, and the precipitate was redispersed in 5 mL of pure water to obtain solution C;

[0059] (4) 0.25 mL of an Au nanoparticle solution with a concentration of 8.8 nM was added to solution C. After stirring for 0.5 h at room temperature, a MXene@Thi / PtAuNPs composite material solution was obtained.

[0060] The preparation principle diagram of MXene@Thi / PtAuNPs composite material in this case is shown in Figure 1 As shown in Figure A.

[0061] Example 2

[0062] The preparation method of the dual-mode aptamer sensor for detecting AFB1 comprises the following steps:

[0063] Ⅰ. Take 6 μL of the MXene@Thi / PtAuNPs composite material solution (concentration 1 mg / mL) prepared in Example 1 and drop it onto the pretreated glassy carbon electrode GCE, and dry it to obtain the first intermediate electrode;

[0064] II. Modify the surface of the first intermediate electrode with 6 μL of 2 μM hairpin DNA solution, where the hairpin DNA is denoted as hDNA, and place it at room temperature for 8 hours. Use gold-sulfur bonds to fix the hDNA on the electrode surface to form a second intermediate electrode;

[0065] III. Modify the surface of the second intermediate electrode with 6 μL of 1 mM mercaptohexanol (MCH) solution, incubate at room temperature for 1 hour to block the non-specific binding sites of gold, and form a third intermediate electrode;

[0066] IV. Modify the surface of the third intermediate electrode with 6 μL of 2 μM ferrocene-labeled aflatoxin B1 aptamer solution, wherein the ferrocene-labeled aflatoxin B1 aptamer is denoted as Fc-Apt, and react at 37°C for 1 hour. Utilize its hybridization complementation with hDNA to assemble Fc-Apt on the electrode to obtain a dual-mode aptamer sensor for detecting AFB1. The sensor is denoted as GCE / MXene@Thi / PtAuNPs / hDNA / MCH / Fc-Apt according to the material assembly order. The preparation principle diagram of the sensor is as shown in the figure. Figure 1 As shown in Figure B.

[0067] Example 3

[0068] The detection method of AFB1 includes the following steps:

[0069] S1. Prepare multiple dual-mode aptamer sensors GCE / MXene@Thi / PtAuNPs / hDNA / MCH / Fc-Apt prepared in Example 2, and drop a series of concentrations of aflatoxin B1 standard solutions onto the sensors. The concentrations of the series of concentrations of aflatoxin B1 standard solutions are: 3×10 -12 g / mL, 1×10 -11 g / mL, 3×10 -11 g / mL, 1×10 -10 g / mL, 1×10 - 9 g / mL, 3×10 -9 g / mL, 1×10 -8 g / mL and 3×10 -8 g / mL, each sensor corresponds to a concentration. After incubation at room temperature for 40 minutes, it was rinsed with 0.1M phosphate buffer solution (pH=7.4) and used as the working electrode. It was combined with the Ag / AgCl (saturated KCl) reference electrode and the platinum wire counter electrode to form a three-electrode system. Ratio electrochemical detection and impedance detection were performed in the three-electrode system. The detection process is as follows: Figure 1 As shown in Figure C;

[0070] Ratio electrochemical detection: Using 0.1 M phosphate buffer solution (pH = 7.4) as the electrolyte, scanning the electrochemical alternating current voltammetry (ACV) curve, the following is obtained: Figure 2 The ACV response diagram corresponding to different concentrations of AFB1 is shown in the left figure. Figure 2 As shown in the left figure, with the increase of AFB1 concentration, I Thi The value of I gradually increases, and Fc The value of gradually decreases, and the oxidation current value of thionine and the oxidation current value of ferrocene are obtained, which are recorded as I Thi and I Fc , with the oxidation current value of thionine I Thi and the oxidation current value of ferrocene I Fc The ratio of is the ordinate, and the aflatoxin B1 concentration corresponding ratio signal I is established. Thi / I Fc The standard linear curve is as follows: Figure 2 As shown in the right figure, the ratio signal I Thi / I Fc Standard linear curve A:I Thi / I Fc =4.844+0.353LogC AFB1 [g / mL](R 2 =0.992), the linear range was 10pg / mL to 30ng / mL, and the detection limit was 6.2pg / mL;

[0071] Impedance detection: Using a 5 mM mixed solution of K3Fe(CN)6 and K4Fe(CN)6 as the electrolyte, the electrochemical impedance spectroscopy (EIS) curve was scanned to obtain the following Figure 3 The EIS response diagram corresponding to different concentrations of AFB1 is shown in the left figure. Figure 3 As shown in the left figure, with the increase of AFB1 concentration, R et The value of gradually decreases, and the impedance signal R is obtained et , R et The electron transfer resistance of the electrode is used to establish the impedance signal R corresponding to the concentration of aflatoxin B1. et The standard linear curve is as follows: Figure 3 As shown in the right figure, the impedance signal R et Standard linear curve B: R et =66.6-86.7LogC AFB1 [g / mL](R 2 =0.996), the linear range was 3pg / mL to 30ng / mL, and the detection limit was 0.8pg / mL.

[0072] S2. The dual-mode aptamer sensor GCE / MXene@Thi / PtAuNPs / hDNA / MCH / Fc-Apt prepared in Example 2 was applied to the spiked detection of different concentrations of aflatoxin B1 in actual corn samples. The detection process was as follows:

[0073] First, the corn bought from the farmers' market was ground into powder, and a certain concentration of AFB1 standard solution was added. The mixed solution was extracted, filtered, and diluted to form a sample solution of a certain concentration. The sample solution was dripped onto the sensor GCE / MXene@Thi / PtAuNPs / hDNA / MCH / Fc-Apt, incubated at room temperature for 40 minutes, and then rinsed with 0.1M phosphate buffer solution (pH=7.4). In the three-electrode system, the rinsed electrode was used as the working electrode, the Ag / AgCl (saturated KCl) electrode was used as the reference electrode, and the platinum wire was used as the counter electrode. Ratio electrochemical detection and impedance detection were performed according to the method in step S1, and the detected ratio signal I Thi / I Fc And the impedance signal R et Substituting the standard linear curve A and the standard linear curve B respectively, the concentration information of aflatoxin B1 in corn can be calculated in the ratio electrochemical detection mode and the impedance detection mode respectively. The results are shown in Table 1.

[0074] Table 1 Results of dual-mode aptasensor detection of AFB1 in corn samples

[0075]

[0076] The results in Table 1 show that the constructed sensor can achieve sensitive and accurate dual-mode detection of aflatoxin B1, and the relative standard deviation is controlled within 5%.

[0077] The aptamer sensors constructed using GCE / MXene (6 μL of 1 mg / mL MXene aqueous solution was directly added to the glassy carbon electrode, and the subsequent preparation was the same as in Example 2), GCE / MXene / PtNPs (6 μL of 1 mg / mL solution B in Example 1 was added to the glassy carbon electrode, and the subsequent preparation was the same as in Example 2), GCE / MXene / AuNPs (MXene aqueous solution was added to 0.25 mL of 8.8 nM Au nanoparticle solution to form a mixed solution containing 1 mg / mL MXene, which was then added to the glassy carbon electrode, and the subsequent preparation was the same as in Example 2), and GCE / MXene / PtAuNPs (GCE / MXene / PtAuNPs was obtained by removing step 3 from the operating process of Example 1, and the subsequent preparation was the same as in Example 2) could not form a dual mode.

[0078] The above description is only a preferred specific implementation manner of the present invention, but the protection scope of the present invention is not limited thereto. Any technician familiar with the technical field can make equivalent replacements or changes according to the technical scheme and inventive concept of the present invention within the technical scope disclosed by the present invention, which should be covered by the protection scope of the present invention.

Claims

1. A method for preparing a dual-mode aptamer sensor for detecting AFB1, characterized in that: The steps include: Ⅰ. Prepare a MXene@Thi / PtAuNPs composite material solution, drop it onto the pretreated glassy carbon electrode GCE, and dry it to obtain the first intermediate electrode; II. Modifying the surface of the first intermediate electrode with a hairpin DNA solution, wherein the hairpin DNA is denoted as hDNA, and leaving it at room temperature to form a second intermediate electrode; III. modifying the surface of the second intermediate electrode with a mercaptohexanol solution and incubating at room temperature to form a third intermediate electrode; IV. Modify the surface of the third intermediate electrode with a ferrocene-labeled aflatoxin B1 aptamer solution, wherein the ferrocene-labeled aflatoxin B1 aptamer is denoted as Fc-Apt, and react at 35 to 40° C. to obtain a dual-mode aptamer sensor for detecting AFB1. The sensor is denoted as GCE / MXene@Thi / PtAuNPs / hDNA / MCH / Fc-Apt according to the material assembly order.

2. The method for preparing the dual-mode aptasensor for detecting AFB1 according to claim 1, characterized in that: The method for obtaining the MXene@Thi / PtAuNPs composite material solution is: (1) Dispersing 2 to 20 mg of MXene in 2 to 20 mL of water and stirring at room temperature for 0.2 to 1 hour to obtain solution A; (2) adding 0.02-0.2 mL of 0.1 M H2PtCl6 solution to solution A, stirring at room temperature for 0.2-1 hour, adding 0.06-0.6 mL of formic acid, continuing to stir at room temperature for 6-12 hours, centrifuging and washing three times, and redispersing the precipitate in 2-20 mL of water to obtain solution B; (3) adding 0.01 to 0.1 mg of thionine to solution B, stirring at room temperature for 6 to 24 hours, centrifuging and washing three times, and then redispersing the precipitate in 2 to 20 mL of pure water to obtain solution C; (4) 0.1 to 1 mL of a 5 to 10 nM Au nanoparticle solution was added to solution C, and the mixture was stirred for 0.2 to 1 hour at room temperature to obtain a MXene@Thi / PtAuNPs composite material solution.

3. The method for preparing the dual-mode aptasensor for detecting AFB1 according to claim 2, characterized in that: The concentration of the MXene@Thi / PtAuNPs composite material solution is 0.05-5 mg / mL; the concentration of the hairpin DNA solution is 1-8 μM; the concentration of the mercaptohexanol solution is 0.01-10 mM; and the concentration of the ferrocene-labeled aflatoxin B1 aptamer solution is 1-8 μM.

4. The method for preparing the dual-mode aptasensor for detecting AFB1 according to claim 3, characterized in that: The volume ratio of the MXene@Thi / PtAuNPs composite material solution, the hairpin DNA solution, the mercaptohexanol solution, and the ferrocene-labeled aflatoxin B1 aptamer solution is 4-8 μL: 4-8 μL: 4-8 μL: 4-8 μL.

5. The method for preparing the dual-mode aptasensor for detecting AFB1 according to any one of claims 1 to 4, characterized in that: The room temperature placement time in step II is 4 to 12 hours; the room temperature incubation time in step III is 0.5 to 2 hours; the reaction temperature in step IV is 37° C. and the reaction time is 1 to 2 hours. 6 . A dual-mode aptamer sensor for detecting AFB1 prepared according to the preparation method according to any one of claims 1 to 5 .

7. A method for detecting AFB1, characterized in that: The steps include: S1. Prepare a plurality of dual-mode aptamer sensors for detecting AFB1 prepared by the preparation method according to any one of claims 1 to 5, drop a series of concentrations of aflatoxin B1 standard solution onto the sensor, each sensor corresponding to one concentration, incubate at room temperature and then rinse, use this as the working electrode, form a three-electrode system with it, a reference electrode and a counter electrode, and perform ratio electrochemical detection and / or impedance detection in the three-electrode system; Ratio electrochemical detection: Using phosphate buffer solution as the electrolyte, scanning the electrochemical AC voltammetry curve, measuring the ratio signal I Thi / I Fc , establish the ratio signal I corresponding to the concentration of aflatoxin B1 Thi / I Fc The standard linear curve is recorded as standard linear curve A; where I Thi is the oxidation current value of thionine, I Fc is the oxidation current value of ferrocene. Impedance detection: Using a mixed solution containing K3Fe(CN)6 and K4Fe(CN)6 as the electrolyte, scan the electrochemical impedance spectrum curve and measure the impedance signal R et , establish the impedance signal R corresponding to the concentration of aflatoxin B1 et The standard linear curve is recorded as standard linear curve B; where R et is the electron transfer resistance of the electrode; S2, dripping the sample solution to be tested onto the dual-mode aptamer sensor, incubating at room temperature and then rinsing, and performing ratio electrochemical detection and / or impedance detection in a three-electrode system according to the steps in S1; Ratiometric electrochemical detection: The ratio signal I measured by the electrochemical AC voltammetry curve is Thi / I Fc Substituting into the standard linear curve A, the concentration of aflatoxin B1 in the sample to be tested based on ratio electrochemical detection can be obtained; Impedance detection: The impedance signal R measured by the electrochemical impedance spectroscopy curve et Substituting it into the standard linear curve B, the concentration of aflatoxin B1 in the sample to be tested based on impedance detection can be obtained.

8. The method for detecting AFB1 according to claim 7, characterized in that: A series of concentrations of aflatoxin B1 standard solution ranged from 10 -12 ~10 -8 g / mL; The room temperature incubation time is 0.5 to 2 hours; The reference electrode is an Ag / AgCl electrode, and the counter electrode is a platinum wire; The concentration of the phosphate buffer solution is 0.01 to 0.1 M, and the pH value is 7.0 to 7.4; The total concentration of the mixed solution containing K3Fe(CN)6 and K4Fe(CN)6 is 1-10 mM.

9. The method for detecting AFB1 according to claim 8, characterized in that: The standard linear curve A:I Thi / I Fc =4.844+0.353LogC AFB1 , R 2 =0.992, the linear range is 10pg / mL~30ng / mL, and the detection limit is 6.2pg / mL; among which C AFB1 Indicates the concentration of aflatoxin B1, in g / mL; I Thi / I Fc It represents the ratio of the oxidation current value of thionine to the oxidation current value of ferrocene; The standard linear curve B: R et =66.6-86.7LogC AFB1 , R 2 =0.996, the linear range is 3pg / mL~30ng / mL, and the detection limit is 0.8pg / mL; among which C AFB1 Indicates the concentration of aflatoxin B1 in g / mL.

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