Method for detecting miRNA-21 through self-powered dual-mode sensor with dual-circulation signal amplification function

Abstract

The invention relates to a self-energized sensor, in particular to a preparation method of a photocatalytic fuel cell portable self-energized dual-mode sensor for dual-cycle signal amplification and application of the photocatalytic fuel cell portable self-energized dual-mode sensor in detection of miRNA-21. The sensor prepared by the invention is composed of a dual-photoelectrode system (BiVO4 / Bi2S3 / ITO photoanode and HOFs / ITO photocathode), a capacitor and a digital multimeter (DMM). The sensor integrates CHA and chemical oxidation reduction cycle double amplification mechanisms, and the detection sensitivity is remarkably improved through the synergistic amplification effect. According to the sensor, a digital multimeter and a smart phone are adopted as detection devices, and PFC-colorimetric dual-mode detection of miRNA is realized. The dual-mode sensor has excellent analysis performance, provides a novel technical scheme for instant detection of tumor cell miRNA, and shows important clinical application value in the field of early diagnosis of cancers.

Description

Technical Field

[0001] The present invention relates to a self-powered sensor, in particular to a preparation method of a portable self-powered dual-mode sensor of a photocatalytic fuel cell with dual-cycle signal amplification, and application of the sensor in detecting miRNA-21. Background Art

[0002] Self-powered photoelectrochemical (PEC) sensing technology is considered an effective approach to achieving sensor portability, miniaturization, and cost-effectiveness, as it can directly generate response signals (such as open-circuit potential, current, or power density) without relying on an external power source. Among them, self-powered PEC sensors based on photocatalytic fuel cells (PFCs) have become a research hotspot due to their ability to directly convert chemical and light energy into electrical signals, laying the foundation for the development of light-induced, self-driven electrochemical sensing platforms. However, traditional single-signal readout modes are susceptible to interference from coexisting factors such as experimental environment fluctuations and improper operation, and often face challenges in accuracy and reliability in practical applications. Compared with single-mode biosensors, multimodal analytical technologies, by integrating the advantages of different detection modes, can not only effectively reduce false positive / false negative errors but also enable cross-validation of detection results based on different signal output mechanisms. Therefore, dual-mode sensing strategies have attracted widespread attention in recent years. As a classic analytical method, colorimetric detection has significant advantages such as ease of operation and intuitive results. Combining PFC with colorimetry to construct a dual-mode detection strategy can fully utilize the synergistic effects of the two and significantly improve detection performance.

[0003] MicroRNA (miRNA) is a class of non-coding single-stranded RNA that plays a vital role in gene expression and regulation and is often used as an important diagnostic and prognostic biomarker. Among them, miRNA-21 has been found to be abnormally expressed in many types of cancer, including lung cancer, ovarian cancer, and breast cancer. However, due to the short sequence, low abundance, easy degradation, and high sequence similarity between family members of miRNA, it is very challenging to detect low levels of miRNA-21 in actual samples. Therefore, the development of sensitive miRNA-21 detection methods is of great significance for the clinical diagnosis of diseases, especially cancer.

[0004] Among the related patents that have been disclosed for the detection of miRNA-21, CN202010542514.5 is a method for photoelectrochemical detection of miRNA-21 content based on bismuth vanadate, which combines the photoelectric signal excitation of the BiVO4 modified electrode caused by miRNA-21-mediated rolling circle amplification, endonuclease-specific recognition and hydrolysis reaction, and the specific embedding of methylene blue or Nile blue into double-stranded DNA to construct a signal "enhanced" detection. The label-free photoelectrochemical detection method proposed in this invention has the advantages of low cost, simple and flexible operation, small reagent dosage, high sensitivity, and small background signal. It is expected to become one of the effective methods for detecting miRNA-21. However, the sensor still relies on the electrochemical workstation to collect and process the signal data, which makes it difficult to achieve on-site detection, hindering its practical application.

[0005] CN202211500950.1 A method for photoelectrochemical detection of miRNA-21, comprising: mixing locked walker chain functionalized gold nanoparticles (WP), substrate chain functionalized gold nanoparticles (TP), miRNA-21, a divalent water-soluble manganese salt, and water to produce HRP, utilizing HRP to catalyze the reaction of Na2S2O3 and H2O2 to produce H2S, and simultaneously synthesizing a CuO-Au nanocomposite through Cu-MOF and modifying it on an electrode, utilizing H2S to react with the modified CuO-Au nanocomposite on the electrode to generate a CuS / CuO-Au complex, thereby significantly increasing the photocurrent, thereby establishing a linear relationship between the change in photocurrent and the concentration of miRNA-21, and realizing photoelectrochemical detection of miRNA-21. This method is based on the specific recognition of miRNA-21 and the locked chain, designs DNA Walker signal amplification, and utilizes biocatalytic induction of the CuS / CuO-Au semiconductor heterojunction to amplify the signal, thereby realizing highly sensitive and highly selective photoelectrochemical detection of miRNA-21. However, this method uses a single signal (photocurrent signal) as a quantitative measurement standard, which has the problem that the measurement concentration range is narrow and the measurement results need to be cross-validated. Summary of the Invention

[0006] In response to the shortcomings of the existing technology, the present invention constructs a self-powered photocatalytic fuel cell (PFC) and colorimetric analysis dual-mode sensor based on a dual-cycle signal amplification strategy to achieve highly sensitive detection of miRNA-21.

[0007] The present invention provides a method for detecting miRNA-21 using a self-powered dual-mode sensor with dual-cycle signal amplification, comprising the following steps:

[0008] (1) Preparation of BiVO4 / Bi2S3 / ITO photoanode, comprising the following steps:

[0009] (1.1) Stir 0.01-0.1 g of NH₄VO₃ in 10-100 mL of hot water at 40-50°C for 20-30 minutes to form solution A.

[0010] Stir 0.1-0.5 g Bi(NO3)3·5H2O in 10-100 mL ethylene glycol for 20-30 minutes to form solution B;

[0011] Solution A was slowly added to solution B with continued stirring. After obtaining an orange mixture, the mixture was transferred to a polytetrafluoroethylene-lined autoclave and heated at 120-250°C for 9-10 hours to obtain a yellow product. The product was washed several times with deionized water and anhydrous ethanol, respectively, and dried overnight to obtain BiVO4.

[0012] (1.2) 0.05–0.2 g of BiVO₄ and 0.1–0.5 g of thioacetamide (TAA) were mixed in 10–100 mL of ethanol and stirred for 20–30 minutes. The mixture was then transferred to a polytetrafluoroethylene-lined autoclave and heated at 120°C for 7–8 hours to obtain the product. The product was centrifuged, washed several times with water and ethanol, and dried overnight to obtain BiVO₄ / Bi₂S₃.

[0013] (1.3) Dropping BiVO4 / Bi2S3 solution on the surface of ITO electrode to prepare BiVO4 / Bi2S3 / ITO photoanode;

[0014] (2) Preparation of HOFs / ITO photocathode, comprising the following steps:

[0015] (2.1) Dissolve 5–500 mg of 1,3,6,8-tetrakis-(p-aminophenyl)pyrene (PyTTA) in 1–10 mL of N,N-dimethylformamide (DMF). Slowly add the dissolved solution dropwise to 50–150 mL of vigorously stirred water and stir for 3–5 min.

[0016] (2.2) Slowly add 10-50 mL of anhydrous ethanol to the vigorously stirred water in step (2.1) and continue stirring for 3-5 minutes. After the reaction is completed, centrifuge and wash several times with anhydrous ethanol and water, and finally disperse in ethanol to prepare a HOFs solution;

[0017] (2.3) The obtained HOFs solution was dropped onto the surface of the ITO electrode to prepare the HOFs / ITO photocathode;

[0018] (3) Constructing a colorimetric sensing mode to detect miRNA-21 content, including the following steps:

[0019] (3.1) Take 10 to 100 μL of the solution, the concentration is 0.5 to 5 mg mL- 1Dopamine was dropped into a 96-well microplate and incubated at 25-37°C for 25-30 min and dried;

[0020] At the same time, 10 to 100 μL of a 0.5 to 5 μmol L- 1 The amino-modified P1 probe (commercially purchased DNA chain) was added to a 96-well microplate and incubated at 25–37°C for 0.5–1 h. After incubation, the plates were washed with PBS buffer.

[0021] After washing, use a concentration of 1 to 5 mmol L- 1 The cells were blocked with 6-mercapto-1-hexanol (MCH) for 25-30 min and then washed with PBS buffer;

[0022] (3.2) Different concentrations of target miRNA-21 and two hairpin probe chains H1 and H2 of catalytic hairpin assembly (CHA) were mixed at a molar ratio of 1:1 to 1:5 (H1:H2) (10-100 μL, concentration of 0.5-5 μmol L- 1 ) Incubate at 25-37°C for 1.5-2 hours to trigger the CHA reaction. After the reaction, 10-100 μL of CHA product is added to a 96-well microplate, incubated at 25-37°C for 0.5-1 hour, and washed with PBS buffer;

[0023] After washing, add 50-500 μL of 0.5-5 mmol L- 1 L-ascorbic acid-2-phosphate (AAP) is added to the wells and incubated at a constant temperature for 0.5 to 1 hour. The released ascorbic acid (AA) is transferred from the wells to 10 to 500 μL of the mixed solution and incubated at a constant temperature for 10 to 15 minutes. After incubation, the colorimetric detection is performed.

[0024] The mixed solution contains a concentration of 10 to 100 mmol L- 1 Tris(2-carboxyethyl)phosphine (TCEP), concentration of 10 to 500 mmol L- 1 Na2HPO4, concentration is 10~100mmol L- 1 of bathophenanthroline (BPT), 5% to 20% of Triton-100 (TX-100) and 5 to 50 mmol L- 1 of FeCl3;

[0025] (3.3) Using a smartphone, the color signal converter app was used to measure the image to obtain R, G, and B values. A linear relationship was established between the obtained (R+G) / B value and the target concentration to quantitatively detect the miRNA-21 content.

[0026] (4) Constructing a PFC sensing model to detect miRNA-21 content:

[0027] On the basis of step (3), a dual photoelectrode system was used to combine the prepared BiVO4 / Bi2S3 / ITO photoanode and HOFs / ITO photocathode in a single-chamber quartz cell with PBS as the electrolyte. The mixed solution of step (3) was added to the electrolytic cell, and a 500W xenon lamp was used to vertically irradiate the electrodes, thereby constructing a self-powered sensor.

[0028] A capacitor is used to collect the charge generated between two electrodes and illuminated for a few seconds to complete the charging process.

[0029] The switch is turned from the position connected to the photoelectrode to the position connected to the digital multimeter (DMM). The capacitor discharges the collected charge, generating a transient photocurrent. The DMM is used to record the change in the instantaneous photocurrent value. Finally, the concentration of miRNA-21 in the sample to be tested is calculated through the linear equation.

[0030] The sensor prepared by this invention consists of a dual-photoelectrode system (a BiVO4 / Bi2S3 / ITO photoanode and a HOFs / ITO photocathode), a capacitor, and a digital multimeter (DMM). This sensor utilizes a dual-cycle signal amplification mechanism to significantly enhance detection sensitivity. The first cycle of signal amplification stems from the target miRNA-21 triggering a catalytic hairpin assembly (CHA) reaction, which produces a large amount of double-stranded CHA products (H1 / H2-ALP) through a nucleic acid strand displacement reaction. The second cycle of signal amplification stems from the alkaline phosphatase (ALP)-catalyzed hydrolysis of ascorbic acid (AA), which is then regenerated by TCEP reducing dehydroascorbic acid (DHA), forming a sustainable signal amplification cycle.

[0031] The core of the dual-mode detection mechanism of PFC and colorimetric analysis lies in the dual functionalization of AA: in the photoelectrochemical detection mode, AA, as an efficient electron donor, can undergo oxidation reaction with the photogenerated holes of BiVO4 / Bi2S3 / ITO photoanode, significantly promoting the separation efficiency of photogenerated electron-hole pairs, thereby greatly improving the output signal of PFC; at the same time, in the colorimetric detection mode, AA can convert Fe(BPT)3 into 3 + reduced to Fe(BPT)3 2+, triggering a characteristic color reaction of the solution system (from colorless to pink), and the color intensity is quantitatively related to the AA concentration, achieving precise amplification of the colorimetric signal. This collaborative detection mechanism based on AA-triggered dual-signal output not only ensures the comparability and reliability of the test results, but also significantly improves the accuracy of the detection system through mutual verification between signals. This dual-mode biosensor successfully achieved high-performance detection of miRNA-21. Its unique signal amplification strategy and dual-mode verification mechanism provide a new technical approach for the real-time detection of tumor cell miRNA, showing important application value in the field of early cancer diagnosis. BRIEF DESCRIPTION OF THE DRAWINGS

[0032] Figure 1 The UV-visible absorption spectra of miRNA-21 at different concentrations detected using the colorimetric mode in Example 2 are shown. The inset is a photograph of the solution color change corresponding to the change in miRNA-21 concentration.

[0033] Figure 2 OD in Example 2 534 The relationship curve between nm value and miRNA-21 concentration value, the inset is the linear relationship curve between OD value and the logarithm of miRNA-21 concentration;

[0034] Figure 3 is the linear relationship between the image (R+G) / B value and the miRNA-21 concentration in Example 2;

[0035] Figure 4 This is a digital photograph of the current signal output by a digital multimeter when using the PFC sensing mode to detect miRNA-21 content at different concentrations in Example 3;

[0036] Figure 5 Graph showing the relationship between different miRNA-21 concentrations and photocurrent responses in Example 3. The inset shows the linear relationship between photocurrent response and miRNA-21 concentration. DETAILED DESCRIPTION

[0037] The present invention will be further described below with reference to the embodiments, but the present invention is not limited thereto.

[0038] Example 1

[0039] A method for detecting miRNA-21 using a self-powered dual-mode sensor with dual-cycle signal amplification comprises the following steps:

[0040] (1) Preparation of BiVO4 / Bi2S3 / ITO photoanode;

[0041] 0.0701 g of NH4VO3 was stirred in 40 mL of hot water (45°C) for 30 minutes to form solution A;

[0042] 0.1640 g of Bi(NO3)3·5H2O was stirred in 40 mL of ethylene glycol for 30 minutes to form solution B;

[0043] Solution A was slowly added to solution B with continued stirring to obtain an orange mixture, which was then transferred to a polytetrafluoroethylene-lined autoclave and heated at 180°C for 10 hours to obtain a yellow product. The product was washed three times with deionized water and anhydrous ethanol and dried overnight to obtain BiVO4;

[0044] 0.1 g BiVO4 and 0.2 g TAA were mixed in 30 mL ethanol, stirred for 30 min, and then transferred into a polytetrafluoroethylene-lined autoclave and heated at 120 °C for 8 h to obtain the product. The product was centrifuged, washed three times with water and ethanol, respectively, and air-dried at 60 °C overnight to obtain BiVO4 / Bi2S3.

[0045] The BiVO4 / Bi2S3 solution was dropped onto the surface of the ITO electrode to prepare the BiVO4 / Bi2S3 / ITO photoanode;

[0046] (2) Preparation of HOFs / ITO photocathode;

[0047] Take 50 mg of PyTTA and completely dissolve it in 7.5 mL of DMF. The dissolved solution is slowly added dropwise to 100 mL of vigorously stirred water and stirred for 5 min.

[0048] Then, 40 mL of anhydrous ethanol was slowly added to the vigorously stirred water and the stirring was continued for 5 min. After the reaction, the mixture was washed by centrifugation with anhydrous ethanol and water for 6 times, and finally dispersed in 20 mL of ethanol to prepare a HOFs solution.

[0049] The obtained HOFs solution was dropped onto the surface of the ITO electrode to prepare the HOFs / ITO photocathode;

[0050] (3) construct a colorimetric sensing model to detect miRNA-21 content;

[0051] First, take 50 μL, the concentration is 1 mg·mL- 1 Dopamine was dropped into a 96-well microplate, incubated at 37°C for 30 min, and dried;

[0052] At the same time, 20 μL of 1 μmol L- 1 The amino-modified P1 probe (DNA chain purchased from the market) was added to a 96-well microplate and incubated at 37°C for 1 h. After washing with PBS buffer, the probe was diluted with 2 mmol L- 1 The cells were blocked with MCH () for 30 min and washed three times with PBS buffer;

[0053] Different concentrations of target miRNA-21 and CHA probes H1 and H2 were mixed at a molar ratio of 1:1 (20 μL, 1 μmol L- 1 ) were incubated at 37°C for 2 hours. After triggering the CHA reaction, 20 μL of CHA product was added to a 96-well microplate and incubated at 37°C for 1 hour. After washing with PBS buffer, 100 μL of 1.0 mmol L- 1 The AAP was incubated at 37 °C for 1 h, and the released AA was transferred from the wells to 100 μL of a mixed solution (the mixed solution contained 6 μL of a 50 mmol L- 1 of tris(2-carboxyethyl)phosphine, 82 μL, a concentration of 100 mmol L- 1 of Na2HPO4, 4μL, concentration of 10mmol L- 1 of red phenanthroline, 6 μL, 10% Triton-100, 2 μL, 10 mmol L- 1 of FeCl3), incubated at 50°C for 10 min and then detected in colorimetric mode;

[0054] Using a smartphone, the color signal converter app was used to measure the image to obtain the R, G, and B values. The obtained (R+G) / B value was then used to establish a linear relationship with the target concentration to quantitatively detect the miRNA-21 content. (4) A PFC sensing mode was constructed to detect the miRNA-21 content.

[0055] Based on the operation of step (3), a dual photoelectrode system was used to prepare BiVO4 / Bi2S3 / ITO photoanode and HOFs / ITO photocathode in a single-chamber quartz cell with a concentration of 10 mmol L- 1 The mixed solution of step (3) was added into the electrolytic cell, and a 500W xenon lamp was used to vertically irradiate the electrodes, thereby constructing a self-powered sensor;

[0056] A capacitor is used to collect the charge generated between the two electrodes and illuminated for 100 seconds to complete the charging. When the switch is turned from the photoelectrode position to the DMM, the capacitor discharges the collected charge, generating a transient photocurrent. The change in the instantaneous photocurrent value is recorded using the DMM.

[0057] Example 2

[0058] The colorimetric sensing mode constructed in Example 1 was used to detect miRNA-21 at different concentrations. Figure 1 As shown in Figure 2, the absorbance at 534 nm increased with the increase of miRNA-21 concentration. Figure 1As can be seen from left to right in the illustration, the solution color changes from nearly colorless to pink as the concentration of miRNA-21 changes. This is because the increase in miRNA-21 concentration gradually increases the amount of AA produced in the system, and the pink Fe(BPT)3 generated during the chemical redox cycle 2+ The amount of iodine gradually increases, so the absorbance gradually increases, and its linear detection range is 0.01~50nmol L -1 ,like Figure 2 As shown in the inset, the absorbance is linearly related to the logarithm of miRNA-21 concentration within this concentration range.

[0059] The linear equation is I = 0.05545 × log C [miRNA-21] +0.2169(R 2 =0.9943), and the detection limit was 0.0082 nmol L -1 .

[0060] These results demonstrate that the color change caused by miRNA-21 concentration can be visually distinguished. Subsequently, a smartphone was introduced to measure the image using a color signal converter app to obtain R, G, and B values. The resulting (R+G) / B value was then linearly correlated with the target concentration to quantitatively detect miRNA-21.

[0061] like Figure 3 As shown, (R+G) / B and target concentration are in the range of 0.01 to 10 nmol L -1 The regression equation can be expressed as (R+G) / B=0.03231C. [miRNA-21] +2.257(R 2 =0.9929), and the detection limit was 0.0069 nmol L -1 .

[0062] Example 3

[0063] The PFC sensing mode constructed in Example 1 was used to analyze and measure different concentrations of miRNA-21. Figure 4-5 shown.

[0064] like Figure 4 As shown in the figure, the photocurrent signal intensity gradually increases with the increase of miRNA-21 concentration, which can be attributed to the following two aspects:

[0065] (I) As the concentration of miRNA-21 increases, the amount of ALP introduced by CHA gradually increases, thereby increasing the amount of AA enzymatically hydrolyzed by AAP;

[0066] (II) The oxidation product of AA (DHA) can be repeatedly reduced to AA through a TCEP-mediated chemical redox cycle, and AA can enhance the photocurrent signal as an electron donor.

[0067] Figure 5 Figure 2 is the relationship between different miRNA-21 concentrations and photocurrent response. The miRNA-21 concentrations are 0, 0.001, 0.005, 0.01, 0.05, 0.1, 0.5, 1, 5, 10, and 50 nmol L -1 ,from Figure 5 As can be seen from the illustration, the linear range is 0.001 to 50 nmol L -1 , and within this concentration range, the photocurrent signal intensity is linearly related to the logarithm of the miRNA-21 concentration.

[0068] The linear equation is I = 4.289 × log C[miRNA-21] +37.96(R 2 =0.9923), and the detection limit was 0.18 pmol L -1 (wherein the detection limit LOD = 3SD / K, SD is the standard deviation of the photocurrent signal measured 10 times for blank samples, and K is the slope of the linear equation).

[0069] This paper constructs a self-powered biosensor based on a dual-cycle signal amplification strategy. This sensor integrates CHA and chemical redox cycling amplification mechanisms, significantly improving detection sensitivity through a synergistic amplification effect. The sensor utilizes a digital multimeter and a smartphone as detection devices, enabling dual-mode PFC-colorimetric detection of miRNA. This dual-mode sensor exhibits excellent analytical performance, providing a novel technical solution for the immediate detection of miRNA in tumor cells and demonstrating significant clinical application value in the field of early cancer diagnosis.

Claims

1. A method for detecting miRNA-21 using a self-powered dual-mode sensor with dual-cycle signal amplification, comprising the following steps: (1) Preparation of BiVO4 / Bi2S3 / ITO photoanode, comprising the following steps: (1.1) Stir NH4VO3 in hot water to form solution A; Stir Bi(NO3)3·5H2O in ethylene glycol to form solution B; Solution A was slowly added to solution B with continued stirring. After obtaining an orange mixture, it was transferred to a polytetrafluoroethylene-lined autoclave and heated for a period of time to obtain a yellow product. The product was washed several times with deionized water and anhydrous ethanol, respectively, and dried overnight to obtain BiVO4. (1.2) BiVO4 and thioacetamide were mixed in ethanol, stirred, and transferred to a polytetrafluoroethylene-lined autoclave. The mixture was heated to obtain a product. The product was centrifuged, washed several times with water and ethanol, and dried overnight to obtain BiVO4 / Bi2S3. ( 1.3) Dropping BiVO4 / Bi2S3 solution onto the surface of an ITO electrode to prepare a BiVO4 / Bi2S3 / ITO photoanode; (2) Preparation of HOFs / ITO photocathode, comprising the following steps: (2.1) Dissolve PyTTA completely in DMF and slowly add the dissolved solution dropwise to vigorously stirred water with stirring. (2.2) Slowly add anhydrous ethanol to the vigorously stirred water in step (2.1) and continue stirring. After the reaction is completed, centrifuge and wash several times with anhydrous ethanol and water, and finally disperse in ethanol to prepare a HOFs solution; (2.3) The obtained HOFs solution was dropped onto the surface of the ITO electrode to prepare the HOFs / ITO photocathode; (3) Constructing a colorimetric sensing mode to detect miRNA-21 content, including the following steps: (3.1) Dopamine was dripped into a 96-well microplate, incubated at a constant temperature, and then dried; Meanwhile, the amino-modified P1 probe was added to a 96-well microplate and incubated at a constant temperature, and then washed with PBS buffer; After washing, the cells were blocked with 6-mercapto-1-hexanol and then rinsed with PBS buffer; (3.2) Incubate different concentrations of the target miRNA-21 with two CHA hairpin probe chains H1 and H2 at a constant temperature to trigger the CHA reaction. After the reaction, add the CHA product to a 96-well microplate and incubate at a constant temperature. After incubation, wash with PBS buffer; After washing, L-ascorbic acid-2-phosphate is added to the wells and incubated at a constant temperature. After incubation, the released ascorbic acid is transferred from the wells to a mixed solution containing tris(2-carboxyethyl)phosphine, Na2HPO4, bathophenanthroline, 5% to 20% Triton-100 and FeCl3, and incubated at a constant temperature. After incubation, detection is performed in a colorimetric mode; (3.3) Using a smartphone, the color signal converter app was used to measure the image to obtain R, G, and B values. A linear relationship was established between the obtained (R+G) / B value and the target concentration to quantitatively detect the miRNA-21 content. (4) A PFC sensing mode was constructed to detect the miRNA-21 content: On the basis of step (3), a dual photoelectrode system was used to combine the prepared BiVO4 / Bi2S3 / ITO photoanode and HOFs / ITO photocathode in a single-chamber quartz cell with PBS as the electrolyte. The mixed solution of step (3) was added to the electrolytic cell, and a 500W xenon lamp was used to vertically irradiate the electrodes, thereby constructing a self-powered sensor. A capacitor is used to collect the charge generated between two electrodes and illuminated for a few seconds to complete the charging process. The switch is turned from the position connected to the photoelectrode to the position connected to the digital multimeter. The capacitor discharges the collected charge to generate a transient photocurrent. The digital multimeter is used to record the change in the instantaneous photocurrent value. Finally, a linear equation is obtained through the relationship between the concentration of miRNA-21 and the photocurrent value, and the concentration of miRNA-21 in the sample to be tested is calculated.

2. The method for detecting miRNA-21 using a self-powered dual-mode sensor with dual-cycle signal amplification according to claim 1, wherein: In step (1.1), the ratio of NH4VO3 to hot water is 0.01-0.1 g:10-100 mL, the hot water temperature is 40-50°C, and the stirring time is 20-30 minutes; The ratio of Bi(NO3)3·5H2O to ethylene glycol is 0.1-0.5g:10-100mL, and the stirring time is 20-30 minutes; The orange mixture was then transferred to a polytetrafluoroethylene-lined autoclave and heated at 120-250° C. for 9-10 hours to obtain a yellow product.

3. The method for detecting miRNA-21 using a self-powered dual-mode sensor with dual-cycle signal amplification according to claim 1, wherein: In step (1.2), the ratio of BiVO4 to thioacetamide to ethanol is 0.05-0.2 g:0.1-0.5 g:10-100 mL, the stirring time is 20-30 minutes, and the mixture is heated in an autoclave at 120° C. for 7-8 hours to obtain the product.

4. The method for detecting miRNA-21 using a self-powered dual-mode sensor with dual-cycle signal amplification according to claim 1, wherein: In step (2.1), the ratio of PyTTA to DMF is 5-500 mg:1-10 mL, the amount of water added is 50-150 mL, and the stirring time is 3-5 minutes; In step (2.2), the amount of anhydrous ethanol added is 10-50 mL. After adding anhydrous ethanol, stirring is continued for 3-5 minutes.

5. The method for detecting miRNA-21 using a self-powered dual-mode sensor with dual-cycle signal amplification according to claim 1, wherein: The concentration of dopamine in step (3.1) is 0.5-5 mg·mL -1 , the dosage is 10-100 μL, and dopamine is incubated at 25-37°C for 25-30 min; The concentration of P1 probe was 0.5-5 μmol L -1 , use 10-100 μL, incubate at 25-37°C for 0.5-1h; The concentration of 6-mercapto-1-hexanol is 1-5 mmol L -1 The closure time is 25 to 30 minutes.

6. The method for detecting miRNA-21 using a self-powered dual-mode sensor with dual-cycle signal amplification according to claim 1, characterized in that: In step (3.2), the molar ratio of the two hairpin probe chains H1 and H2 of CHA is 1:1 to 1:5; The concentrations of H1 and H2 were both 0.5–5 μmol L -1 , the dosage is 10-100 μL, incubate at 25-37°C for 1.5-2h; The amount of CHA product used was 10–100 μL, and the CHA product was incubated at a constant temperature of 25–37°C for 0.5–1 h; The concentration of L-ascorbic acid-2-phosphate is 0.5-5 mmol L -1 , add 50-500 μL, and incubate at a constant temperature of 35-37°C for 0.5-1h; The mixed solution contains a concentration of 10 to 100 mmol L -1 Tris(2-carboxyethyl)phosphine, concentration of 10 to 500 mmol L -1 Na2HPO4, concentration is 10~100mmol L -1 of red phenanthroline, 5% to 20% Triton-100 and 5 to 50 mmol L -1 The amount of FeCl3 mixed solution used is 10-500 μL.

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