SrTiO3-In2S3 heterojunction photosensitive material and sensor and application thereof

Abstract

The invention discloses a SrTiO3-In2S3 heterojunction photosensitive material and a sensor and application thereof, the photosensitive material adopts a two-step hydrothermal method to successfully construct an II-type heterojunction, and the separation efficiency of photon-generated carriers is remarkably improved, so that the photocurrent response is enhanced; a photosensitive material is modified on the surface of a tin oxyfluoride electrode to form a composite electrode, then the composite electrode and a micro-fluidic chip construct a micro-fluidic photoelectrochemical immunosensor, and a specific antibody is fixed to serve as a biological recognition element. According to the sensor, trace consumption of samples and reagents is realized, and a controllable reaction environment is provided. When the sensor is used for detecting pepsinogen I, the sensor shows a wide linear range, high sensitivity and excellent selectivity and stability, the lower detection limit is as low as 0.1 pg.mL <-1 >, the upper limit can reach 1 mu g.mL <-1 >, and the sensor has a wide application prospect in the field of clinical disease marker detection.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of detection, in particular to a SrTiO3-In2S3 heterojunction photosensitive material, and a sensor prepared from the material and application of the sensor. BACKGROUND

[0002] Pepsinogen (PG) is a kind of cell-secreted substance related to digestion in the stomach, including two isozymes, PG I and PG II. It is mainly secreted by mucous cells, main cells of the corpus and fundus of the stomach. Most of the pepsinogen enters the stomach cavity to be converted into pepsin, thereby completing the digestion function, and only 1% of the pepsinogen penetrates the gastric mucosal capillary into the blood circulation and stably exists. The secretion of pepsinogen is changed when the stomach atrophy occurs. Therefore, the level of PG I in serum can be used as a reliable indicator of gastric mucosal atrophy. When atrophic gastritis occurs, the secretion of PG is also decreased due to the decrease in secretion capacity, so the decrease in serum PG can indicate the progression degree of atrophic gastritis. Pepsinogen I (PG I) is secreted by the main cells and neck mucous cells of the fundus gland. PGI is mainly a pointer reflecting the function of the acid-secreting gland cells, so when the gastric acid secretion is relatively high, PGI will increase; if the gastric acid secretion is reduced or the gastric mucosal gland atrophy occurs, PGI will decrease. When PG I ≤ 70 ng / mL, the gastric mucosal cells are atrophic, and atrophic gastritis is mostly seen, especially attention is paid to high risk factors of atrophic gastritis, intestinal metaplasia, dysplasia, and gastric cancer. When PG I > 240 ng / mL, the gastric mucosa is damaged, and the active phase PGI increases significantly when the gastric mucosa is attacked or damaged. Helicobacter pylori infection, superficial gastritis, erosive gastritis, gastric ulcer, and duodenal ulcer can all cause an increase in serum PG I. Therefore, the determination of the concentration of serum PG I protein in the early stage has obvious help for screening gastric diseases. The content of PG I in the blood is at the nanogram level, so it is of great significance to accurately determine the low concentration of PG I, and a high-sensitivity method is needed for detection. The purpose is to detect serum pepsinogen I (PG I) by using chemiluminescence (CLIA), flow fluorescence luminescence (FFIA), and enzyme-linked immunoassay (ELISA) three different methods.

[0003] The core advantage of chemiluminescence method (CLIA) is high sensitivity, which can detect extremely low concentration of target substances, wide linear range, no need for multiple dilutions of samples, high degree of automation and adaptation to batch detection. Its shortcomings are more prominent. First, the cost is high, the core luminescent reagent is expensive and sensitive to temperature and light, and needs strict cold chain storage and transportation, which is easy to be invalid due to improper storage. At the same time, it depends on special analyzer, and the equipment purchase and maintenance cost makes it difficult for small laboratories to bear. Secondly, the specificity is insufficient, and hemoglobin, bilirubin and other substances in the sample can easily cause non-specific luminescence. Some reaction systems may also produce cross-reactions with similar antigens or antibodies, resulting in false positive results and affecting the accuracy of detection.

[0004] The core advantage of flow fluorescence luminescence method (FFIA) is that it can realize multiple detection, and can analyze multiple target substances in a single sample at the same time. It has high sensitivity, wide linear range and is suitable for multi-index synchronous screening demand. Its shortcomings are first, the cost of reagents and equipment is high, the core reagents such as fluorescent microspheres and specific probes are expensive, and the purchase and maintenance cost of special flow fluorescence detector is high, which is difficult for small institutions to bear. Secondly, the specificity is easy to be disturbed. The fluorescence spectrum of different probes may overlap and cause cross interference. Impurities in the sample may also affect the binding specificity of the microsphere, resulting in deviation of the results. Thirdly, the operation and data interpretation complexity is high, which requires professional personnel to master the skills of sample processing, instrument calibration and multi-index data splitting. Moreover, the reagent stability is poor, and the temperature and humidity requirements for storage environment are strict, which may lead to performance decline due to improper storage.

[0005] The advantage of enzyme-linked immunoassay (ELISA) is simple operation and low cost, which does not require complex and expensive instruments, and the reagent has good stability and is easy to store and transport, which is suitable for primary laboratories and large-scale sample screening. Its shortcomings are more obvious. First, the sensitivity is relatively low, which is difficult to detect extremely low concentration of target substances, and the adaptation to microanalysis demand is insufficient. Secondly, the specificity is easy to be disturbed, and non-specific antibodies and cross-reacting substances in the sample may cause false positive or false negative results. Thirdly, the reaction steps are more and the time is longer, the repeatability is poor when operated manually, and the linear range is narrow, so the high concentration sample needs to be diluted multiple times for accurate detection, which increases the operation complexity and error risk.

[0006] In addition, other detection methods for PG I protein have also been developed in recent years, such as latex-enhanced immunoturbidimetry. The advantages of this method are simple and rapid operation, strong automation adaptability, no need for complex pretreatment, relatively moderate cost, and suitability for routine detection of batch samples. The disadvantages of this method are as follows: first, the sensitivity is lower than that of chemiluminescence and other technologies, and the detection capability for extremely low concentration target substances is limited, which is difficult to meet the demand of microanalysis; second, the specificity is easily interfered, and components such as blood lipids and hemoglobin in the sample may cause non-specific turbidity changes, resulting in result deviation; third, the linear range is narrow, and high concentration samples need to be diluted multiple times, which increases the operation steps and may introduce errors; meanwhile, the stability of reagents is greatly affected by temperature and pH value, and after long-term storage, latex particles are prone to aggregation, affecting the detection accuracy; and finally, the light path precision of the detection instrument is relatively high, and ordinary equipment may not be able to guarantee the repeatability of the results.

[0007] In current biosensor detection technology, optical detection and electrochemical detection are the most widely used methods, while photoelectrochemical detection is relatively less used. Photoelectrochemical (PEC) biosensors have many advantages and are versatile. First, they have excellent detection performance. They convert signals based on the separation and transfer of photo-generated carriers, have extremely low background current, and have significant signal amplification. Their sensitivity is much higher than that of traditional electrochemical sensors, and they can accurately detect picomolar or even femtomolar levels of biological molecules such as nucleic acids, proteins, and antigens, meeting the detection needs of trace biomarkers. Second, they have obvious advantages in detection specificity. By modifying specific biological recognition elements (such as antibodies, nucleic acid probes, and enzymes) on the surface of photoelectric active materials, they can achieve high selectivity in recognizing target analytes. Moreover, they do not require complex labeling steps or only require simple labeling, reducing signal interference and biological activity loss during the labeling process and further improving detection accuracy. Third, they are extremely flexible in application scenarios. Their simple structure and small size make them easy to miniaturize, integrate, and array. In addition, they have low energy consumption and controllable cost. The detection process only requires external light excitation, and the core photoelectric active material has relatively low preparation cost and can be synthesized on a large scale. Moreover, the sensor has good biocompatibility and can maintain stable detection performance in complex biological samples (such as serum, urine, and tissue fluid) without being easily disturbed by the sample matrix. Combined with fast response speed and short detection cycle, it can effectively improve detection efficiency and has strong application value and broad development prospects in various biological detection fields. The advantages of type II heterojunction between strontium titanate (SrTiO3) and indium sulfide (In2S3) are particularly notable. The band structure matching degree of the two is high, the conduction band minimum and valence band maximum of SrTiO3 are higher than those of In2S3, forming a typical type II band arrangement. Under light, photo-generated electrons can transfer from the conduction band of In2S3 to the conduction band of SrTiO3, and photo-generated holes can migrate from the valence band of SrTiO3 to the valence band of In2S3, achieving efficient spatial separation of photo-generated carriers, significantly suppressing the recombination of electron-hole pairs, and significantly improving the separation efficiency and lifetime of carriers. Meanwhile, SrTiO3 has good chemical stability, electron transport performance, and wide bandgap characteristics, while In2S3 has a suitable narrow bandgap width and strong visible light absorption capacity. The combination of the two can widen the light response range of the heterojunction, effectively utilize visible light or even part of near-infrared light, and improve light utilization efficiency. In addition, the interface between the two is tightly bonded, with low lattice mismatch, which can reduce interface defect states and promote the rapid transfer of carriers at the interface. Both have good biocompatibility and environmental friendliness, strong adaptability in photoelectrocatalysis, photosensing, and biological detection, and relatively flexible synthesis process, which can be prepared by various methods such as hydrothermal and sol-gel, making it easy to control the morphology and structure of the heterojunction and further optimize its photoelectric performance.Furthermore, this photocatalyst has not yet been found to be synthesized in type II heterojunctions or applied in sensors.

[0008] The core advantages of microfluidic chips are: extremely low sample and reagent consumption, which can significantly reduce costs and make them suitable for rare samples; efficient mass and heat transfer in microchannels, which can shorten experimental procedures to minutes, and can integrate multiple steps such as pretreatment, reaction, and detection to achieve automated, high-throughput analysis and reduce human error; small and portable size, which facilitates rapid on-site detection, low waste emissions and environmental protection, while precise control of reaction conditions to improve the repeatability and accuracy of results, making them suitable for applications in multiple fields.

[0009] To overcome the shortcomings of existing detection technologies, there is an urgent need to combine microfluidic integration technology with photoelectrochemical (PEC) detection platforms to obtain sensors with good stability, fast response rate, high specificity, small size, low material consumption, and excellent cost performance. Summary of the Invention

[0010] In view of this, one objective of the present invention is to provide a SrTiO3-In2S3 heterojunction photosensitive material; a second objective of the present invention is to provide a microfluidic photoelectrochemical immunosensor; a third objective of the present invention is to provide a method for preparing the aforementioned microfluidic photoelectrochemical immunosensor; a fourth objective of the present invention is to provide a method for detecting pepsinogen I; and a fifth objective of the present invention is to provide the use of the aforementioned SrTiO3-In2S3 heterojunction photosensitive material in the preparation of photoelectrochemical sensors for detecting biomarkers.

[0011] To achieve the above objectives, the present invention provides the following technical solution: 1. A SrTiO3-In2S3 heterojunction photosensitive material, prepared by the following method: Strontium titanate (SrTiO3) nanomaterials were synthesized using a hydrothermal method. In2S3 was synthesized on the surface of the SrTiO3 nanomaterial using a hydrothermal method to form a type II heterojunction. Specifically, SrTiO3 nanopowder, InCl3•4H2O and thiourea were dissolved in water, and the mixture was stirred, subjected to hydrothermal reaction and dried to obtain the SrTiO3-In2S3 heterojunction photosensitive material.

[0012] 2. A microfluidic photoelectrochemical immunosensor, comprising: A microfluidic chip having at least one microfluidic channel; A composite electrode, which is bonded within the microfluidic channel, the composite electrode comprising a tin oxyfluoride electrode and a SrTiO3-In2S3 heterojunction photosensitive material as described in claim 1 modified on its surface. A biometric element, fixed to the surface of the heterojunction photosensitive material of the composite electrode, is used to specifically bind to the target antigen.

[0013] Preferably, the biological recognition element is fixed by the following method: first, fixing a chitosan / glutaraldehyde mixed solution on the surface of the composite electrode to activate groups; then, fixing the monoclonal antibody corresponding to the target antigen on the surface of the activated electrode; finally, blocking the non-specific binding sites using a bovine serum albumin solution.

[0014] Preferably, the spraying quality of the SrTiO3-In2S3 heterojunction photosensitive material on the tin oxide electrode is 10 mg.

[0015] 3. A preparation method of the microfluidic photoelectrochemical immunosensor, comprising the following steps: (1) preparing a microfluidic chip: photoetching a preset microfluidic channel pattern on a silicon wafer; (2) preparing a composite electrode: cleaning a tin oxide electrode, and spraying the SrTiO3-In2S3 heterojunction photosensitive material on the surface of the electrode; (3) bonding: bonding the composite electrode obtained in step (2) with the microfluidic chip obtained in step (1); (4) biological modification: fixing a biological recognition element on the surface of the bonded composite electrode.

[0016] 4. A detection method of pepsinogen I, using the microfluidic photoelectrochemical immunosensor, comprising the following steps: pumping a sample to be detected containing the pepsinogen I antigen into the microfluidic channel of the sensor, so that it is incubated with the biological recognition element on the surface of the electrode at 37°C; after cleaning with a phosphate buffer solution, pumping an ascorbic acid solution as an electrolyte; applying a bias voltage of -0.5~0.5 V, detecting the photocurrent signal under light, and the reduction value of the photocurrent intensity is negatively correlated with the concentration of the pepsinogen I antigen.

[0017] Preferably, the linear range of the detection method is 0.1 pg·mL -1 to 1 μg·mL -1 , and the lower detection limit is 0.1 pg·mL -1 .

[0018] 5. The use of the SrTiO3-In2S3 heterojunction photosensitive material in the preparation of a photoelectrochemical sensor for detecting a biomarker.

[0019] Preferably, the biomarker is pepsinogen I.

[0020] The beneficial effects of the present application are that the present application provides a SrTiO3-In2S3 heterojunction photosensitive material, a two-step synthesis method taking SrTiO3 as a precursor, preparation of a SrTiO3-In2S3 heterojunction material, the synthesis strategy effectively improves the photoelectric conversion efficiency of the system, realizes significant amplification of the sensing signal, and further reduces the detection limit; a new type of immunosensor integrated with a SrTiO3-In2S3 heterojunction photosensitive material microfluidic channel is used for high-sensitivity detection of pepsinogen I (PGI) antigen. By designing a new single-channel microfluidic structure, a precise reaction space is provided for antigen-antibody specific binding, while the sample and reagent injection amount is greatly reduced, the consumable consumption is reduced, and a wider response range (0.1 pg·mL -1 -1 μg·mL -1 ) is exhibited, and has the outstanding characteristics of high selectivity, excellent stability, and low consumable consumption. BRIEF DESCRIPTION OF DRAWINGS

[0021] In order to make the purpose, technical scheme and beneficial effects of the present application clearer, the present application provides the following drawings for illustration: Figure 1 SEM image and element distribution of SrTiO3; Figure 2 SEM image and element distribution of In2S3; Figure 3 SEM image and element distribution of SrTiO3-In2S3 heterojunction; Figure 4 XRD pattern of SrTiO3, In2S3 and SrTiO3-In2S3; Figure 5 Different element XPS pattern of SrTiO3; Figure 6 Different element XPS pattern of In2S3; Figure 7 Different element XPS pattern of SrTiO3-In2S3 heterojunction composite material; Figure 8 UV and model Schottky diagram of SrTiO3 and In2S3; Figure 9 Mechanism of e − / h + transfer in PEC immunosensor; Figure 10 Microfluidic chip sample diagram designed by solidwork2018; Figure 11 Establishment of a photoelectrochemical microfluidic sensor schematic diagram; Figure 12For the difference in photocurrent of different quality of heterojunction materials sprayed; Figure 13 For the impedance curve of different materials; Figure 14 For the I-t graph of the photoelectrode sprayed with different materials under the conditions of 0.5 M ascorbic acid and light; Figure 15 For the change of photocurrent of different concentrations of antigens; Figure 16 For the linear relationship between the change value of photocurrent intensity and the logarithm of PGⅠ concentration; Figure 17 For the IT graph of the sensor testing different samples; Figure 18 For the stability performance test graph of the immunosensor at different time periods; Figure 19 For the IT graph of the immunosensor testing clinical samples. DETAILED DESCRIPTION

[0022] The present application will be further described below in conjunction with the drawings and specific examples, so that those skilled in the art can better understand the present application and implement it, but the examples are not limiting to the present application.

[0023] Example 1, material synthesis Synthesis of strontium titanate (SrTiO3), the specific preparation method is as follows: (1) At room temperature, 26.4 mmol of ethanolamine and 5 mmol of tetrabutyl titanate were mixed in a 50 mL polytetrafluoroethylene liner, and a uniform transparent light yellow liquid was obtained after magnetic stirring for 120 min.

[0024] (2) 18 mL of ultrapure water was added to the above mixed solution, and magnetic stirring was carried out for 10 min.

[0025] (3) 5 mmol of SrO was added and magnetic stirring was carried out for 90 min until the SrO was completely dispersed in the mixed solution, and then appropriate amount of ultrapure water was added to ensure that the final reaction solution had a volume of 20 mL, and the filling amount of the reaction kettle was 40%.

[0026] (4) The polytetrafluoroethylene liner containing the Sr-Ti-O mixed solution was sealed in a stainless steel reaction kettle and moved into a muffle furnace for hydrothermal reaction at 120 ℃ for 36 h, and then naturally cooled to room temperature.

[0027] (5) Take out the inner lining containing SrTiO3 polyhedron, pour off the supernatant, and wash the precipitate by alternately ultrasonic dispersing with anhydrous ethanol and ultrapure water, and then centrifugal separation, repeat three times, and increase 1 vol.% dilute nitric acid solution cleaning once during the process to remove the SrCO3 impurities in the reaction product.

[0028] (6) Place the precipitate obtained by cleaning in a vacuum drying oven at 70 ℃ for 24 h to obtain strontium titanate nanomaterial, abbreviated as STO.

[0029] In2S3 synthesis: 0.05 M of InCl3·4H2O and 0.1 M of CH4N2S were sequentially dissolved in 30 mL of deionized water to obtain a clear solution, which was transferred to a polytetrafluoroethylene sealed autoclave; the autoclave was sealed and reacted in a drying oven at 180 ℃ for 4 hours, and then naturally cooled to room temperature. The sample was taken out, washed with water and anhydrous ethanol for 3 times, and dried at 60 ℃ overnight. The obtained sample was named as In2S3 (IS).

[0030] SrTiO3-In2S3 heterojunction formation: IS was synthesized on the surface of STO to form a type II heterojunction. 0.2 g of SrTiO3 nanometer powder, 0.15 g of InCl3·4H2O and 0.12 g of thiourea were added to 30 mL of water. Dissolve and stir at room temperature for 30 min, transfer to a polytetrafluoroethylene sealed autoclave, and react in a drying oven at 180 ℃ for 4 hours. After cooling to room temperature, the inner lining was taken out and dried in a 60 ℃ drying oven for 12 h to obtain SrTiO3-In2S3 heterojunction.

[0031] In order to understand the structure of single photocatalyst SrTiO3, In2S3 and SrTiO3-In2S3 heterojunction, SEM characterization test was performed. The SEM image and element distribution of SrTiO3 are shown in Figure 1 The results show that the SEM image of SrTiO3 is irregular polyhedron, and the elements of Sr, Ti and O are uniformly distributed, with a total amount of 100%.

[0032] The SEM image of In2S3 is shown in Figure 2 The results show that In2S3 is coexistence of spherical and cubic, and In and S elements are uniformly distributed in spherical In2S3, and S element is less distributed in cubic shape, with a total amount of 100%.

[0033] The SEM image of SrTiO3-In2S3 is shown in Figure 3The results show that SrTiO3-In2S3 exhibits irregular polyhedron coexisting with SrTiO3 spherical or cubic In2S3, and the elements distribution is consistent with the position relationship of the two different materials closely combined, and there is no cross contamination between each other. The total amount of each element in the heterojunction material is 100%.

[0034] The powder XRD spectra of SrTiO3, In2S3 synthesized by one-step hydrothermal method and SrTiO3-In2S3 composite samples synthesized by two-step method are shown in FIG. 2. Figure 4 X-ray diffraction analysis, the characteristic diffraction peaks of SrTiO3 are at 32.4°, 40.4°, 46.7°, 57.9° and 67.8°, each diffraction peak can completely correspond to the standard card (PDF #01-1018), and has good crystallinity. The characteristic diffraction peaks of In2S3 are at 27.7°, 33.5°, 43.8° and 47.9°, which can correspond to the (311), (400), (511) and (440) crystal planes of β-In2S3; each diffraction peak can completely correspond to the standard card (PDF #05-0731), and has good crystallinity. For the SrTiO3-In2S3 sample, the characteristic diffraction peaks of SrTiO3 and In2S3 appear at the same time, indicating that the SrTiO3-In2S3 composite material is successfully prepared.

[0035] SrTiO3 was analyzed. As shown in FIG. 3, Figure 5 three components appear in the left O 1s spectrum, O L (the lattice oxygen) binding energy is about 530 eV, which corresponds to the bridge oxygen (lattice oxygen of Sr-Ti-O in perovskite structure) in the lattice of the sample, which is the normal bonding oxygen in the crystal structure. In the middle Sr 3d spectrum, the Sr 3d orbit splits into 3d5 / 2 and 3d3 / 2 (peak area ratio is about 3:2) due to spin-orbit coupling. The binding energy of the two peaks in the figure corresponds to the +2 valence state of Sr, indicating that Sr exists in a stable +2 valence state in the sample without other valence state hybridization. The right Ti 2p spectrum splits into 2p3 / 2 and 2p1 / 2 (peak area ratio is about 2:1). The main peak in the figure corresponds to the characteristic binding energy of Ti 4+ . This indicates that Ti mainly exists in +4 valence in the sample, and there is also a small amount of Ti 3+ .

[0036] Subsequently, In2S3 was analyzed. As shown in FIG. 4, Figure 6As shown, the left In 3d orbit splits into 3d5 / 2 and 3d3 / 2 (peak area ratio about 3:2) due to spin-orbit coupling. The 3d5 / 2 peak in the figure is the characteristic peak of In, and the binding energy position corresponds to the +3 valence chemical state of In; the simultaneously appearing 3p1 / 2 peak is another orbital characteristic peak of In, further proving the existence of In element in the sample and the consistency of its chemical state, indicating that In mainly exists in the form of +3 valence and is stable without other valence hybridization. The right S 2p orbit splits into 2p3 / 2 and 2p1 / 2 (peak area ratio about 2:1). The main peak 2p3 / 2 corresponds to a chemical state of S, and the characteristic peak of -SH appears in the figure, indicating that there are two chemical environments of S in the sample: one is S bonded with metal, and the other is surface-adsorbed mercapto (-SH). The existence of such multiple chemical states reflects the bonding diversity of S in the material, which may be related to the surface modification or defect structure of the material.

[0037] Finally, the heterojunction material is analyzed by XPS, Sr is only +2 valence, and the chemical state is stable; Ti is mainly +4 valence, accompanied by a small amount of Ti 3+ , which is related to the charge compensation of oxygen vacancies; O exists in the form of lattice oxygen, oxygen vacancies and surface-adsorbed oxygen, indicating that the material has oxygen vacancy defects; In: only +3 valence, and the chemical state is stable; S: mainly exists in the form of chemical state bonded with In, and the chemical state is single. It is proved that the heterojunction composite system of SrTiO3 and In2S3 is successfully constructed.

[0038] Optical absorption performance is a key factor for testing the photocatalytic performance of semiconductors, which can be characterized by UV-visible absorption spectrum (UV), such as Figure 7 As shown, the lauc curve can be obtained according to the Kubelka-Munk function, and the band gap energy (Eg) of SrTiO3 and In2S3 is calculated. The band gap energy can be calculated by formula (1): (αhν) n =A(hν-Eg) (1) Where α is the absorption coefficient, h and v are Planck's constant and light frequency respectively, k is a constant, and the band gap energy can be represented by Eg. For a direct band gap semiconductor, n is 2. The final calculation result shows that the Eg values of IS and ZIS are 2.97 and 2.1 eV respectively. In order to further explore the catalytic mechanism of SrTiO3-In2S3 heterojunction, platinum sheet was used as the counter electrode, Ag / AgCl as the reference electrode, and 0.2M sodium sulfate solution as the electrolyte solution, and Mott-schottky plots test was carried out on the heterojunction. As shown in Figure 8Since the tangent slope is positive, it can be inferred that SrTiO3 and In2S3 are n-type semiconductors. According to equations (2) and (3), the flat band potential of IS and ZIS relative to the Ag / AgCl electrode (vs. Ag / AgCl) is -0.68 and -0.99 eV, respectively, and the flat band potential relative to the normal hydrogen electrode (NHE) is -0.48 and -0.79 eV, respectively.

[0039] E(RHE)=E(Ag / AgCl)+0.0591pH+0.2 (2) E(RHE)=E(NHE)+0.0591pH (3) E g =E VB -E CB (4) According to equation (4), the valence band potential of SrTiO3 and In2S3 is 2.29 eV and 1.11 eV, respectively.

[0040] Under UV irradiation, after the two semiconductor materials SrTiO3 and In2S3 absorb photons with matching energy, electron transition occurs and generates photo-generated electron-hole pairs, i.e. photo-generated electrons jump from the valence band (VB) to the conduction band (CB). According to the calculated data of the valence band top (E_VB) and conduction band bottom (E_CB) energy levels of the two materials, the CB and VB energy levels of In2S3 and SrTiO3 in this study show an increasing distribution characteristic in turn. During the migration process of photo-generated carriers, photo-generated electrons on the conduction band of SrTiO3 transfer to the conduction band of In2S3, while photo-generated holes on the valence band of In2S3 migrate to the valence band of SrTiO3, and finally photo-generated electrons are transported to the fluorine tin oxide (FTO) electrode through the conduction band, forming a detectable photoelectrochemical (PEC) signal. During the incubation of PGⅠ antigen and the sensor, the specific binding of the antigen to the antibody fixed on the electrode surface leads to an increase in steric hindrance effect, which in turn causes a significant decrease in photocurrent intensity. It is worth noting that the thin nanosheet structure effectively shortens the charge diffusion path of photo-generated holes in the water environment, improving the carrier migration efficiency; at the same time, the photo-generated holes generated in SrTiO3 and In2S3 are captured by ascorbic acid (AA) solution, significantly inhibiting the recombination of photo-induced electron-hole pairs (e - / h - ). The above results demonstrate that the SrTiO3-In2S3 composite material forms an n-n type Ⅱ heterojunction structure, which provides favorable conditions for the efficient separation of photo-generated carriers, helping to improve the PEC sensing performance. Figure 9

[0041] Example 2, preparation of microfluidic channel and sensor and performance test ​The microfluidic chip diagram is drawn using solidwork2018 version, as shown in Figure 10 The microfluidic chip is composed of a sample inlet 1, a sample outlet 2 and a reaction chamber 3. The sample to be tested is injected from the left side 1, reacted in the left side reaction chamber, and discharged from the right side sample outlet after the reaction is completed.

[0042] Then according to the microfluidic chip structure, the microfluidic chip is prepared, and a 1cm×1cm silicon wafer is selected. The specific steps are as follows: 1) Cleaning: sequentially clean with acetone, alcohol, deionized water, and dry under nitrogen.

[0043] 2) The pre-designed microfluidic channel pattern is engraved on the silicon wafer. The photoresist used here is used as the patterning of PDMS microfluidic, with a thickness of 0.5mm. Negative photoresist SU82075 is selected, and spin coating method and immersion development method are used. Alkaline aqueous solution is used as the developer of ultraviolet photoresist, leaving the circuit pattern.

[0044] Establishment of microfluidic photoelectrochemical immunosensor: a 3 cm×3 cm fluorine tin oxide (FTO) electrode is used, which is sequentially placed in acetone, ethanol and deionized water, and is cleaned for 5 minutes by ultrasonic cleaning technology to remove impurities on the surface of the electrode. The SrTiO3-In2S3 heterojunction photosensitive material prepared by hydrothermal synthesis is sprayed on the surface of the FTO electrode after photoetching, and then bonded with the microfluidic channel to form a SrTiO3 / In2S3 / FTO composite electrode. The SrTiO3 / In2S3 / FTO photoanode is assembled on the upper chamber of the microfluidic channel, and the biological cathode is placed in the lower chamber to realize the specific recognition of the analyte pepsinogen I (PG I). The specific process is shown in Figure 11 During the detection process, after the biological anode completes the catalytic reaction, phosphate buffer solution (PBS) is pumped into the chip through the sample inlet to clean the biological cathode, and ascorbic acid (AA) solution is pumped in as electrolyte to enhance the light absorption performance of the system. After the upper and lower chambers are filled with electrolyte, the 4200 type semiconductor test analyzer is used to record the electrochemical response signal between the biological cathode and the photoanode. Then, 8 μL of 10 mM / 2 mM chitosan (CTS) / glutaraldehyde (GA) mixed solution is delivered to the STO / IS / FTO electrode area by using a common needleless needle tube, and the electrode surface groups are activated by room temperature reaction for 2 hours. -1The PG I monoclonal antibody was immobilized on the surface of the activated electrode, and incubated at 4°C overnight. The next day, the electrode was washed with 0.1 M PBS to remove unbound antibodies, and 8 μL of 1% bovine serum albumin (BSA) solution was pumped in to block the non-specific binding sites on the electrode surface at 37°C for 2 hours. Then, the unbound biomolecules were eluted again using 0.1 M PBS. Finally, the prepared sensor was stored in a refrigerator at 4°C for standby detection.

[0045] To study the influence of different thicknesses of heterojunction materials on the detection effect. Different thicknesses of heterojunction materials were sprayed on FTO glass. When the material was too thin, the incident light could not be fully absorbed, resulting in insufficient photo-generated carriers. However, when the thickness of the layer was increased, the photo-generated carriers had to diffuse to the electrode interface to be collected. When the diffusion length of the carriers was exceeded, the carriers far from the interface were lost due to recombination. The relationship between the photocurrent and the spraying thickness showed an inverted U-shaped curve. The spraying thickness was tested. Three pieces of FTO glass of the same size and resistance were sprayed with different amounts of heterojunction materials. The spraying amounts were 10 mg, 15 mg, and 20 mg, respectively. As shown in Figure 12 , the photocurrent was the largest when the spraying amount was 10 mg, so the subsequent experiments were all based on a spraying amount of 10 mg.

[0046] Electrochemical impedance spectroscopy (EIS) technology can be used to characterize the dynamic changes in the electrochemical properties of the electrode interface during the preparation of the immunosensor. In the Nyquist plot of EIS, the linear part in the low-frequency region is related to the diffusion process of the electrode interface, while the semicircular part in the high-frequency region corresponds to the electron transfer resistance (R et ), the value of which directly reflects the degree of hindrance to the electron transfer process at the electrode interface, so the changes in R et can be used to monitor the real-time modification process on the electrode surface. In the high-frequency region, Zview3 software was used for fitting. The EIS curve shown in Fig. 13 showed a typical semicircular feature, indicating that the equivalent circuit of the electrode interface of the immunosensor conformed to the parallel circuit model. The semicircle diameter of the SrTiO3-In2S3 modified FTO electrode was the smallest, and the resistance was the smallest; the resistance of In2S3 and the resistance of SrTiO3 increased in turn, indicating that the heterojunction material was successfully constructed.

[0047] SrTiO3, In2S3, and SrTiO3-In2S3 were sprayed on FTO glass and tested for photocurrent. As shown in Figure 14 , the photocurrent of the SrTiO3-In2S3 heterojunction material was the highest, and the photocurrents of In2S3 and SrTiO3 decreased in turn. This laid the foundation for subsequent testing.

[0048] Example 3, detection of immunosensor The prepared sensor was contacted with different concentrations of pepsinogen I (PGI) antigen solution by pumping, and incubated at 37°C for 0.5 hours. The time-current (IT) curve method was used to systematically explore the photoelectric current response of the microfluidic photoelectrochemical immunosensor to different concentrations of PGI antigen in 0.1 M ascorbic acid (AA) electrolyte solution.

[0049] 1) Sensitivity test The PGI antigen was diluted into different gradients (2 μg·mL -1 , 1 μg·mL -1 , 100 ng·mL -1 , 10 ng·mL -1 , 1 ng·mL -1 , 100 pg·mL -1 , 10 pg·mL -1 , 1 pg·mL -1 , 100 fg·mL -1 , 10 fg·mL -1 ) and introduced into the prepared photoelectrochemical (PEC) microfluidic sensor by pumping, and incubated in a 37°C incubator for 30 min. After incubation, the sensor was washed with phosphate buffer solution (PBS), and then the photoelectric current test was carried out with the help of a semiconductor tester, with a bias voltage range of -0.5~0.5 V, each group of experiment was repeated 3 times, and the relevant test data was recorded for subsequent analysis. The results are shown in Figure 15 The results show that the photoelectric current intensity decreases regularly with the increase of PGI antigen concentration. The essence of this phenomenon is that with the increase of PGI concentration, more PGI antigen specifically binds to the antibody (Ab) fixed on the electrode surface, forming an antigen-antibody complex that hinders the photoelectrochemical (PEC) reaction process, thereby causing the photoelectric current signal to weaken.

[0050] In addition, in the range of 0.1 pg·mL -1 -1 μg·mL -1 , a good linear relationship between the change of photoelectric current intensity (I) and the logarithm of PGI concentration was obtained ( Figure 16 ). The fitting curve is y = -0.6345x + 5.8929 (R 2 = 0.989, the detection limit of PGI is 0.1 pg·mL -1 . The PEC immunosensor constructed for PGI detection has better analysis performance compared with other detections.

[0051] 2) Specificity test To evaluate the comprehensive performance of the prepared photoelectrochemical (PEC) biosensor, further tests were conducted to study the specificity and anti-interference. Different antigens (heart failure markers ST2, placental growth factor PLGF, vascular endothelial growth factor VEGF, and des-gamma-carboxy prothrombin DCP) with a concentration of 1 ng·mL -1 were pumped into the prepared PEC microfluidic sensor and incubated in a 37°C constant temperature incubator for 30 min. After washing with PBS, the photocurrent was tested using a semiconductor tester with a bias voltage of -0.5-0.5 V, repeated 3 times, and the data was recorded, as shown in Figure 17 By comparing the photocurrent response signal changes of different samples, the results showed that only in the PLGF single solution and the mixed system containing PLGF, the photocurrent response showed a significant decrease, while other interfering substances and mixed systems without PLGF had no significant effect on the photocurrent signal, indicating that the PEC biosensor had excellent specific recognition ability and good anti-interference performance for PLGF.

[0052] 3) Stability test The prepared PEC microfluidic sensor was tested for photocurrent using a semiconductor tester at different time periods (0 days, 5 days, 9 days, 13 days, and 15 days) with a bias voltage of -0.5-0.5 V, repeated 3 times, and the data was recorded. Optionally, PLGF antigen with a concentration of 1 ng·mL -1 was pumped into the prepared PEC microfluidic sensor and incubated in a 37°C constant temperature incubator for 30 min. After washing with PBS, the photocurrent was tested using a semiconductor tester with alternating light and darkness for 20 times, a cycle of 400 seconds, a bias voltage of -0.5-0.5 V, repeated 3 times, and the data was recorded. The results are shown in Figure 18 The results showed that after the prepared immunosensor was treated with 10 ng·mL -1 antigen and stored at 4°C, the photocurrent response performance was tested at different time points, and the results showed that the sensor could maintain a relatively stable photocurrent output level after 15 days of storage without significant decay, confirming its good storage stability and performance durability.

[0053] 4) Clinical detection test of immunosensor To evaluate the practical application performance of the constructed photoelectrochemical (PEC) immunosensor, the detection of clinical samples was verified. In clinical practice, the evaluation standard for pepsinogen I (PGI) level is: when PGI ≤ 70 ng·mL -1When PGⅠ > 240 ng / mL, it indicates atrophy of gastric mucosal cells, which is commonly seen in atrophic gastritis, and it is necessary to be alert to the risk of high-risk lesions such as atrophic gastritis, intestinal metaplasia, dysplasia, and gastric cancer; -1 At this time, it indicates that there is damage to the gastric mucosa. During the active phase of gastric mucosal attack or damage, PGⅠ levels will significantly increase; while 70 ng / mL -1 <PGⅠ≤240 ng ·mL -1 This indicates a normal state. Since PGⅠ protein is universally expressed in vivo, this experiment did not include negative and positive control samples; only quantitative detection was performed. Six clinical samples were selected (concentrations of 42.64 ng / mL). -1 63.72 ng / mL -1 96.35 ng / mL -1 190.26 ng / mL -1 235.12 ng / mL -1 250.48 ng / mL -1 (Concentration was determined using chemiluminescence immunoassay) After diluting 100 times, the solutions were pumped into the prepared PEC microfluidic sensors and incubated at 37°C for 30 min. After washing with PBS, the photocurrent was measured using a semiconductor analyzer with a bias voltage between -0.5 and 0.5 V. This was repeated three times, and the data were recorded. The results are as follows: Figure 19 As shown in the table, all six samples were effectively detected. Table 1 lists the concentration comparison data between the detected and actual values ​​of PEC. Statistical analysis shows that the relative error of all measurement data was controlled within 10%, confirming the high accuracy of the detection method and indicating that the prepared PEC immunosensor has good potential for clinical application.

[0054] Table 1. Comparison of Clinical Sample Test Data Sample Standard concentration ng · mL -1 ]]> measured concentration ng mL -1 ]] Relative error 1 42.64 42.66 0.05% 2 63.72 62.1 2.54% 3 96.35 97.55 1.24% 4 190.26 189.37 0.47% 5 235.12 240.15 2.14% 6 250.48 263.7 5.28% The method of the present invention was compared with the published PG1 biosensor, and the results are shown in Table 2.

[0055] Table 2 Comparison of the present invention with the published PG1 biosensor Detection method Detection range Detection limit Reference Ultrasensitive fluorescence immunoassay of pepsinogen I based on enzyme-triggered decomposition of AuNCs / MnO 0.05 ng mL -1 - 200 ng mL -1 ]]> 0.013 ng·mL -1 ]] Zhang H, Cai B, Cai F, Lian M, Wang Y. Ultrasensitive fluorescence immunoassay of pepsinogen I based on enzyme-triggered decomposition of AuNCs / MnO. Anal Methods. 2023 Dec 21;16(1):122-127. doi: 10.1039 / d3ay01821k. PMID: 38086630.2 Electrochemical immunoassay for gastric cancer biomarker pepsinogen I detection based on PdAgPt / MoS 0.5 ng mL -1 - 200 ng mL -1 ]]> 0.173 ng mL -1 ]] He C, Qiu Z, Jin F, Weng L, Chen L, Wang L, Jiang S, Shi J. Electrochemical immunoassay for gastric cancer biomarker pepsinogen I detection based on PdAgPt / MoS. Biomed Mater. 2025 Jan 10;20(2). doi:10.1088 / 1748-605X / ad9fc7. PMID: 39681086.2 Microfluidic photoelectrochemical immunosensor based on SrTiO3-In2S3 heterojunction material 0.1 pg-mL -1 -1 ug-mL -1 ]]> 0.0915 pg-mL -1 ]] The present invention In summary, the application realizes the integration of photoelectrode and microfluidic system by using tin oxyfluoride (FTO) as electrode substrate, selecting SrTiO3 and In2S3 as photosensitive components, preparing SrTiO3-In2S3 heterojunction material by improved hydrothermal synthesis method, and spraying it on the photoanode area of the new single-channel microfluidic chip. The system uses SrTiO3-In2S3 heterojunction nanomaterial to functionally modify the photoanode, and the target analyte pepsinogen I (PGI) is fixed on the surface of the biological cathode. In the detection process, only the enzymatic biological cathode and the sample are incubated to catalyze the target reaction, and the photoanode does not directly contact the sample matrix, effectively avoiding the non-specific interference of the electron donor in the actual sample, significantly improving the stability and anti-interference ability of the sensor. Research has confirmed that the PEC detection platform constructed has excellent characteristics in sensitivity, selectivity, stability and detection linear range, and other key sensor performance indicators. So far, there has been no related report on the application of such technology in pepsinogen antigen detection. Therefore, a new PGI photoelectrochemical biosensor based on SrTiO3-In2S3 heterojunction nanophotocatalyst and PEC platform is successfully prepared in this paper, which has the outstanding advantages of good stability, fast response rate, strong specificity, small size, less consumption of materials and economic efficiency, providing a new technical solution for the detection of trace biomarkers.

[0056] The above-mentioned embodiments are only preferred embodiments of the present application, and the protection scope of the present application is not limited thereto. Any equivalent replacement or transformation of the present application based on the present application is within the protection scope of the present application. The protection scope of the present application is subject to the claims.

Claims

1. A SrTiO3-In2S3 heterojunction photosensitive material, characterized in that, The SrTiO3-In2S3 heterojunction photoactive material is prepared by the following method: SrTiO3 nanomaterial is synthesized by a hydrothermal method; In2S3 is synthesized on the surface of the SrTiO3 nanomaterial by a hydrothermal method to form a type II heterojunction, and the SrTiO3-In2S3 heterojunction photoactive material is obtained after stirring, hydrothermal reaction and drying.

2. A microfluidic photoelectrochemical immunosensor, characterized in that, It comprises: a microfluidic chip having at least one microfluidic channel; a composite electrode bonded in the microfluidic channel, the composite electrode comprising a tin oxyfluoride electrode and the SrTiO3-In2S3 heterojunction photoactive material of claim 1 modified on the surface of the electrode; a biological recognition element fixed on the surface of the heterojunction photoactive material of the composite electrode for specific binding of target antigens. 3.The microfluidic photoelectrochemical immunosensor according to claim 2, wherein, The biological recognition element is fixed by first activating groups on the surface of the composite electrode with a chitosan / glutaraldehyde mixed solution, then fixing monoclonal antibodies corresponding to target antigens on the surface of the activated electrode, and finally blocking non-specific binding sites with a bovine serum albumin solution.

4. The microfluidic photoelectrochemical immunosensor according to claim 2 or 3, wherein, The SrTiO3-In2S3 heterojunction photoactive material is sprayed on the tin oxyfluoride electrode at a mass of 10 mg.

5. A method for preparing a microfluidic photoelectrochemical immunosensor according to any one of claims 2 to 4, characterized in that, It comprises the following steps: (1) preparing a microfluidic chip: photoetching a preset microfluidic channel pattern on a silicon wafer; (2) preparing a composite electrode: cleaning a tin oxyfluoride electrode and spraying the SrTiO3-In2S3 heterojunction photoactive material of claim 1 on the surface of the electrode; (3) bonding: bonding the composite electrode obtained in step (2) with the microfluidic chip obtained in step (1); (4) biological modification: fixing a biological recognition element on the surface of the bonded composite electrode.

6. A method for detecting pepsinogen I, characterized by, The microfluidic photoelectrochemical immunosensor of any one of claims 2-4 is used, comprising the following steps: a sample to be tested containing pepsinogen I antigen is pumped into the microfluidic channel of the sensor and incubated with the biological recognition element on the surface of the electrode at 37°C; after washing with a phosphate buffer, an ascorbic acid solution is pumped in as an electrolyte; a bias voltage of -0.5-0.5 V is applied, and the photocurrent signal is detected under light, and the decrease in photocurrent intensity is negatively correlated with the concentration of pepsinogen I antigen.

7. The method of claim 6, wherein: The linear range of the detection method is 0.1 pg mL -1 to 1 pg mL -1 The lower limit of detection is 0.1 pg mL -1 The limit of detection (LOD) is 0.0915 pg mL -1 (S / N = 3).

8. Use of the SrTiO3-In2S3 heterojunction photoactive material of claim 1 in the preparation of a photoelectrochemical sensor for detecting biomarkers.

9. Use according to claim 8, characterized in that, The biomarker is pepsinogen I.

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