Preparation method of Hg < 2 + > sensor based on graphene-gold nanoparticle composite nanomaterial and Hg < 2 + > detection method

Abstract

The invention discloses a preparation method of an Hg < 2 + > sensor based on a graphene-gold nanoparticle composite nanomaterial and an Hg < 2 + > detection method.The preparation method of the Hg < 2 + > sensor based on the graphene-gold nanoparticle composite nanomaterial comprises the following steps that S1, a substrate silicon wafer is cleaned; s2, growth and transfer of graphene; s3, preparing the graphene-gold nanoparticle composite nano material sensor; according to the Hg < 2 + > sensor based on the graphene-gold nanoparticle composite nanomaterial prepared by the invention, by combining AuNPs with graphene, the problems of irreversible self-aggregation, low colloidal stability and the like when graphene is independently used are effectively solved; the excellent conductivity and catalytic performance of AuNPs and the high specific surface area and good biocompatibility of graphene generate a synergistic effect, and the electrical property and detection sensitivity of the sensor are remarkably improved.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of sensors, and particularly relates to a Hg 2+ sensor preparation method and Hg 2+ detection method based on a graphene-gold nanoparticle composite nanomaterial. BACKGROUND

[0002] Mercury ion (Hg 2+ ) as a highly toxic heavy metal ion has high toxicity, difficult degradability and biological accumulation. Once it enters the environment, it will continuously enrich through the food chain, posing a serious threat to the ecological system and human health. Hg 2+ can damage the nervous system, digestive system, respiratory system and other systems of the human body, and cause various diseases. Therefore, it is of great practical significance to realize high-sensitivity, high-selectivity, rapid and accurate detection of Hg 2+ in the environment.

[0003] At present, the methods for detecting Hg 2+ mainly include atomic absorption spectrometry, atomic fluorescence spectrometry, inductively coupled plasma mass spectrometry and the like. Although these traditional detection methods have high detection accuracy, they usually require complex sample pretreatment processes, expensive instrument equipment and professional operators, and have high detection cost and long time consumption, which is difficult to meet the demand of on-site rapid detection.

[0004] Biosensors have attracted widespread attention in the field of heavy metal ion detection due to their advantages of simple operation, rapid response and low cost. Graphene, as a new type of nanomaterial with excellent electrical properties, large specific surface area and good biocompatibility, is widely used in the construction of biosensors. However, the sensors constructed by using graphene alone have problems such as irreversible self-aggregation, low colloidal stability, poor reliability and repeatability, and non-specific adsorption, which limit the further improvement of their detection performance.

[0005] Gold nanoparticles (AuNPs) have unique surface plasmon resonance effect, excellent electrical conductivity and catalytic performance, and good biocompatibility. The combination of AuNPs and graphene to form a composite nanomaterial can effectively improve the above-mentioned defects of graphene and synergistically improve the sensitivity and stability of the sensor. In addition, Hg 2+ can specifically bind to DNA sequences rich in thymine (T) bases to form a stable "T-Hg 2+ -T" hairpin structure, which provides a molecular recognition basis for constructing a highly selective Hg 2+ biosensor. However, the current Hg 2+ biosensors based on the "T-Hg 2+The optimization of the sensor, the DNA hairpin structure is insufficient, especially by regulating the number and distribution of specific bases in the DNA sequence to optimize the hairpin structure and Hg 2+ The study of binding efficiency is less, resulting in a large space for improving the detection performance of the sensor.

[0006] Therefore, a kind of Hg 2+ Sensor and its preparation method and detection method become the technical problems to be solved in the field of heavy metal ion detection.

[0007] Therefore, a kind of Hg 2+ Sensor preparation method and Hg 2+ Detection method. SUMMARY

[0008] The purpose of the present application is to provide a kind of Hg 2+ Sensor preparation method and Hg 2+ Detection method to solve the problems raised in the above background art.

[0009] To achieve the above purpose, the present application provides the following technical solutions:

[0010] A kind of efficient graphene-carbon nanotube transistor sensor preparation method, comprising the following steps:

[0011] S1, the cleaning of substrate silicon wafer

[0012] S1.1, preparation of Piranha cleaning solution: mix concentrated H2SO4 and H2O2 according to the volume ratio of 7:3, first slowly pour concentrated H2SO4 into the culture dish, then use dropper to slowly add H2O2 along the wall of the culture dish into concentrated H2SO4, gently shake the culture dish to fully mix them;

[0013] S1.2, silicon wafer immersion cleaning: use tweezers to slowly put the cut silicon wafer with the size of 1.5cm×1.5cm into the culture dish containing Piranha cleaning solution, shake the culture dish to ensure that all silicon wafers are completely immersed and not overlapped, adjust the temperature of constant temperature magnetic stirrer to 85-95 DEG C, immerse the silicon wafer at this temperature for 2h, until no bubbles are generated on the surface of the silicon wafer;

[0014] S1.3, silicon wafer cleaning and drying: after the cleaning solution is naturally cooled to room temperature, pour it into the waste liquid barrel, add ultrapure water to the culture dish, repeatedly clean the silicon wafer 3 times, each time for 10min, after cleaning, blow the silicon wafer dry with nitrogen, and seal it in IPC vacuum drying dish for standby;

[0015] S2, Growth and transfer of graphene

[0016] S2.1, Copper foil pretreatment: cut the copper foil slightly smaller than the quartz boat to avoid wrinkles and bending, and then soak the cut copper foil in acetic acid to remove the oxide on the surface of the copper foil and achieve surface flattening;

[0017] S2.2, Chemical vapor deposition method for growing graphene: place the pretreated copper foil into the quartz boat and send it into the furnace tube, turn on the vacuum pump to exhaust most of the air in the furnace to a pressure of 3-4 Pa;

[0018] Open the hydrogen valve and adjust the H2 flow rate to 10 sccm, and at the same time use the programmed temperature rising method to raise the temperature in the furnace to 1000℃; after the temperature reaches 1000℃, preheat for 15 min, then introduce CH4, set the CH4 flow rate to 4 sccm, and simultaneously adjust the H2 flow rate to 8 sccm, maintain this flow rate ratio for 35 min, turn off the CH4, and cool in a 10 sccm hydrogen environment for 2-3 h to obtain a copper foil with grown graphene, take out the copper foil with graphene grown on the surface and put it into a glass container for standby;

[0019] S2.3, Transfer of graphene: cut the copper foil with graphene grown on it into a shape slightly larger than the silicon wafer, drop 0.1-0.15 mL of PMMA glue with a molecular weight of 495000 and a concentration of A6 on the surface of the copper foil, spin coat at a speed of 4000 r / min for 45 s, and then bake at 180℃ for 2 min; coat the graphene silicon wafer with Scotch tape on a glass plate, cut the glue frame according to the standard, and then paste the glue frame to the lower edge of the copper foil, and cut off the excess copper sheet and glue frame;

[0020] Transfer the graphene with the glue frame to the FeCl3 solution containing an appropriate amount of HCl solution, HCl prevents Fe 3+ from hydrolyzing, seal and dissolve for 6 h until the copper foil is completely dissolved, replace it with ultrapure water and soak for more than 1 day, and replace the ultrapure water in time during the process;

[0021] Take out the sample and paste it on a clean silicon wafer, place it in a constant temperature magnetic stirrer at 45℃ to dry the moisture, cut off the glue frame with a blade to leave the transferred graphene, and then put it into an acetone solution for more than 1 day to remove the surface PMMA, and then observe the glue removal effect under a microscope and seal it for standby;

[0022] S3, Preparation of graphene-gold nanoparticle composite nanomaterial sensor

[0023] S3.1, gold nanoparticles modified graphene: 100 μL of gold nanoparticles solution with a concentration of 50 μg / mL is vertically dropped on the transferred graphene surface, and the gold nanoparticles solution is completely dried at room temperature to obtain a graphene device with gold nanoparticles attached;

[0024] S3.2, reactive ion etching: the electron tube is preheated for 5 min, the nitrogen cylinder is opened to fill nitrogen for 20 s, the gold nanoparticle-attached graphene device is placed, and the nitrogen valve is closed; after completing the gas path cleaning, O2 is introduced, the O2 flow rate is adjusted to stabilize the internal pressure of the reaction chamber, the etching power is set to 120 W, the O2 flow rate is 60 mL / min, the working gas pressure is maintained at 10 Pa, O2 etching is performed for 1 min, and an etched graphene strip is obtained;

[0025] S3.3, electrode evaporation: the mask plate is covered on the etched graphene strip and fixed in position, 80 nm of Cr and 600 nm of Au are evaporated by using a resistance evaporation film machine; the sample is inverted on the card plate, the baffle is twisted and a glass cover is added, the compression cooler and the evaporation machine are turned on, and the evaporation machine is pumped to absolute vacuum before evaporation;

[0026] First, Cr is evaporated, the evaporation power is turned on after switching the metal, the current is set to about 200 A, and evaporation is performed for 8-9 min; after Cr evaporation is completed, the current is adjusted to 0 and Au is switched, and the current is adjusted to 150 A until Au evaporation is completed;

[0027] The reaction program is sequentially closed, the molecular pump reading is decreased to 0, the exhaust valve is opened to break the vacuum, electrode evaporation is completed, and a Hg 2+ sensor based on graphene-gold nanoparticle composite nanomaterial is obtained.

[0028] S3.4, DNA modification: DSAI solution is added to the surface of the above-prepared graphene-gold nanoparticle composite nanomaterial sensor, and the reaction is carried out in the dark for 11 h, then 100 μL of DNA4 solution with a concentration of 40 nM is added, the DNA4 sequence is 5'-CCACCACTTTTTTTTTGGGGTTTTTTTTT-3', and the reaction is carried out for 45 min to complete the DNA modification, and a graphene-gold nanoparticle composite nanomaterial-based Hg 2+ sensor for detecting Hg 2+ is obtained.

[0029] A method for detecting Hg 2+ by using a graphene-gold nanoparticle composite nanomaterial-based Hg 2+ sensor, comprising the following steps:

[0030] T1, sensor initial state detection: the gate voltage of the semiconductor parameter instrument is set to -8-8V, the source-drain voltage is set to -50mV, the prepared Hg 2+ sensor based on graphene-gold nanoparticle composite nanomaterial is detected, and the initial current value I0 is recorded;

[0031] T2, sample detection: different concentrations of Hg 2+ samples are added dropwise on the surface of the sensor, the reaction is carried out at room temperature for 1h, the gate voltage and the source-drain voltage of the semiconductor parameter instrument are kept unchanged, the current value I of the sensor is detected again, and the current change amount I0-I is recorded;

[0032] T3, result analysis: according to the linear relationship between the current change amount I0-I and the concentration of Hg 2+ , the concentration of Hg 2+ in the sample solution is calculated combined with the detected current change amount; wherein when the concentration of Hg 2+ is in the range of 0.05-1nM, the linear equation of the current change amount and the concentration of Hg 2+ is y=0.0425x+0.39475, R 2 =0.99902, and the detection limit is 0.31pM;

[0033] T4, actual water sample pretreatment: for the actual water sample of the Yangtze River, the water sample is filtered by using a water system filter with a pore size of 0.22um, impurity particles in the water sample are removed, and then Hg 2+ detection is carried out according to the above steps T2 and T3.

[0034] Compared with the prior art, the present application has the following beneficial effects:

[0035] 1, the Hg 2+ sensor based on graphene-gold nanoparticle composite nanomaterial prepared by the present application effectively solves the problems of irreversible self-aggregation and low colloidal stability of graphene when used alone, the excellent conductivity and catalytic performance of AuNPs and the high specific surface area and good biocompatibility of graphene produce a synergistic effect, which significantly improves the electrical performance and detection sensitivity of the sensor; experiments show that the conductivity of the sensor of the present application is improved by about 10 times compared with the graphene sensor without AuNPs modification;

[0036] 2, the present application inserts 4 G bases as a bridging segment between the arms of the DNA hairpin structure rich in multiple 'T' bases (i.e. DNA4 sequence is used), accurately regulates the number of reactive base sites of the arms of the hairpin structure, optimizes the binding efficiency of the DNA hairpin structure and Hg 2+ , so that the sensor has high sensitivity to Hg 2+The detection limit is as low as 0.31 pM, which is much better than the existing part Hg 2+ The sensor has extremely high detection sensitivity.

[0037] 3、The sensor has excellent selectivity to Hg²⁺. 2+ , K + , Fe 2+ , Mg 2+ , Na + , Pb 2+ , Ca 2+ , Mn 2+ , Zn 2+ and other metal ions, only Hg 2+ produces a significant current response, which can accurately identify Hg 2+ and effectively avoid the interference of other ions.

[0038] 4、The Hg 2+ detection method is simple to operate, does not need a complex sample pretreatment process (only needs simple filtration for actual water samples), has short detection time, fast response, and low detection cost, and can be suitable for on-site rapid detection; meanwhile, the method shows good accuracy and reliability in actual water sample (such as Yangtze River water) detection.

[0039] 5、In the sensor preparation process, by optimizing the AuNPs concentration (selecting 50 μg / mL), the reaction time of DSAI and the sensor (selecting 11 h), the reaction time of DSAI and DNA (selecting 45 min) and other experimental parameters, the performance stability and detection accuracy of the sensor are further ensured, which provides a strong guarantee for the large-scale production and practical application of the sensor. DETAILED DESCRIPTION

[0040] The accompanying drawings are included to provide a further understanding of the application, and constitute a part of the specification, which together with the embodiments of the application, are used to explain the application, and do not constitute a limitation on the application.

[0041] In the drawings:

[0042] Figure 1 is a schematic diagram of the interaction principle of DTT and Hg 2+ ;

[0043] Figure 2 is a characterization diagram of the prepared AuNPs modified graphene sensor of the application;

[0044] Figure 3 is a characterization diagram of the prepared AuNPs modified graphene sensor of the application;

[0045] Figure 4The figure shows the optimization results of the reaction time of the graphene sensor modified with DSAI and AuNPs in this invention.

[0046] Figure 5 This is a graph showing the optimized binding time of DSAI to DNA in this invention.

[0047] Figure 6 A comparison chart of current detected by a semiconductor parameter meter for different GFET devices of the present invention;

[0048] Figure 7 This invention relates to a DNA3 sensor based on graphene-AuNPs composite nanomaterials for detecting Hg. 2+ Sensitivity analysis of current and Hg 2+ Concentration relationship;

[0049] Figure 8 This invention relates to a DNA3 sensor based on graphene-AuNPs composite nanomaterials for detecting Hg. 2+ Sensitivity analysis and linear fitting curve;

[0050] Figure 9 This invention relates to a DNA4 sensor based on graphene-AuNPs composite nanomaterials for detecting Hg. 2+ Sensitivity analysis diagram, current versus Hg 2+ Concentration relationship;

[0051] Figure 10 This invention relates to a DNA4 sensor based on graphene-AuNPs composite nanomaterials for detecting Hg. 2+ Sensitivity analysis plot, linear fitting curve;

[0052] Figure 11 This invention relates to a DNA5 sensor based on graphene-AuNPs composite nanomaterials for detecting Hg. 2+ Sensitivity analysis diagram, current versus Hg 2+ Concentration relationship;

[0053] Figure 12 This invention relates to a DNA5 sensor based on graphene-AuNPs composite nanomaterials for detecting Hg. 2+ Sensitivity analysis plot, linear fitting curve;

[0054] Figure 13 This invention relates to a DNA6 sensor based on graphene-AuNPs composite nanomaterials for detecting Hg. 2+ Sensitivity analysis diagram, current versus Hg 2+ Concentration relationship;

[0055] Figure 14 This invention relates to a DNA6 sensor based on graphene-AuNPs composite nanomaterials for detecting Hg. 2+sensitivity analysis chart, linear fitting curve;

[0056] Figure 15 The selective analysis chart of the graphene-AuNPs composite nanomaterial DNA4 sensor for Hg 2+

[0057] Figure 16 The response chart of the graphene-AuNPs composite nanomaterial DNA sensor after adding Hg 2+

[0058] Figure 17 The detection results of the Hg 2+ solution with known concentration. DETAILED DESCRIPTION

[0059] The technical solutions in the embodiments of the present application will be clearly and completely described below in conjunction with specific embodiments. Obviously, the described embodiments are only some of the embodiments of the present application, but not all the embodiments. Based on the embodiments in the present application, all other embodiments obtained by those skilled in the art without creative work fall within the scope of protection of the present application.

[0060] The preparation method of the high-efficiency graphene-carbon nanotube transistor sensor in the embodiment comprises the following steps:

[0061] S1, cleaning of the substrate silicon wafer

[0062] S1.1, preparation of Piranha cleaning solution: mix concentrated H2SO4 and H2O2 according to the volume ratio of 7:3, first slowly pour the concentrated H2SO4 into a culture dish, then use a dropper to slowly add H2O2 along the wall of the culture dish into the concentrated H2SO4, and gently shake the culture dish to fully mix the two;

[0063] S1.2, silicon wafer immersion cleaning: slowly put the cut silicon wafer with a size of 1.5 cm x 1.5 cm into the culture dish containing the Piranha cleaning solution with tweezers, shake the culture dish to ensure that all the silicon wafers are fully immersed and do not overlap each other, adjust the temperature of the constant-temperature magnetic stirrer to 85-95℃, and immerse the silicon wafer at this temperature for 2h until no bubbles are generated on the surface of the silicon wafer;

[0064] S1.3, silicon wafer cleaning and drying: after the cleaning solution is naturally cooled to room temperature, pour it into a waste liquid tank, add ultrapure water to the culture dish, and repeatedly clean the silicon wafer for 3 times, each time for 10 min, after cleaning, blow the silicon wafer dry with nitrogen, and seal it in an IPC vacuum drying dish for standby;

[0065] S2, growth and transfer of graphene

[0066] ​​S2.1, Copper foil pretreatment: cut the copper foil to a size slightly smaller than the quartz boat to avoid wrinkles and bending, and then soak the cut copper foil in acetic acid to remove the oxide on the surface of the copper foil and achieve surface flattening;

[0067] S2.2, Chemical vapor deposition method for growing graphene: place the pretreated copper foil into the quartz boat and send it into the furnace tube, turn on the vacuum pump to reduce the pressure in the furnace to 3-4 Pa to remove most of the air;

[0068] Open the hydrogen valve and adjust the H2 flow rate to 10 sccm, and at the same time use the programmed temperature rising method to raise the temperature in the furnace to 1000℃; after the temperature reaches 1000℃, preheat for 15 min, then introduce CH4, set the CH4 flow rate to 4 sccm, and simultaneously adjust the H2 flow rate to 8 sccm, maintain this flow rate ratio for 35 min, turn off CH4, and cool in a 10 sccm hydrogen environment for 2-3 h to obtain a copper foil with grown graphene, take out the copper foil with graphene grown on the surface and put it into a glass container for standby;

[0069] S2.3, Transfer of graphene: cut the copper foil with graphene grown on it to a size slightly larger than the silicon wafer, drop 0.1-0.15 mL of PMMA glue with a molecular weight of 495000 and a concentration of A6 on the surface of the copper foil, spin coat at a speed of 4000 r / min for 45 s, and then bake at 180℃ for 2 min; coat the graphene silicon wafer with Scotch tape on a glass plate, cut the glue frame to the size of the graphene silicon wafer, and then paste the glue frame to the lower edge of the copper foil and cut off the excess copper sheet and glue frame;

[0070] Transfer the graphene with the glue frame to a FeCl3 solution containing an appropriate amount of HCl solution, HCl prevents Fe 3+ from hydrolyzing, and the copper foil is completely dissolved after 6 hours of sealing and etching, and then use ultrapure water to soak and clean for more than 1 day, and replace the ultrapure water in time during the process;

[0071] Take out the sample and paste it on a clean silicon wafer, place it in a constant temperature magnetic stirrer at 45℃ to dry the moisture, use a blade to cut off the glue frame and leave the transferred graphene, and then put it into an acetone solution for more than 1 day to remove the surface PMMA, and then observe the glue removal effect under a microscope and seal it for standby;

[0072] S3, Preparation of graphene-gold nanoparticle composite nanomaterial sensor

[0073] S3.1, Gold nanoparticle modified graphene: use a pipette to take 100 μL of gold nanoparticle solution with a concentration of 50 μg / mL and vertically drop it on the surface of the transferred graphene, and let it stand at room temperature until the gold nanoparticle solution is completely dried, to obtain a graphene device with gold nanoparticles attached;

[0074] S3.2 Reactive Ion Etching: Preheat the electron tube for 5 minutes, open the nitrogen cylinder and fill with nitrogen for 20 seconds, place the graphene device with gold nanoparticles attached, and then close the nitrogen valve; after completing the gas path cleaning, introduce O2, adjust the O2 flow rate to stabilize the internal pressure of the reaction chamber, set the etching power to 120W, the O2 flow rate to 60mL / min, maintain the working gas pressure at 10Pa, and etch with O2 for 1 minute to obtain the etched graphene strips;

[0075] S3.3 Evaporation of electrodes: Cover the etched graphene strip with a mask and fix its position. Use a resistance evaporation deposition machine to evaporate 80nm Cr and 600nm Au. Place the sample upside down on the card plate, twist the baffle and add a glass cover. Turn on the compressor cooler and the evaporation machine. Before evaporation, evaporate the evaporation machine to absolute vacuum.

[0076] First, deposit Cr by vapor deposition. After switching metals, turn on the evaporation power supply and set the current to about 200A. Vaporize for 8-9 minutes. After Cr is deposited, adjust the current to 0 and then switch to Au metal. Adjust the current to 150A until Au is deposited.

[0077] The reaction program was sequentially shut down, and once the molecular pump reading dropped to 0, the vent valve was opened to break the vacuum, completing the electrode deposition and obtaining Hg based on graphene-gold nanoparticle composite nanomaterials. 2+ sensor;

[0078] S3.4, DNA Modification: DSAI solution was added to the surface of the prepared graphene-gold nanoparticle composite nanomaterial sensor, and the reaction was carried out in the dark for 11 hours. Then, 100 μL of a 40 nM DNA4 solution was added. The DNA4 sequence was 5'-CCACCACTTTTTTTTTGGGGTTTTTTTTT-3'. The reaction was carried out for 45 minutes to complete the DNA modification, yielding a sensor suitable for Hg testing. 2+ Hg detected based on graphene-gold nanoparticle composite nanomaterials 2+ sensor.

[0079] A Hg based on graphene-gold nanoparticle composite nanomaterial prepared by the method of claim 1 2+ Sensor detects Hg 2+ The method includes the following steps:

[0080] T1. Sensor Initial State Detection: Set the gate voltage of the semiconductor parameter analyzer to -8 to 8V and the source-drain voltage to -50mV. Detect the prepared Hg based on graphene-gold nanoparticle composite nanomaterials. 2+ The sensor detects and records the initial current value I0;

[0081] T2. Sample Detection: Different concentrations of Hg are dropped onto the sensor surface.2+ The sample solution was reacted at room temperature for 1 hour. The gate voltage and source-drain voltage of the semiconductor parameter instrument were kept constant. The current value I of the sensor was detected again, and the change in current I0-I was recorded.

[0082] T3. Result Analysis: Based on the current change I0-I and Hg 2+ The linear relationship between concentration and the measured change in current was used to calculate the Hg concentration in the sample solution. 2+ The concentration of Hg; where, when Hg 2+ When the concentration is in the range of 0.05–1 nM, the change in current is related to Hg. 2+ The linear equation for concentration is y = 0.0425x + 0.39475, R0 2 =0.99902, detection limit is 0.31pM;

[0083] T4. Pretreatment of Actual Water Samples: For actual Yangtze River water samples, a water system filter with a pore size of 0.22 μm is used to filter the water sample to remove impurities and particles. Then, Hg is processed according to steps T2 and T3 above. 2+ Testing.

[0084] Example 1: Hg based on graphene-gold nanoparticle composite nanomaterials 2+ Sensor fabrication

[0085] S1. Cleaning of the substrate silicon wafer

[0086] Preparation of Piranha cleaning solution: In a fume hood, slowly pour 70 mL of concentrated H2SO4 into a clean polytetrafluoroethylene petri dish, then slowly add 30 mL of H2O2 along the wall of the petri dish to the concentrated H2SO4 while gently shaking the petri dish to ensure that the two liquids are fully mixed and avoid local overheating.

[0087] Silicon wafer immersion cleaning: Using tweezers, slowly place the pre-cut 1.5cm×1.5cm monocrystalline silicon wafers (99.99% purity) into a petri dish containing Piranha cleaning solution, ensuring that the silicon wafers are completely submerged in the cleaning solution and that the silicon wafers do not contact or overlap each other. Place the petri dish on a thermostatic magnetic stirrer, set the temperature to 90℃, and heat and immerse for 2 hours. During this period, observe the formation of bubbles on the surface of the silicon wafer. When no more bubbles emerge from the surface of the silicon wafer, it indicates that the organic contaminants on the surface of the silicon wafer have been basically removed.

[0088] Silicon wafer cleaning and drying: Turn off the thermostatic magnetic stirrer and allow the cleaning solution to cool naturally to room temperature. Slowly pour it into a dedicated waste liquid container. Add sufficient ultrapure water (prepared by a UPF-10L laboratory ultrapure water system, resistivity 18.2 MΩ・cm) to the petri dish. Gently shake the petri dish to clean the silicon wafer. After soaking for 10 minutes, discard the cleaning water. Repeat this cleaning process 3 times. After cleaning, use a nitrogen gun (nitrogen purity 99.999%) to blow dry the moisture on the surface of the silicon wafer with a gentle airflow. Be careful to avoid excessive nitrogen flow that may scratch the surface of the silicon wafer. Place the dried silicon wafer in an IPC vacuum desiccator and seal it for later use.

[0089] S2, Graphene Growth and Transfer

[0090] Copper foil pretreatment: Cut copper foil with a thickness of 25μm and a purity of 99.99% to a size slightly smaller than the quartz boat (quartz boat size is 10cm×2cm×1cm). During the cutting process, use tweezers to handle it gently to avoid wrinkles, bends or scratches on the copper foil. Put the cut copper foil into a beaker containing a 5% acetic acid solution and soak for 30 minutes. During this period, gently shake the beaker once every 10 minutes to enhance the removal effect of oxides on the surface of the copper foil. After soaking, wash the copper foil with ultrapure water 3 times, 5 minutes each time, and then blow it dry with nitrogen to obtain a smooth and clean copper foil.

[0091] Graphene growth by chemical vapor deposition: The pretreated copper foil is laid flat in a quartz boat, ensuring that the copper foil is placed flat in the quartz boat and does not contact the quartz boat wall. The quartz boat is slowly pushed into the center of the tube of the tube furnace. The tube furnace door is closed, the vacuum pump is turned on, and the pressure in the tube furnace is evacuated to 3-4 Pa. This vacuum is maintained for 10 minutes to fully remove the air from the tube furnace.

[0092] Open the hydrogen valve and adjust the gas mass flow controller to stabilize the H2 flow rate at 10 sccm. Simultaneously, turn on the programmable temperature system of the tube furnace and set the heating rate to 10℃ / min to raise the furnace temperature to 1000℃. Once the temperature reaches 1000℃, maintain the H2 flow rate and preheat at a constant temperature for 15 minutes to further remove any remaining impurities on the copper foil surface. Then, open the methane valve and adjust the CH4 flow rate to 4 sccm, while simultaneously adjusting the H2 flow rate to 8 sccm. Maintain this gas flow ratio for 35 minutes to allow CH4 to decompose on the copper foil surface and grow graphene. After growth is complete, close the methane valve, maintain the H2 flow rate at 10 sccm, and turn on the cooling system of the tube furnace to allow the furnace temperature to cool naturally to room temperature at a rate of 5℃ / min (approximately 2–3 hours). Once the temperature has dropped to room temperature, close the hydrogen valve and vacuum pump, open the tube furnace door, remove the copper foil with graphene grown on its surface, place it in a clean glass container, seal it, and store it for later use.

[0093] Graphene transfer: Cut the copper foil with graphene grown on it into 2cm×2cm squares (slightly larger than the 1.5cm×1.5cm silicon wafer) and lay them flat on a clean glass slide with the graphene side facing up. Use a pipette to pick up 0.12mL of PMMA adhesive with a molecular weight of 495000 and a concentration of A6, and slowly drop it onto the graphene on the copper foil surface, ensuring that the PMMA adhesive evenly covers the entire graphene surface. Place the glass slide in a spin coater, set the spin coater speed to 4000r / min, and the spin coater time to 45s. After spin coater is completed, place the glass slide in a constant temperature drying oven at 180℃ and bake for 2min to allow the PMMA adhesive to fully cure and bond tightly with the graphene.

[0094] Cut the Scoth tape into a frame (1.5cm × 1.5cm) that matches the size of the silicon wafer, and stick it on a clean glass plate. Then, gently attach the copper foil with PMMA / graphene covering the surface (graphene side facing the frame) to the frame. Use tweezers to gently press the edges of the frame to ensure that the copper foil is tightly bonded to the frame. Then, use scissors to cut off the excess copper foil outside the frame.

[0095] Prepare a 0.5 mol / L FeCl3 solution and add an appropriate amount of concentrated HCl (volume ratio 1:100) to prevent Fe from being absorbed. 3+ Hydrolysis: Pour the prepared solution into a petri dish. Place the PMMA / graphene / copper foil sample (copper foil side down) with the adhesive frame attached into the FeCl3 solution. Seal the petri dish and allow it to stand at room temperature for 6 hours to etch. During this period, gently shake the petri dish once every 2 hours to accelerate the dissolution of the copper foil. After the copper foil is completely dissolved (with no obvious copper foil residue in the solution), carefully remove the PMMA / graphene sample with tweezers and transfer it to a petri dish containing ultrapure water for immersion and cleaning. Change the ultrapure water every 4 hours and continue cleaning for more than 1 day to thoroughly remove residual Fe from the sample surface. 3+ and Cl - .

[0096] Gently attach the cleaned PMMA / graphene sample (graphene side down) to the pretreated silicon wafer surface. Use tweezers to gently press the sample edges to ensure tight adhesion between the sample and the silicon wafer. Then, place the silicon wafer on a 45°C constant-temperature magnetic stirrer and slowly stir to dry the moisture on the sample surface. After drying, carefully cut off the adhesive frame at the edge of the sample with a blade, leaving only the PMMA / graphene portion on the silicon wafer. Place the silicon wafer in a petri dish containing acetone solution and soak it at room temperature for more than one day to remove the PMMA adhesive from the graphene surface. After soaking, remove the silicon wafer and observe the adhesive removal effect on the graphene surface under an optical microscope to ensure that the PMMA adhesive has been completely removed. Then, place the silicon wafer in an IPC vacuum desiccator and seal it for later use.

[0097] S3. Fabrication of graphene-gold nanoparticle (AuNPs) composite nanomaterial sensors

[0098] Gold nanoparticles modified graphene: Gold nanoparticles (AuNPs) with a particle size of 10 nm were purchased from Xi'an Qiyue Biological Reagent Co., Ltd., and diluted with ultrapure water to a concentration of 50 μg / mL. 100 μL of AuNPs solution of this concentration was pipetted and vertically dropped onto the surface of the silicon wafer (graphene area) on which graphene was transferred, ensuring that the AuNPs solution uniformly covered the entire graphene surface. The silicon wafer was placed in a dust-free environment at room temperature and allowed to stand until the AuNPs solution was completely dried, thus obtaining the graphene device with AuNPs attached.

[0099] Reactive Ion Etching (RIE): Turn on the low-noise lock-in amplifier (MF-DEV5811) and the reactive ion etching instrument, preheat the electron tube for 5 minutes, place the graphene device with AuNPs attached into the sample chamber of the etching instrument, close the sample chamber door, open the nitrogen cylinder, and fill the sample chamber with nitrogen for 20 seconds to clean the residual gas in the sample chamber. Close the nitrogen valve, open the oxygen valve, adjust the O2 flow rate to stabilize the internal pressure of the sample chamber at 10 Pa, set the etching power to 120 W, start the etching program, and etch with O2 for 1 minute to remove graphene and other impurities outside the template to obtain etched graphene strips. After etching is completed, turn off the etching instrument and the oxygen valve, and take out the sample for later use.

[0100] Evaporation electrode: Fix the etched graphene strip sample on the sample stage of the evaporation machine. Cover the sample surface with a mask (matching the size of the graphene strip), ensuring the mask is accurately positioned and firmly fixed. Turn on the compressor cooler and vacuum pump of the resistance evaporation coating machine to evaporate the vacuum chamber to absolute vacuum (pressure below 1×10⁻⁶). -4 First, place the Cr metal wire into the evaporation boat, turn on the evaporation power supply, and slowly adjust the current to 200A to start Cr deposition. The deposition time is 8-9 minutes to ensure that the Cr coating thickness reaches 80nm. After the Cr deposition is completed, turn off the evaporation power supply and wait for the evaporation boat to cool down. Then, replace it with Au metal wire, turn on the evaporation power supply again, adjust the current to 150A, and start Au deposition until the Au coating thickness reaches 600nm. After the deposition is completed, turn off the evaporation power supply and vacuum pump. After the vacuum chamber pressure returns to atmospheric pressure, open the vacuum chamber door, take out the sample, and obtain the AuNPs modified graphene sensor.

[0101] DNA modification: Tetraphenylene monoquaternary ammonium salt (DSAI) was purchased from Xi'an Qiyue Biotechnology Co., Ltd., and dissolved and diluted to a concentration of 1 pM using dimethyl sulfoxide (DMSO, Shanghai Aladdin Biochemical Technology Co., Ltd.). 20 μL of this DSAI solution was pipetted onto the surface of the AuNPs-modified graphene sensor, and the sensor was placed in a light-protected environment for 11 h. Subsequently, DNA4 (sequence 5'-CCACCACTTTTTTTTTGGGGTTTTTTTTT-3') purified by HPLC was purchased from Sangon Biotech (Shanghai) Co., Ltd., and diluted to a concentration of 40 nM using ultrapure water. 100 μL of this DNA4 solution was pipetted onto the sensor surface, and the reaction was carried out at room temperature for 45 min to complete the DNA modification, ultimately obtaining Hg based on graphene-gold nanoparticle composite nanomaterials. 2+ sensor.

[0102] Example 2: Hg based on the above sensor 2+ Detection

[0103] T1. Sensor Initial State Detection: The gate voltage of the semiconductor parameter analyzer (B1500A Keysight, Suzhou Measurement & Trading Co., Ltd.) was set to -8 to 8V, and the source-drain voltage was set to -50mV. The Hg prepared in Example 1 was then used... 2+ The sensor is connected to the probe station (P-150, Chengdu Cexin Technology Co., Ltd.) of the semiconductor parameter analyzer. The parameter analyzer is started to detect the initial current value of the sensor and record it as I0.

[0104] T2, Standard Hg 2+ Solution testing: Mercuric nitrate (Hg(NO3)2) was purchased from Shandong Xiya Chemical Industry Co., Ltd., and Hg solutions with concentrations of 1 pM, 5 pM, 10 pM, 50 pM, 100 pM, 500 pM, 1 nM, 5 nM, and 10 nM were prepared using ultrapure water. 2+ Standard solutions were prepared by pipetting 100 μL of different concentrations of Hg. 2+ A standard solution was added dropwise to the sensor surface and allowed to react at room temperature for 1 hour. After the reaction, the voltage parameter of the semiconductor parameter analyzer was kept constant, and the current value of the sensor was measured again and recorded as I. The change in current I0-I was calculated. Based on the detection results, the change in current was plotted against Hg. 2+ The concentration relationship curve yields the result when Hg 2+ When the concentration is in the range of 0.05–1 nM, the linear equation is y = 0.0425x + 0.39475, R0 2 =0.99902, detection limit is 0.31pM;

[0105] T3. Selective Detection: Reagents including manganese chloride (MnCl2·4H2O), copper sulfate (CuSO4·5H2O), lead acetate (Pb(CH3COO)2·3H2O), calcium chloride (CaCl2·2H2O), ferrous sulfate (FeSO4·7H2O), sodium nitrate (NaNO3), potassium chloride (KCl), zinc chloride (ZnCl2), and magnesium chloride (MgCl2·6H2O) were purchased from Sinopharm Chemical Reagent Co., Ltd., etc. Cu chloride solutions with a concentration of 10 nM were prepared using ultrapure water. 2+ K + Fe 2+ Mg 2+ Na + Pb 2+ Ca 2+ Mn 2+ Zn 2+ Following step T2, these ion solutions were added dropwise to the sensor surface, and the changes in sensor current were detected and recorded. The results showed that only when the sensor is in contact with Hg... 2+ When applied, the current intensity decreases significantly more than with other ions, demonstrating that the sensor has excellent selectivity for Hg²⁺.

[0106] T4. Actual Water Sample Testing: Collect Yangtze River water samples and filter them using a water system filter with a pore size of 0.22 μm to remove impurities. Take 100 μL of the filtered water sample and add it to the sensor surface as described in step T2. Detect the change in sensor current and calculate the Hg concentration in the water sample using a linear equation. 2+ The concentration of Hg in the Yangtze River water samples was [data missing]. 2+ The concentration was 103 pM, which meets the national standard requirement of 199.4 pM. Meanwhile, Hg was added to the filtered Yangtze River water samples at concentrations of 0.05 nM, 0.1 nM, 0.5 nM, and 1 nM, respectively. 2+ The standard solution was used for a spiked recovery experiment, and the results were as follows: Figure 17 As shown in the table, the recovery rate ranged from 98.36% to 124%, and the relative standard deviation (RSD) ranged from 2.175% to 3.195%, demonstrating that the sensor has good accuracy and reliability in actual water sample testing.

[0107] Example 3: DTT on Hg 2+ Verification of the regulatory role of detection

[0108] The Hg²⁺ sensor was prepared according to the method of Example 1, and the initial current value I0 of the sensor was detected according to step T1 of Example 2.

[0109] Use a pipette to draw 100 μL of 10 nM Hg solution. 2+The solution was dropped onto the sensor surface and reacted at room temperature for 1 hour. The current value I1 of the sensor was then detected, and the change in current I0-I1 was recorded.

[0110] Dithiothreitol (DTT) was purchased from Sigma-Aldrich (Shanghai) Trading Co., Ltd. A 10 mM DTT solution was prepared using ultrapure water. 100 μL of this DTT solution was pipetted and added dropwise to the above-mentioned solution containing Hg. 2+ After the sensor surface has been reacted, the current value I2 of the sensor is detected after 30 minutes at room temperature, and the change in current I2-I1 is recorded.

[0111] Use a pipette to draw another 100 μL of 10 nM Hg solution. 2+ The solution was dropped onto the sensor surface and reacted at room temperature for 1 hour. The current value I3 of the sensor was then detected, and the change in current I2-I3 was recorded.

[0112] The results showed that adding Hg 2+ The sensor current decreased significantly (I0-I1 was relatively large); after adding DTT, the current rebounded significantly (I2-I1 was positive); Hg was added again. 2+ Subsequently, the current decreased again (I2-I3 was relatively large), proving that DTT can react with Hg through thiol groups. 2+ Combined, regulating Hg 2+ The interaction with DNA, which in turn affects the current response of the sensor, can be used to reduce Hg. 2+ Non-specific adsorption improves detection accuracy.

[0113] It will be apparent to those skilled in the art that the present invention is not limited to the details of the exemplary embodiments described above, and that the invention can be implemented in other specific forms without departing from the spirit or essential characteristics of the invention. Therefore, the embodiments should be considered in all respects as exemplary and non-limiting, and the scope of the invention is defined by the appended claims rather than the foregoing description. Thus, it is intended that all variations falling within the meaning and scope of equivalents of the claims be included within the present invention.

[0114] Furthermore, it should be understood that although this specification describes embodiments, not every embodiment contains only one independent technical solution. This narrative style is merely for clarity. Those skilled in the art should consider the specification as a whole, and the technical solutions in each embodiment can also be appropriately combined to form other embodiments that can be understood by those skilled in the art.

Claims

1. A Hg based on graphene-gold nanoparticle composite nanomaterials 2+ The sensor fabrication method is characterized by, Includes the following steps: S1. Cleaning of the substrate silicon wafer S1.1 Preparation of Piranha cleaning solution: Mix concentrated H2SO4 and H2O2 at a volume ratio of 7:

3. First, slowly pour the concentrated H2SO4 into the petri dish, and then use a dropper to slowly add H2O2 along the wall of the petri dish to the concentrated H2SO4. Gently shake the petri dish to mix the two thoroughly. S1.2, Silicon wafer immersion cleaning: Use tweezers to slowly place the cut silicon wafers, which are 1.5cm×1.5cm in size, into a petri dish containing Piranha cleaning solution. Shake the petri dish to ensure that all silicon wafers are completely submerged and do not overlap. Adjust the temperature of the thermostatic magnetic stirrer to 85-95℃ and immerse the silicon wafers at this temperature for 2 hours until no more bubbles are generated on the surface of the silicon wafers. S1.3, Silicon wafer cleaning and drying: After the cleaning solution has cooled to room temperature, pour it into the waste liquid bucket, add ultrapure water to the petri dish, and clean the silicon wafer repeatedly 3 times, each time for 10 minutes. After cleaning, blow the silicon wafer dry with nitrogen gas and put it into an IPC vacuum desiccator and seal it for later use. S2, Graphene Growth and Transfer S2.1 Copper foil pretreatment: Cut copper foil to a size slightly smaller than the quartz boat to avoid wrinkles and bends. Soak the cut copper foil in acetic acid to remove oxides from the surface of the copper foil and achieve surface smoothing. S2.2 Chemical vapor deposition method for graphene growth: The pretreated copper foil is placed into a quartz boat and sent into the tube furnace tube. The vacuum pump is turned on to pump the pressure inside the furnace to 3-4 Pa to remove most of the air. Open the hydrogen valve, adjust the H2 flow rate to 10 sccm, and simultaneously raise the furnace temperature to 1000℃ using a programmed temperature rise method. After the temperature reaches 1000℃, preheat for 15 minutes, then introduce CH4, set the CH4 flow rate to 4 sccm, and simultaneously adjust the H2 flow rate to 8 sccm. Maintain this flow rate ratio for 35 minutes, then turn off CH4 and cool in a 10 sccm hydrogen environment for 2-3 hours to obtain a copper foil with graphene grown on it. Take out the copper foil with graphene grown on its surface and place it in a glass container for later use. S2.3 Graphene Transfer: Cut the copper foil with graphene grown on it into a shape slightly larger than the silicon wafer. Drop 0.1-0.15 mL of PMMA adhesive with a molecular weight of 495,000 and a concentration of A6 onto the surface of the copper foil. Spin coat it at 4000 r / min for 45 s, and then bake it at 180℃ for 2 min. Stick Scoth tape onto a glass plate. Cut a frame for the adhesive tape based on the silicon wafer coated with graphene. Stick the frame to the lower edge of the copper foil and cut off the excess copper foil and frame. Graphene with a frame attached was transferred to a FeCl3 solution containing an appropriate amount of HCl. The HCl prevented Fe from being absorbed. 3+ Hydrolysis, sealing and etching for 6 hours until the copper foil is completely dissolved, then soaking and cleaning with ultrapure water for more than 1 day, changing the ultrapure water in time during the process; The sample was retrieved and attached to a clean silicon wafer. It was then placed on a 45°C constant temperature magnetic stirrer to dry the moisture. The glue frame was cut off with a blade, leaving the transferred graphene. The graphene was then soaked in acetone solution for more than one day to remove the PMMA on the surface. The glue removal effect was observed under a microscope and then sealed for later use. S3. Fabrication of graphene-gold nanoparticle composite nanomaterial sensors S3.1 Gold nanoparticle-modified graphene: Use a pipette to take 100 μL of a 50 μg / mL gold nanoparticle solution and drop it vertically onto the transferred graphene surface. Let it stand at room temperature until the gold nanoparticle solution is completely dried to obtain a graphene device with gold nanoparticles attached. S3.2 Reactive Ion Etching: Preheat the electron tube for 5 minutes, open the nitrogen cylinder and fill with nitrogen for 20 seconds, place the graphene device with gold nanoparticles attached, and then close the nitrogen valve; after completing the gas path cleaning, introduce O2, adjust the O2 flow rate to stabilize the internal pressure of the reaction chamber, set the etching power to 120W, the O2 flow rate to 60mL / min, maintain the working gas pressure at 10Pa, and etch with O2 for 1 minute to obtain the etched graphene strips; S3.3 Evaporation of electrodes: Cover the etched graphene strip with a mask and fix its position. Use a resistance evaporation deposition machine to evaporate 80nm Cr and 600nm Au. Place the sample upside down on the card plate, twist the baffle and add a glass cover. Turn on the compressor cooler and the evaporation machine. Before evaporation, evaporate the evaporation machine to absolute vacuum. First, deposit Cr by vapor deposition. After switching the metal, turn on the evaporation power supply, set the current to about 200A, and vapor deposit for 8 to 9 minutes. After Cr is deposited, the current is adjusted to 0 and then switched to metal Au. The current is adjusted to 150A until Au deposition is complete. The reaction program was sequentially shut down, and once the molecular pump reading dropped to 0, the vent valve was opened to break the vacuum, completing the electrode deposition and obtaining Hg based on graphene-gold nanoparticle composite nanomaterials. 2+ sensor; S3.4, DNA Modification: DSAI solution was added to the surface of the prepared graphene-gold nanoparticle composite nanomaterial sensor, and the reaction was carried out in the dark for 11 hours. Then, 100 μL of a 40 nM DNA4 solution was added. The DNA4 sequence was 5'-CCACCACTTTTTTTTTGGGGTTTTTTTTT-3'. The reaction was carried out for 45 minutes to complete the DNA modification, yielding a sensor suitable for Hg testing. 2+ Hg detected based on graphene-gold nanoparticle composite nanomaterials 2+ sensor.

2. A Hg based on graphene-gold nanoparticle composite nanomaterial prepared by the method of claim 1 2+ Sensor detects Hg 2+ The method is characterized by, Includes the following steps: T1. Sensor Initial State Detection: Set the gate voltage of the semiconductor parameter analyzer to -8 to 8V and the source-drain voltage to -50mV. Detect the prepared Hg based on graphene-gold nanoparticle composite nanomaterials. 2+ The sensor detects and records the initial current value I0; T2. Sample Detection: Different concentrations of Hg are dropped onto the sensor surface. 2+ The sample solution was reacted at room temperature for 1 hour. The gate voltage and source-drain voltage of the semiconductor parameter instrument were kept constant. The current value I of the sensor was detected again, and the change in current I0-I was recorded. T3. Result Analysis: Based on the current change I0-I and Hg 2+ The linear relationship between concentration and the measured change in current was used to calculate the Hg concentration in the sample solution. 2+ The concentration of Hg; where, when Hg 2+ When the concentration is in the range of 0.05–1 nM, the change in current is related to Hg. 2+ The linear equation for concentration is y = 0.0425x + 0.39475, R0 2 =0.99902, detection limit is 0.31pM; T4. Pretreatment of Actual Water Samples: For actual Yangtze River water samples, a water system filter with a pore size of 0.22 μm is used to filter the water sample to remove impurities and particles. Then, Hg is processed according to steps T2 and T3 above. 2+ Testing.

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