A dual-responsive hydrogel and its preparation method and use

Abstract

The application discloses a preparation method of a hydrogen peroxide and zinc ion double-response hydrogel precursor, and the method comprises the following steps: firstly, preparing a Fe-MOF suspension, and then mixing the Fe-MOF suspension with a buffer solution of acrylamide and N,N-methylene bisacrylamide, and a solution containing an oxidant and tetramethylethylenediamine to obtain a hydrogel precursor. Then, the hydrogel precursor is dried to obtain a hydrogel. The application further discloses the use of the hydrogel precursor and the hydrogel. According to the preparation method, the reaction time is short, the preparation method is simple, and the method has certain universality. The prepared hydrogel material has good mechanical properties, a large specific surface area, a three-dimensional network structure, good electrical conductivity, and the property of fast response to changes, and can be used in electrochemical sensors, and has wide application prospects in the fields of biological detection, disease diagnosis, food hygiene and the like.

Description

Technical Field

[0001] This invention relates to the field of sensor material preparation, and more specifically, to an acrylamide dual-response hydrogel doped with zinc finger peptides and Fe-MOF, its preparation method and uses, wherein the dual-response hydrogel can generate rapid dual responses to both hydrogen peroxide and zinc ions. Background Technology

[0002] Hydrogels possess a large specific surface area and a unique three-dimensional network structure, enabling them to hold a large number of molecules with different functions and thus be endowed with various functionalities, such as good conductivity, strong electrochemical signals, and excellent catalytic activity. Simultaneously, the microenvironment of hydrogels can enhance the stability and bioactivity of biomolecules. Furthermore, the high permeability of hydrogels can accelerate the transport of small molecules and ions, as well as the rapid transfer of electrons. These unique properties make hydrogels highly promising for constructing electrochemical immunosensing interfaces. Because the three-dimensional porous structure of conductive hydrogels not only increases the effective electrochemical area but also accelerates mass transport and electron transfer, it can significantly enhance the electrochemical signal. Responsive hydrogels are an interesting type of hydrogel whose physical properties can switch between different states by applying external stimuli such as temperature, pH, pressure, and ions. Due to their unique physical or chemical properties, smart responsive hydrogels can be applied to the design and assembly of various electrochemical immunosensors to achieve superior performance. However, a significant drawback of responsive hydrogels is their relatively long response time to state changes and low response signal intensity. Switching times can range from tens of minutes to several hours, resulting in long detection times. Furthermore, the release rate of a single reactive hydrogel is insufficient, resulting in a low current signal for the analyte and a narrow detection range when applied to a sensor.

[0003] A preparation process for a dual-network, temperature-responsive ion-conducting hydrogel has been reported in the prior art (Qian Pang, Hongtao Hu, Haiqi Zhang, Bianbian Qiao, and Lie Ma, ACS Appl. Mater. Interfaces 2022, 14, 23, 26536–26547). The hydrogel uses a polyvinylpyrrolidone (PVP) / tannic acid (TA) / Fe... 3+ A cross-linked network was introduced into an N,N-methylenediacrylamide-crosslinked poly(N-isopropylacrylamide-co-acrylamide) network. This resulted in good tensile strength, rapid temperature response, and good electrical conductivity. Furthermore, PVP / TA / Fe... 3+ The introduction of cross-linked networks endows hydrogels with excellent stretchability and conductivity. Hydrogel-based sensors can detect ambient temperature in real time and can be applied to implantable temperature sensors for detecting abnormal hyperthermia caused by fever or infection.

[0004] However, the above technology still has the following drawbacks: 1) The conductive hydrogel has low sensitivity to temperature response signals. 2) The readout signal of the hydrogel within the linear temperature range is easily affected by the phase transition temperature and is prone to large errors.

[0005] Another technique (T. Yao, J. Feng, C. Chu, Z. Ma, H. Han, Sens. Actuators BChem., 2021, 348.) proposes a novel cascade controlled-release system based on pH-responsive ZIF-8 capsules and enzyme-responsive hyaluronic acid hydrogels for the electrochemical detection of tumor markers. Glucose in the tube acts as the "primary trigger," and under the catalysis of glucose oxidase modified on immunomagnetic beads, it can be efficiently converted into glucose, providing a large number of protons. Due to the acidic conditions, the hyaluronidase encapsulated in ZIF-8 is released. Upon addition of hyaluronidase to the electrode covered by a hyaluronic acid hydrogel containing methylene blue molecules, the hydrogel is sequentially disintegrated, leading to the release of methylene blue. Under optimal conditions, this method shows a detection range of 1 × 10⁻⁶ for prostate-specific antigen. -3 -1×10 2 ng mL -1 The detection limit is extremely low, at 37.27 fg / mL. -1 .

[0006] However, the above technologies still have the following drawbacks: 1) The gel response time is as long as 2 hours, which is too long and limits the application range. 2) The intensity of the gel response signal is greatly affected by a single factor, making it difficult to improve the detection sensitivity. If a sensing substrate with a sufficiently fast release rate can be found, it will advance the progress of electrochemical sensing.

[0007] In view of this, the present invention is hereby proposed. Summary of the Invention

[0008] To address the aforementioned issues, this invention designs and synthesizes a novel dual-response hydrogel, which is synthesized from acrylamide and methylenebisacrylamide and contains zinc finger peptide (ZFP) and Fe-MOF, enabling it to exhibit dual responses to hydrogen peroxide and zinc ions.

[0009] According to a first aspect of the present invention, one object of the present invention is to provide a method for preparing a hydrogel precursor that is dual-responsive to hydrogen peroxide and zinc ions, the method comprising the following steps:

[0010] (1) Preparation of Fe-MOF suspension

[0011] Iron salts are dissolved in a surfactant solution to form solution A; polyphenols are dissolved in an alkaline solution containing surfactants to form solution B; solutions A and B are then mixed and stirred, and subjected to a high-temperature hydrothermal reaction to form Fe-MOF. The resulting precipitate is centrifuged, washed with water and absolute ethanol, and then redispersed in water to form a suspension C of Fe-MOF.

[0012] (2) Dissolve acrylamide and N,N-methylenebisacrylamide in a buffer solution, add 0.5 μM zinc finger peptide aqueous solution, and stir continuously to form solution D; disperse the oxidant and tetramethylethylenediamine in water to form solution E;

[0013] (3) Mix the solutions D and E obtained in step (2) with the Fe-MOF suspension C obtained in step (1) to obtain the hydrogel precursor.

[0014] In step (1):

[0015] The iron salt may be selected from at least one of ferric chloride hexahydrate, ferric sulfate, and ferric nitrate, preferably ferric chloride hexahydrate.

[0016] The solvent in solvent A can be selected from at least one of deionized water, ethanol and methanol, with deionized water being preferred.

[0017] In solution A, the molar mass concentration of the iron salt is 0.01–0.1 mol / L.

[0018] The surfactant may be selected from at least one of polyvinylpyrrolidone (molecular weight 25KD-40KD), quaternized polyacrylamide (molecular weight 20KD-50KD), polyacrylic acid (molecular weight 2KD-5KD), and polyethylene oxide (molecular weight 20KD-50KD), preferably polyvinylpyrrolidone, and more preferably polyvinylpyrrolidone with a molecular weight of 25-40KD.

[0019] The polyphenol is selected from at least one of ellagic acid, tannic acid, flavonoids, phenolic acids, and polyphenol amides, with ellagic acid being preferred.

[0020] The mass ratio of the iron salt to the surfactant is 0.03:1 to 0.1:1, preferably 0.065:1.

[0021] In solution B, the mass ratio of the polyphenol to the surfactant is 0.02:1 to 0.1:1, preferably 0.068:1.

[0022] The alkali in solution B is selected from at least one of sodium hydroxide, potassium hydroxide, lithium hydroxide, and ammonia water, preferably sodium hydroxide; the molar concentration of the alkali is 20-50 mM, preferably 22.5 mM.

[0023] The solvent in solvent B may be selected from at least one of deionized water, ethanol and methanol, with deionized water being preferred.

[0024] The volume ratio of solution A to solution B is 0.1:1 to 2.0:1, preferably 1:1.

[0025] The high-temperature hydrothermal reaction temperature is between 100-180℃, preferably 130℃.

[0026] The hydrothermal reaction time is between 2 and 8 hours, preferably 4 hours.

[0027] The mass concentration of Fe-MOF in the Fe-MOF suspension C is between 10 and 50 g / L, preferably 20 g / L.

[0028] In step (2):

[0029] The pH of the buffer solution can be 3.0 to 8.0, preferably 6.0.

[0030] The buffer solution is selected from phosphate buffer, carbonate buffer, citrate buffer, and acetate buffer, preferably phosphate buffer (PBS).

[0031] In solution D, the mass ratio of acrylamide to N,N-methylenebisacrylamide is 10:1 to 20:1, preferably 14:1.

[0032] In solution D, the mass concentration of acrylamide is between 100 g and 300 g / L, preferably 140 g / L.

[0033] In solution E, the oxidant can be selected from at least one of hydrogen peroxide, ammonium persulfate, potassium permanganate, and peracetic acid, preferably ammonium persulfate;

[0034] The mass concentration of the oxidant is 50-200 g / L, preferably 100 g / L.

[0035] In solution E, the ratio of tetramethylethylenediamine to water is between 0.01:1 and 0.1:1, preferably 0.05:1.

[0036] In step (3)

[0037] The volume ratio of the suspension C, solution D and solution E is between 0.05:10:1 and 2:10:5, preferably 1:10:2.

[0038] According to a second aspect of the present invention, a second object of the present invention is to provide a hydrogel precursor that is dual-responsive to hydrogen peroxide and zinc ions, prepared by the above method.

[0039] According to a third aspect of the invention, a third object of the invention is to provide a hydrogel formed by drying the said dual-response hydrogel precursor.

[0040] According to a fourth aspect of the present invention, a fourth object of the present invention is to provide a method for preparing the hydrogel, comprising drying the hydrogel precursor according to the present invention to obtain the hydrogel, wherein the drying temperature is between 25 and 50°C, preferably 37°C, and the corresponding time is 15 min;

[0041] According to a fifth aspect of the invention, a fifth object of the invention is to provide an electrochemical sensor platform modified by the responsive hydrogel, the electrochemical sensor platform comprising an electrode, a sodium alginate-nickel ion-graphene oxide gel layer and the hydrogel layer according to the invention.

[0042] According to a sixth aspect of the present invention, a sixth object of the present invention is to provide a method for preparing the electrochemical sensor platform, comprising:

[0043] First, the electrode surface is cleaned, then a sodium alginate-nickel ion-graphene oxide gel layer is formed on the electrode surface, and then the hydrogel precursor according to the present invention is drop-coated onto the electrode surface and dried to obtain the electrochemical sensor platform.

[0044] According to a seventh aspect of the invention, a seventh object of the invention is to provide the electrochemical sensor platform for use as an immune sensor, preferably for detecting the concentration of the tumor marker precursor gastrin-releasing peptide (ProGRP).

[0045] Beneficial effects

[0046] (1) The hydrogel preparation method provided by this invention has the following advantages: short reaction time, simple preparation method and certain universality (it can be applied to the design of other dual-response hydrogels). The prepared novel hydrogel material has good mechanical properties, large specific surface area, three-dimensional network structure, good conductivity and rapid response to changes.

[0047] (2) It is used in electrochemical sensors and has broad application prospects in fields such as biological detection, disease diagnosis, and food hygiene. Attached Figure Description

[0048] To more clearly illustrate the specific embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the specific embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are some embodiments of the present invention. For those skilled in the art, other drawings can be obtained from these drawings without creative effort.

[0049] Figure 1 A scanning electron microscope image of the dual-response hydrogel prepared in Example 1;

[0050] Figure 2 This is a photograph of the Fe-MOF prepared in Example 1.

[0051] Figure 3 The image shows the X-ray photoelectron spectrum of the dual-response hydrogel obtained in Example 1.

[0052] Figure 4 This is a scanning electron microscope image of the dual-response hydrogel prepared in Example 2.

[0053] Figure 5 The graph shows the changes in hydrogen peroxide and zinc ion concentrations of the dual-response hydrogel prepared in Example 1.

[0054] Figure 6 This is a standard curve of electrochemical signal versus tumor marker concentration in an application example. Detailed Implementation

[0055] The present invention will now be described in detail. Before proceeding with the description, it should be understood that the terminology used in this specification and the appended claims should not be construed as limited to its general or dictionary meaning, but rather should be interpreted according to the meaning and concept corresponding to the technical aspects of the invention, based on the principle that the inventors are allowed to appropriately define the terms for the best interpretation. Therefore, the description presented herein is merely a preferred example for illustrative purposes and is not intended to limit the scope of the invention. It should be understood that other equivalents or modifications can be obtained from it without departing from the spirit and scope of the invention.

[0056] The biresponsive hydrogel according to the present invention is synthesized from acrylamide and methylenebisacrylamide, and contains zinc finger peptide (ZFP) and Fe-MOF, and is capable of responding to Zn. 2+ A dual reaction occurs with H₂O₂. Zinc ions act as one of the signal amplification switches, specifically interacting with cysteine ​​and histidine in the ZFP of the hydrogel to form numerous exposed pores. H₂O₂ acts as another signal amplification switch, drawing energy from Fe... 3+ To Fe 2+ The transformation. In Fe-MOF, Fe... 2+ and Fe 3+ The coordination abilities of the Fe-MOF and Fe-MOF are completely different. When the valence state changes, the Fe-MOF structure is disrupted, and this transformation directly affects the stability of the MOF. The coordination of iron ions with catechol groups forms Fe-MOF, in which the iron ions exist stably in the trivalent form. This is different from Fe... 3+ In comparison, Fe 2+The coordination ability of the molecule is significantly insufficient. The above transformation improves the conductivity of the hydrogel. A quantitative relationship between the glucose oxidase-loaded ZIF-8 and the analyte content was established. GOx loaded on ZIF-8 catalyzes the conversion of glucose to gluconic acid and H2O2. ZIF-8 is degraded through local acidification, leading to the degradation of Zn. 2+ Release. Dual-responsive hydrogel for Zn 2+ It exhibits a significant dual reaction with H2O2, resulting in a faster response and higher sensitivity compared to single-reaction hydrogels.

[0057] The electrochemical sensor platform according to the present invention consists of a signal layer (sodium alginate-nickel ion-graphene oxide (SA-Ni)). 2+ The electrochemical sensor platform consists of a silencing layer (a dual-response hydrogel) and a silencing layer. It can be used as, for example, an immunosensor, and has broad application prospects in fields such as biological detection, disease diagnosis, and food hygiene.

[0058] In the preparation method of the hydrogen peroxide and zinc ion dual-responsive hydrogel precursor according to the present invention, the mass ratio of the iron salt and the surfactant in step (1) is 0.03:1 to 0.1:1, preferably 0.065:1. If the ratio is too low, the Fe-MOF size is too large and it is not suitable for doping into the gel. If the ratio is too high, the Fe-MOF size is too small and its influence on the gel is low when it changes.

[0059] In solution B, the mass ratio of the polyphenol to the surfactant is 0.02:1 to 0.1:1, preferably 0.068:1. If the ratio is too low, the Fe-MOF size is too large and it is not suitable for doping into the gel. If the ratio is too high, the Fe-MOF size is too small and its influence on the gel is low when it changes.

[0060] The mass concentration of Fe-MOF in the Fe-MOF suspension C is between 10 and 50 g / L, preferably 20 g / L. If the concentration of Fe-MOF is too high, it will affect the stability of the gel, while if the concentration of Fe-MOF is too low, the change in Fe-MOF will have little effect on the gel.

[0061] In solution D, the mass ratio of acrylamide to N,N-methylenebisacrylamide is 10:1 to 20:1, preferably 14:1. If the ratio is too high, the degree of cross-linking of the gel will be low, and the polymerization time will be long.

[0062] The mass concentration of the oxidant is 50-200 g / L, preferably 100 g / L. An appropriate concentration of oxidant can promote the rapid polymerization of the gel. Too low a concentration of oxidant has a low promoting effect, while too high a concentration of oxidant leaves more residue after the reaction, affecting the stability of the gel.

[0063] In solution E, the ratio of tetramethylethylenediamine to water is between 0.01:1 and 0.1:1, preferably 0.05:1. An appropriate concentration of tetramethylethylenediamine can promote the rapid polymerization of the gel. Too low a concentration of tetramethylethylenediamine has a low promoting effect, while too high a concentration of tetramethylethylenediamine leaves more residue after the reaction, affecting the stability of the gel.

[0064] The volume ratio of suspension C, solution D and solution E in step (3) is between 0.05:10:1 and 2:10:5, preferably 1:10:2. An unsuitable ratio will affect the gel stability and responsiveness. A volume ratio within the above range will help to obtain the best gel stability and responsiveness.

[0065] In the preparation method of the hydrogel according to the present invention, the drying temperature needs to be accurately controlled between 25 and 50°C, preferably 37°C. If the temperature is too high, it will affect the water retention of the gel; if the temperature is too low, a longer drying time is required.

[0066] Unless otherwise expressly stated, numerical ranges throughout the application include any subranges therein and any numerical values ​​incremented by the smallest subunit of a given value. Unless otherwise expressly stated, numerical values ​​throughout the application represent approximate measures or limitations on the range of embodiments including minor deviations from a given value and having approximately the mentioned value as well as having the mentioned precise value. Except in the detailed description of the working embodiments provided at the end, all numerical values ​​of parameters (e.g., quantities or conditions) in this application (including the appended claims) should in all cases be understood to be modified by the term “approximately,” regardless of whether “approximately” actually precedes the numerical value. “Approximately” indicates that the stated numerical value allows for slight inaccuracies (some close to precision at that value; approximately or reasonably close to the value; approximate). If the inaccuracy provided by “approximately” is not understood in this common sense in the art, then “approximately” as used herein at least indicates a variation that can be produced by common methods of measuring and using these parameters. For example, “approximately” can include variations less than or equal to 10%, less than or equal to 5%, less than or equal to 4%, less than or equal to 3%, less than or equal to 2%, less than or equal to 1%, or less than or equal to 0.5%.

[0067] In this document, all features or conditions defined in the form of numerical ranges or percentage ranges are for the sake of brevity and convenience only. Accordingly, descriptions of numerical ranges or percentage ranges should be considered as covering and specifically disclosing all possible secondary ranges and individual values ​​within those ranges, particularly integer values. For example, a range description of "1 to 8" should be considered as specifically disclosing all secondary ranges such as 1 to 7, 2 to 8, 2 to 6, 3 to 6, 4 to 8, 3 to 8, etc., particularly secondary ranges defined by all integer values, and should be considered as specifically disclosing individual values ​​within those ranges such as 1, 2, 3, 4, 5, 6, 7, 8, etc. Unless otherwise specified, the foregoing interpretation applies to all content throughout this invention, regardless of its scope.

[0068] If a quantity or other numerical value or parameter is expressed as a range, a preferred range, or a series of upper and lower limits, it should be understood that this document has specifically disclosed all ranges consisting of any upper or preferred value of that range and the lower or preferred value of that range, regardless of whether such ranges are separately disclosed. Furthermore, when a range of numerical values ​​is mentioned herein, unless otherwise stated, the range shall include its endpoints and all integers and fractions within the range.

[0069] In this document, numerical values ​​are to be understood as having a precision with significant digits, provided that the purpose of the invention can be achieved. For example, the number 40.0 should be understood to cover a range from 39.50 to 40.49.

[0070] Unless otherwise stated, all polymer molecular weights in this article are expressed as number-average molecular weights.

[0071] The reagents and solvents disclosed below were purchased from Sigma-Aldrich China and were all of analytical grade. The ultrapure water used in the experiments was obtained using a Pastikar PSDK-30-E ultrapure water system.

[0072] X-ray photoelectron spectroscopy (XPS) data were measured using an Escalab 250 Thermofisher series instrument from the USA; electrochemical tests were performed using a CHI 660E electrochemical workstation series instrument from Shanghai Chenhua Instruments Co., Ltd.

[0073] The following embodiments are merely examples illustrating implementations of the present invention and do not constitute any limitation on the present invention. Those skilled in the art will understand that modifications made without departing from the spirit and concept of the present invention fall within the protection scope of the present invention. Unless otherwise specified, the reagents and instruments used in the following embodiments are commercially available products. Unless otherwise specified, the experimental methods used are conventional methods.

[0074] Example 1

[0075] Solution A: Dissolve 13.5 mg of ferric chloride hexahydrate in 10 mL of an aqueous solution containing 0.2 g of polyvinylpyrrolidone (PVP) with a molecular weight of 30 KD.

[0076] Solution B: Dissolve 34 mg of ellagic acid in 10 mL of 22.5 mM NaOH aqueous solution containing 0.2 g PVP.

[0077] Solution B was added dropwise to solution A, and the mixture was stirred vigorously for 10 minutes. Then, 0.5 g of PVP was dissolved in the solution, and the mixture was stirred for 20 minutes. The mixture was then transferred to a Teflon autoclave and heated at 130°C for 4 hours. The resulting precipitate was centrifuged, washed three times with water and absolute ethanol, and redispersed in 10 mL of water to form a Fe-MOF suspension C.

[0078] Solution D: Dissolve 0.28 g acrylamide and 0.02 g N,N-methylenebisacrylamide in 2 mL of phosphate buffered saline (PBS, pH 6.0) and stir gently for 10 minutes. Then, add 0.2 mL of 5 μM zinc finger peptide aqueous solution and continue stirring.

[0079] Solution E: Disperse 0.3 g of ammonium persulfate and 0.2 mL of tetramethylethylenediamine into 2 mL of water to prepare solution E, which serves as a coagulant.

[0080] Add 10 μL of suspension C and 20 μL of solution E to solution A and stir for 2 minutes. Then store the precursor at 4°C for later use.

[0081] Building a sensor platform:

[0082] A 0.2% (w / w) aqueous solution of sodium alginate (SA) was mixed with a 0.2% (w / w) suspension of graphene oxide (GO), and diluted with water to 8 mL. The mixture was stirred vigorously for 30 minutes. The suspension was stored at 4°C.

[0083] SA-GO suspension was added dropwise to the surface of a glassy carbon electrode and dried at 37°C. An aqueous solution of nickel nitrate was then added dropwise and incubated to obtain sodium alginate-Ni. 2+ - Graphene oxide (SA-Ni) 2+ -GO) gel layer. Gently rinse the GCE with water to remove unreacted solution, then drop the prepared dual-response hydrogel precursor suspension onto the surface and dry at 37°C to assemble the electrochemical sensor platform.

[0084] The specific testing steps are as follows:

[0085] 10 μL of immunomagnetic bead probe suspension, 10 μL of GOx@ZIF-8-Au-Ab2 immunoprobe suspension, and 10 μL of aqueous solutions containing different concentrations of ProGRP were mixed in a centrifuge tube and then sonicated for 25 minutes. After magnetic separation and washing three times with PBS (pH=7.4), 50 μL of 50 mM glucose aqueous solution was added to the centrifuge tube and mixed, and then incubated at room temperature for 70 minutes.

[0086] Drop 5 μL of the supernatant from the centrifuge tube onto the prepared sensor platform and incubate at room temperature for 7 minutes. Gently rinse the electrode surface with PBS (pH 7.4) to remove unreacted solution. Perform square wave voltammetry (SWV) tests using a three-electrode system in PBS (pH 7.4).

[0087] Figure 1 The image shown is a scanning electron microscope image of the dual-response hydrogel prepared in Example 1. It can be seen that the surface of the hydrogel exhibits a porous structure after drying.

[0088] Figure 2 This is a photograph of the Fe-MOF prepared in Example 1. The Fe-MOF structure presents as rhomboid sheets with a length of approximately 1 μm.

[0089] Figure 3 The X-ray photoelectron spectrum of the dual-response hydrogel obtained in Example 1 shows peaks corresponding to the elements, indicating that the sample contains the expected elements. As can be seen from the figure, the material obtained according to the preparation method of the present invention contains C, O, and Fe elements.

[0090] Figure 5 The current response of the dual-response hydrogel prepared in Example 1 was observed in response to changes in (A) hydrogen peroxide concentration, (B) zinc ion concentration, and (C) mixtures of hydrogen peroxide and zinc nitrate in groups 1, 2, 3, 4, 5, 6, 7, and 8, where the hydrogen peroxide concentrations were 0.5, 1, 1.5, 2, 2.5, 3, 3.5, and 4 mM, and the corresponding zinc ion concentrations were 10, 30, 50, 70, 90, 110, 130, and 150 μM. It can be found that... Figure 5 When hydrogen peroxide and zinc ions are applied simultaneously, the signal intensity and sensitivity are significantly higher than when hydrogen peroxide or zinc ions are applied alone, demonstrating the dual-response performance of the hydrogel.

[0091] Example 2

[0092] Except that the volumes of suspension C, solution D, and solution E are in a ratio of 2:10:3, the dual-response hydrogel material was prepared in the same manner as in Example 1. Furthermore, the sensor platform construction method and specific testing procedures were also performed in the same manner as in Example 1.

[0093] Figure 4 The image shown is a scanning electron microscope image of the dried hydrogel prepared in Example 2. The surface is relatively smooth and the pore structure is not obvious.

[0094] Application examples

[0095] The zinc-doped nickel hydroxyl oxide modified electrode obtained in Example 1 was used to detect the concentration of the tumor marker progastrin-releasing peptide (ProGRP). The specific steps included:

[0096] 1) Preparation of GOx@ZIF-8-Au-Ab2: 10 mL of 20 mM Zn(NO)32-6H2O and 6 mg glucose oxidase (GOx) were added to 10 mL of 1.4 M 2-methylimidazole aqueous solution and stirred overnight. The resulting GOx@ZIF-8 nanoparticles were then centrifuged, washed three times with water, and redispersed in 2 mL of water. 1 mL of 0.04 wt% NaAuCl4 aqueous solution was added to 1 mL of the GOx@ZIF-8 suspension and stirred for 15 minutes. Then, 200 μL of 0.3 mg mL-1 NaBH4 (4 °C) was added to the suspension and gently stirred for 1 hour. GOx@ZIF-8 with gold nanoparticles on the surface was obtained by in-situ reduction. GOx@ZIF-8-Au was centrifuged, washed four times with water, and then dispersed in 2 mL of water. 100 μL of 1 mg mL-1 NaBH4 was added to 1 mL of GOx@ZIF-8-Ab2. -1 The ProGRP-labeled antibody (Ab2) was incubated with 2 mL of the above suspension at 4 °C for 12 hours with stirring. Then, 2 mL of 1 wt% BSA was added to the suspension to block the remaining active sites of Au. After centrifuging and washing four times with water, the precipitate was collected and redispersed in 2 mL of water.

[0097] 2) Preparation of immunomagnetic bead probes: First, add 1 mL of 5 mg / mL immunomagnetic bead probes. -1 The suspension of carboxylated magnetic particles was shaken at 37°C for 30 minutes with 1 mL of a mixed aqueous solution containing 0.4 M 1-ethyl-3'-dimethylaminopropylcarbodiimide (EDC) and 0.1 M N-hydroxysuccinimide (NHS). Then, the above suspension was mixed with 1 mL of 200 μg mL... -1 The ProGRP-coated antibody (Ab1) was mixed and incubated at 37°C for 1 hour. Next, 1 mL of 1 wt% BSA aqueous solution was added to the suspension as an inhibitor, and incubated for 30 minutes. Finally, the obtained immunomagnetic bead probes were redispersed in 1 mL of PBS (pH 7.4) and stored at 4°C.

[0098] 3) Detection in the immunosensor: The sensor platform prepared in Example 1 was used for the experiment. 10 μL of immunomagnetic bead probe suspension, 10 μL of GOx@ZIF-8-Au-Ab2 immunosensor suspension, and 10 μL of aqueous solutions containing different concentrations of ProGRP were mixed in a centrifuge tube and then sonicated for 25 minutes. After magnetic separation and washing three times with PBS (pH=7.4), 50 μL of 50 mM glucose aqueous solution was added to the centrifuge tube and mixed, then incubated at room temperature for 70 minutes. 5 μL of the supernatant from the centrifuge tube was dropped onto the immunosensor platform and kept at room temperature for 7 minutes. The electrode surface was gently rinsed with PBS (pH=7.4) to remove unreacted solution. Square wave voltammetry (SWV) was performed using a three-electrode system in PBS (pH=7.4). The electrochemical signal was measured by varying the added antigen concentration, and a standard curve of electrochemical signal versus concentration was plotted, as shown in the figure. Figure 6 As shown, the sensor's linear range is from 100 fg to 100 ng, with a detection limit of 14.24 fg, and the standard curve equation is y = 0.8389x + 13.67. This demonstrates that the sensor can achieve sensitive detection of ProGRP.

[0099] The above description is merely a specific embodiment of the present invention, but the scope of protection of the present invention is not limited thereto. Any variations or substitutions that can be easily conceived by those skilled in the art within the technical scope disclosed in the present invention should be included within the scope of protection of the present invention. Therefore, the scope of protection of the present invention should be determined by the scope of the claims.

Claims

1. A method for preparing a hydrogel precursor that is dual-responsive to hydrogen peroxide and zinc ions, the method comprising the following steps: (1) Preparation of Fe-MOF suspension Iron salt is dissolved in a surfactant solution to form solution A; The polyphenols were dissolved in an alkaline solution containing a surfactant to form solution B. Then, solutions A and B are mixed and stirred, and a high-temperature hydrothermal reaction is carried out to form Fe-MOF. The precipitate is centrifuged, washed with water and absolute ethanol, and then redispersed in water to form a suspension C of Fe-MOF. The iron salt is selected from at least one of ferric sulfate, ferric nitrate, and ferric chloride hexahydrate; the solvent in solution A is selected from at least one of deionized water, ethanol, and methanol; the molar concentration of the iron salt in solution A is 0.01~0.1 mol / L; the surfactant is selected from at least one of polyvinylpyrrolidone with a molecular weight of 25KD-40KD, quaternized polyacrylamide with a molecular weight of 20KD-50KD, polyacrylic acid with a molecular weight of 2KD-5KD, and polyethylene oxide with a molecular weight of 20KD-50KD; the polyphenol is ellagic acid; and the mass ratio of the iron salt to the surfactant is 0.03:1~0.1:

1. The mass ratio of the polyphenol to the surfactant in solution B is 0.02:1 to 0.1:1; The alkali in solution B is selected from at least one of sodium hydroxide, potassium hydroxide, lithium hydroxide, and ammonia water; the molar concentration of the alkali is 20~50mM; The solvent in solution B is selected from at least one of deionized water, ethanol, and methanol; The volume ratio of solution A to solution B is 0.1:1 - 2.0:

1. The high-temperature hydrothermal reaction temperature is between 100-180℃; The hydrothermal reaction time is between 2 and 8 hours; The mass concentration of Fe-MOF in the Fe-MOF suspension C is between 10 and 50 g / L; (2) Dissolve acrylamide and N,N-methylenebisacrylamide in a buffer solution, add 0.5µM zinc finger peptide aqueous solution, and stir continuously to form solution D; disperse the oxidant and tetramethylethylenediamine in water to form solution E; The pH of the buffer solution is 3.0~8.0; The buffer solution is selected from phosphate buffer, carbonate buffer, citrate buffer or acetate buffer; In solution D, the mass ratio of acrylamide to N,N-methylenebisacrylamide is 10:1 to 20:

1. In solution D, the mass concentration of the acrylamide is between 100 g and 300 g / L; In solution E, the oxidant is selected from at least one of hydrogen peroxide, ammonium persulfate, potassium permanganate, and peracetic acid; The mass concentration of the oxidant is 50~200g / L; In solution E, the volume ratio of tetramethylethylenediamine to water is between 0.01:1 and 0.1:

1. (3) Mix the solutions D and E obtained in step (2) with the Fe-MOF suspension C obtained in step (1) to obtain the hydrogel precursor; The volume ratio of the suspension C, solution D, and solution E is between 0.05:10:1 and 2:10:

5.

2. The preparation method according to claim 1, characterized in that, In step (1), The iron salt is ferric chloride hexahydrate; The surfactant is polyvinylpyrrolidone with a molecular weight of 25-40 KD; The mass ratio of the iron salt to the surfactant is 0.065:1; In solution B, the mass ratio of the polyphenol to the surfactant is 0.068:1; The base in solution B is sodium hydroxide; the molar concentration of the base is 22.5 mM. The solvent in solution B is deionized water; The volume ratio of solution A to solution B is 1:

1. The high-temperature hydrothermal reaction temperature is 130℃; The hydrothermal reaction time is 4 hours; The mass concentration of Fe-MOF in the Fe-MOF suspension C is 20 g / L; In step (2): The pH of the buffer solution is 6.0; The buffer solution is phosphate buffer solution (PBS); In solution D, the mass ratio of acrylamide to N,N-methylenebisacrylamide is 14:1; In solution D, the mass concentration of the acrylamide is 140 g / L; In solution E, the oxidant is ammonium persulfate; The mass concentration of the oxidant is 100 g / L; In solution E, the volume ratio of tetramethylethylenediamine to water is 0.05:1; In step (3) The volume ratio of the suspension C, solution D, and solution E is 1:10:

2.

3. A hydrogel precursor that is responsive to both hydrogen peroxide and zinc ions, prepared by the preparation method according to claim 1 or 2.

4. A hydrogel formed by drying the hydrogel precursor according to claim 3.

5. A method for preparing a hydrogel, comprising drying a hydrogel precursor prepared according to the method of claim 1 or 2 to obtain a hydrogel, wherein the drying temperature is between 25 and 50°C and the corresponding time is 15 min.

6. The method for preparing the hydrogel according to claim 5, wherein the drying temperature is 37°C.

7. An electrochemical sensor platform, the electrochemical sensor platform comprising an electrode, a sodium alginate-nickel ion-graphene oxide gel layer and a hydrogel layer formed from the hydrogel according to claim 4.

8. The method for preparing the electrochemical sensor platform according to claim 7, comprising: First, the electrode surface is cleaned, then a sodium alginate-nickel ion-graphene oxide gel layer is formed on the electrode surface, and then the hydrogel precursor prepared according to the preparation method of claim 1 or 2 is drop-coated onto the electrode surface and dried to obtain the electrochemical sensor platform.

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