Electrochemical biosensor for detecting IgG in blood and construction method thereof

Abstract

The invention provides an electrochemical biosensor for detecting IgG in blood and a construction method thereof, and belongs to the technical field of electrochemical biosensors, and the construction method comprises the following steps: mixing aniline, perchloric acid and water, adding polystyrolsulfon acid to obtain an electro-deposition solution, then immersing a glassy carbon electrode into the electro-deposition solution, and carrying out ultrasonic treatment; a polyaniline / glassy carbon electrode is obtained through chronopotentiometry electro-deposition; chloroauric acid trihydrate, sodium chloride and water are mixed to obtain a mixed aqueous solution, the polyaniline / glassy carbon electrode is immersed in the mixed aqueous solution, a gold nanoparticle / polyaniline / glassy carbon electrode is obtained through cyclic voltammetry electrodeposition, a lasso peptide solution is dropwise added to the working surface of the gold nanoparticle / polyaniline / glassy carbon electrode for incubation, and the lasso peptide / gold nanoparticle / polyaniline / glassy carbon electrode is obtained. And constructing to obtain the electrochemical biosensor. By means of the overall hydrophilicity and electric neutrality of the polypeptide, non-specific protein adsorption can be effectively resisted, the detection sensitivity and accuracy are improved, and ultra-low pollution immunoglobulin G detection in blood is achieved.

Description

Technical Field

[0001] This invention relates to the field of electrochemical biosensor technology, and in particular to an electrochemical biosensor for detecting IgG in blood and its construction method. Background Technology

[0002] Immunoglobulin G (IgG) is one of the key immunoglobulins in the human body, forming an important immune defense system along with IgA, IgM, IgD, and IgE. It not only effectively neutralizes pathogens and enhances the phagocytic capacity of phagocytes, but also possesses the unique property of crossing the placenta, allowing it to be passed from mother to infant and providing passive immune protection for up to six months. It is a core substance for maintaining human immune function and ensuring early immunization safety in infants and young children. Furthermore, changes in IgG levels are closely related to human health. When the body lacks IgG, problems such as decreased immune function (e.g., poor vaccine efficacy, increased risk of autoimmune diseases), recurrent infections (e.g., respiratory, ear, and gastrointestinal infections), and delayed growth and development in children may occur. Therefore, accurate detection of IgG in the blood is of great significance for disease diagnosis, immune status assessment, and clinical treatment guidance, and has become an important research direction in the field of bioassay.

[0003] Currently, the detection of IgG in blood mainly relies on two common methods: enzyme-linked immunosorbent assay (ELISA) and immunofluorescence assay (IFA). ELISA, with its high specificity, is widely used in clinical testing, achieving quantitative detection of IgG through a series of steps including sample processing, antibody coating, washing, and substrate color development. IFA, based on fluorescence labeling technology, uses fluorescence microscopy to observe fluorescence intensity and distribution, completing qualitative or semi-quantitative analysis of IgG. Furthermore, electrochemical biosensors, due to their advantages of simple operation, rapid response, and high sensitivity, show broad application prospects in the field of biomarker detection and are gradually becoming an important area of ​​research in IgG detection technology.

[0004] However, existing detection methods and technologies still have significant shortcomings, making it difficult to meet the demands for high efficiency and accuracy in clinical testing. On the one hand, the ELISA procedure is complex and time-consuming, requiring precise control at every step, including sample processing and antibody coating. The entire testing process often takes several hours, resulting in low efficiency. Furthermore, IFA results are highly dependent on the subjective judgment of the testing personnel, with significant differences in interpretation of fluorescence intensity and distribution among different individuals, easily leading to result deviations. On the other hand, although electrochemical biosensors have significant advantages, in the complex environment of blood, their sensing interface is susceptible to non-specific adsorption of interfering proteins—these proteins adhere indiscriminately to the sensing interface through hydrophobic interactions or electrostatic attraction, severely affecting detection sensitivity and accuracy and limiting their practical application.

[0005] In addition, current anti-fouling materials used to address non-specific adsorption (such as polyethylene glycol and zwitterionic polymers) also have shortcomings: polyethylene glycol is easily oxidized and enzymatically degraded under physiological conditions and cannot be used for a long time; zwitterionic polymers, due to their high surface charge density, are easily affected by the electric field during detection and undergo deswelling, which in turn damages the sensor performance.

[0006] Therefore, there is an urgent need for an electrochemical biosensor that is highly sensitive, low-pollution, highly stable, and suitable for detecting IgG in blood. Summary of the Invention

[0007] To overcome the shortcomings of existing technologies, the purpose of this invention is to provide an electrochemical biosensor for detecting IgG in blood and its construction method. By utilizing the overall hydrophilicity and electroneutrality of polypeptides, it can effectively resist non-specific protein adsorption, improve detection sensitivity and accuracy, and achieve ultra-low pollution immunoglobulin G detection in blood.

[0008] To achieve the above objectives, the present invention provides the following solution: On one hand, the present invention provides a method for constructing an electrochemical biosensor for detecting IgG in blood, comprising the following steps: S1. Aniline, perchloric acid and water are mixed, and polystyrene sulfonic acid is added to the mixture to obtain an electrodeposition solution. Then, a glassy carbon electrode is immersed in the electrodeposition solution and electrodeposition is performed by chronopotential method to obtain a polyaniline / glassy carbon electrode. S2. Chloroauric acid trihydrate, sodium chloride and water are mixed to obtain a mixed aqueous solution, and the polyaniline / glassy carbon electrode is immersed in the mixed aqueous solution. Gold nanoparticles / polyaniline / glassy carbon electrode is obtained by cyclic voltammetry electrodeposition. S3. Dissolve the lasso peptide in water to prepare a lasso peptide solution, and drop the lasso peptide solution onto the working surface of the gold nanoparticle / polyaniline / glassy carbon electrode for incubation to obtain the lasso peptide / gold nanoparticle / polyaniline / glassy carbon electrode, thus constructing an electrochemical biosensor for detecting IgG in blood.

[0009] Preferably, in S1, the concentration of aniline in the electrodeposition solution is 0.1 M, and the concentration of perchloric acid is 0.03 M; the current density of the chronopotentiometric method is 10 μA·cm. -2 The electrodeposition time was 1 hour.

[0010] Preferably, in S2, the concentration of chloroauric acid trihydrate in the mixed aqueous solution is 0.1-5.0 mM, and the concentration of sodium chloride is 0.1-1.0 M; the low potential of electrodeposition in the cyclic voltammetry is -0.1-0.5 V, the high potential is 0.1-1.0 V, the scan rate is 10-100 mV / s, and the number of scans is 2-10.

[0011] Preferably, in S3, the concentration of the lazopeptide solution is 1 μM.

[0012] Preferably, in S3, the drop volume of the lazopeptide solution is 10-60 μL, and the incubation time is 6-12 h.

[0013] Preferably, the amino acid sequence of the latissimus dorsi peptide is C(KEKEKE)EK(AVWGRWH)ECPPPPC, and satisfies the following structural characteristics: The first amino acid, cysteine, to the seventh amino acid, glutamic acid, form a ring structure; The cysteine ​​in the ring structure extends into a branch chain with the amino acid sequence EKECPPPPC. This branch chain extends downward from the top of the ring through the internal space of the ring. The ninth amino acid, lysine, is linked to a recognition branch with the amino acid sequence AVWGRWH. The first amino acid, cysteine, is linked to the eleventh amino acid, cysteine, by a disulfide bond. All amino acids in the lasso peptide are D-type amino acids, and the overall configuration is lasso-like.

[0014] On the other hand, the present invention also provides an electrochemical biosensor for detecting IgG in blood, which is prepared using the above-described method for constructing an electrochemical biosensor for detecting IgG in blood.

[0015] Based on the above, the detection method for the electrochemical biosensor used to detect IgG in blood includes the following steps: Using the electrochemical biosensor as the working electrode, a platinum wire as the counter electrode, and a silver / silver chloride electrode as the reference electrode, a three-electrode system was formed. A standard immunoglobulin G solution was dropped onto the surface of the working electrode, and then the three-electrode system was immersed in a buffer solution with a pH of 7-8. The change in current signal was measured by an electrochemical workstation, and a standard working curve was plotted based on the correspondence between the concentration of standard immunoglobulin G and the current signal. Take the same volume of the blood sample to be tested and drop it onto the surface of the working electrode. Assemble a three-electrode system under the same conditions as the above steps and measure the change in current signal. Calculate the concentration of immunoglobulin G in the blood sample to be tested based on the standard working curve.

[0016] Preferably, the detection of the target immunoglobulin G employs a ratiometric electrochemical signal mode, utilizing the hydrophilicity and electroneutrality of the lasso peptide to achieve ultra-low contamination detection in the blood.

[0017] Preferably, the pH 7-8 buffer solution is a 0.2M phosphate buffer solution.

[0018] According to specific embodiments provided by the present invention, the present invention discloses the following technical effects: (1) The electrochemical biosensor provided by the present invention is based on a polyaniline coating, which can effectively block the direct contact of interfering proteins on the sensing interface and provide a certain ability to accelerate electron transfer, which is beneficial to enhance the anti-fouling performance and some sensitivity of the electrochemical biosensor; at the same time, based on gold nanoparticles, the nanoscale spherical structure can effectively increase the electroactive area and accelerate the electron transfer process, which is beneficial to enhance the sensitivity of the electrochemical biosensor; in addition, based on lasso peptides, the electrochemical biosensor has high hydrophilicity and electroneutrality as well as excellent anti-fouling performance and biocompatibility. Its three-dimensional spatial configuration can effectively resist the adhesion of non-specific proteins. The gold nanoparticles and polyaniline coating accelerate the transfer of electrons and ion transport, which can realize low-fouling, high-sensitivity and accurate detection of immunoglobulin G in blood.

[0019] (2) The electrochemical biosensor provided by the present invention can detect immunoglobulin G in blood. It can achieve sensitive and accurate collection and analysis of information by taking a small amount of blood. The detection time is short, the operation is easy, the detection range is wide, the detection limit is low, the sensitivity is high, and the accuracy is high. It is an efficient and simple detection method for immunoglobulin G.

[0020] (3) The electrochemical biosensor provided by the present invention has a detection range of 0.1 ng / mL to 1 mg / ml for the target immunoglobulin G, a linear equation of ΔI = 6.67 LgC + 81.70, a linear correlation coefficient of 0.996, and a limit of detection of 0.03 ng / ml. Attached Figure Description

[0021] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0022] Figure 1 This is a flowchart illustrating the construction method of an electrochemical biosensor for detecting IgG in blood according to the present invention. Figure 2 This is a scanning electron microscope image of electrodeposited polyaniline and gold nanoparticles in Example 1 of the present invention; Figure 3 This is a static water contact angle diagram of the Lasso / AuNPs / PANI modified electrode in Embodiment 1 of the present invention; Figure 4 This is a comparison diagram of the zeta potentials of lasso peptide, PEG, and branched peptide in Example 1 of the present invention; Figure 5 This is a circular dichroism chromatogram of the latissimus dorsi peptide in Example 1 of the present invention; Figure 6 This is a feasibility analysis diagram from Embodiment 1 of the present invention; Figure 7 This is a current response diagram of the Lasso / AuNPs / PANI modified electrode in Example 1 of the present invention to different dilution concentrations of fetal bovine serum; Figure 8 This is a comparison of the signal change rate of electrodes with different modifications in Example 1 of the present invention after being immersed in 10% FBS solution for half an hour; Figure 9 This is a comparison of the signal change rate of electrodes with different modifications in Example 1 of the present invention after being immersed in 20% FBS solution for half an hour; Figure 10 This is a comparison chart of the signal change rate of electrodes with different modifications in Example 1 of the present invention after being immersed in 100% FBS solution for half an hour; Figure 11 The current signal diagram shows the detection of different concentrations of immunoglobulin G by an electrochemical biosensor based on lazopeptide in PBS and FBS solutions. Figure 12 This is a graph showing the current signal of a dendritic peptide-based electrochemical biosensor detecting different concentrations of immunoglobulin G in PBS and FBS solutions. Detailed Implementation

[0023] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0024] To make the above-mentioned objects, features and advantages of the present invention more apparent and understandable, the present invention will be further described in detail below with reference to the accompanying drawings and specific embodiments.

[0025] The electrochemical biosensor based on a lasso peptide / gold nanoparticles / polyaniline / glassy carbon electrode provided by this invention exhibits high hydrophilicity and electroneutrality. The high hydrophilicity allows for the binding of more free water molecules on the sensor surface, forming a dense hydration layer that resists non-specific protein adsorption, thus preventing interference proteins from affecting the sensor's operation. Furthermore, the fabricated electrochemical biosensor has a biosensing interface with good electroneutrality, effectively preventing electrostatic attraction of charged interfering substances at the sensing interface. This results in excellent anti-fouling performance, enabling highly accurate, stable, and sensitive detection of immunoglobulin G.

[0026] like Figure 1 As shown, this invention provides a method for constructing an electrochemical biosensor for detecting IgG in blood, comprising the following steps: S1. Aniline, perchloric acid and water are mixed, and polystyrene sulfonic acid is added to the mixture to obtain an electrodeposition solution. Then, a glassy carbon electrode is immersed in the electrodeposition solution and electrodeposition is performed by chronopotential method to obtain a polyaniline / glassy carbon electrode.

[0027] The electrodeposition solution contained 0.1 M aniline, 0.03 M perchloric acid, and 0.01 g of polystyrene sulfonic acid (PSS); the chronopotentiometric current density was 10 μA·cm⁻¹. -2 The electrodeposition time was 1 hour, and a polyaniline / glassy carbon electrode PANI / GCE was obtained by electrodeposition.

[0028] S2. Chloroauric acid trihydrate, sodium chloride and water are mixed to obtain a mixed aqueous solution, and the polyaniline / glassy carbon electrode is immersed in the mixed aqueous solution. Gold nanoparticles / polyaniline / glassy carbon electrode is obtained by cyclic voltammetry electrodeposition.

[0029] In the mixed aqueous solution, the concentration of chloroauric acid trihydrate HAuCl4·3H2O was 0.1-5.0 mM, and the concentration of sodium chloride NaCl was 0.1-1.0 M. In the cyclic voltammetry method, the low potential for electrodeposition was -0.1-0.5 V, the high potential was 0.1-1.0 V, the scan rate was 10-100 mV / s, and the number of scan cycles was 2-10. Gold nanoparticles / polyaniline / glassy carbon electrodes AuNPs / PANI / GCE were obtained by electrodeposition.

[0030] S3. Dissolve the lasso peptide in water to prepare a lasso peptide solution, and drop the lasso peptide solution onto the working surface of the gold nanoparticle / polyaniline / glassy carbon electrode for incubation to obtain the lasso peptide / gold nanoparticle / polyaniline / glassy carbon electrode, thus constructing an electrochemical biosensor for detecting IgG in blood.

[0031] The concentration of the lasso peptide solution is 1 μM, the drop volume of the lasso peptide solution is 10-60 μL, and the incubation time is 6-12 h, thereby constructing a lasso peptide / gold nanoparticle / polyaniline / glassy carbon electrode Lasso / AuNPs / PANI / GCE.

[0032] Based on the above, the amino acid sequence of the lasso peptide in step S3 is C(KEKEKE)EK(AVWGRWH)ECPPPPC, and it satisfies the following structural characteristics: The first amino acid, cysteine, to the seventh amino acid, glutamic acid, form a ring structure. A branch chain extends from the cysteine ​​in the ring structure, with the amino acid sequence EKECPPPPC. This branch chain extends downwards from the top of the ring through the internal space, forming a lasso configuration. Simultaneously, the ninth amino acid, lysine, is connected to a recognition branch chain with the amino acid sequence AVWGRWH. The first amino acid, cysteine, and the eleventh amino acid, cysteine, are linked by a disulfide bond. Furthermore, all amino acids in the lasso peptide are D-type amino acids, and the overall configuration is lasso-like. This configuration combines the excellent antifouling effect of the cyclic peptide with the recognition of target analytes through the AVWGRWH amino acid sequence. Additionally, the high steric hindrance of the structure also enhances the antifouling performance of the lasso peptide.

[0033] Based on the above, the detection method for the electrochemical biosensor used to detect IgG in blood includes: A three-electrode system was constructed using a Lasso / AuNPs / PANI / GCE electrode as the working electrode, a platinum wire as the counter electrode, and a silver / silver chloride electrode as the reference electrode. A standard immunoglobulin G solution was added dropwise to the surface of the working electrode, and the three-electrode system was then immersed in a 0.2 M phosphate buffer solution at pH 7-8. Changes in the current signal were measured using an electrochemical workstation, and a standard working curve was plotted based on the correlation between the concentration of standard immunoglobulin G and the current signal. Take the same volume of the blood sample to be tested and drop it onto the surface of the working electrode. Assemble a three-electrode system under the same conditions as the above steps and measure the change in current signal. Calculate the concentration of immunoglobulin G in the blood sample to be tested based on the standard working curve.

[0034] Among them, the detection of target immunoglobulin G adopts a ratiometric electrochemical signal mode, which utilizes the hydrophilicity and electroneutrality of lasso peptides to achieve ultra-low contamination detection in blood.

[0035] The above content will be further described below through specific implementation methods. The described embodiments are only some embodiments of the present invention, and the experimental methods in the embodiments without specific conditions are usually determined in accordance with national standards; the experimental instruments, equipment and reagents without source are all commercially available raw materials.

[0036] In addition, in this embodiment, the current signal was recorded using a Shanghai Chenhua CHI 660E electrochemical workstation. The current signal was tested on a conventional three-electrode electrochemical system, using a conventional glassy carbon electrode (GCE) as the working electrode, a platinum wire as the counter electrode, and silver / silver chloride as the reference electrode, forming a three-electrode system purchased from Shanghai Chenhua. Lasso peptide was purchased from Hangzhou Zhuntai. Other chemical reagents and materials were commercially available products purchased from Sigma-Aldrich.

[0037] Example 1 This embodiment provides a method for constructing an electrochemical biosensor for detecting IgG in blood, including: First, aniline, perchloric acid solution, and water were mixed, and then 0.01 g of Pss was added, where the concentration of aniline was 0.1 M and the concentration of perchloric acid was 0.03 M. The mixture was stirred until homogeneous to obtain a polyaniline deposition solution. Then, 5 ml of the polyaniline deposition solution was placed in a small beaker, and a glassy carbon electrode was immersed in the solution. A chronopotentiometric method was used, with the current density set to 10 μA·cm⁻¹. -2 The electrodeposition time was 1 hour, and PANI / GCE was finally obtained.

[0038] Next, chloroauric acid trihydrate HAuCl4·3H2O, NaCl, and water were mixed, with the concentration of chloroauric acid trihydrate HAuCl4·3H2O being 1 mM and the concentration of NaCl being 0.5 M. The mixture was stirred evenly to obtain a gold nanoparticle deposition solution. Subsequently, 5 ml of the gold nanoparticle deposition solution was placed in a small beaker, and PANI / GCE was immersed in the deposition solution. By cyclic voltammetry, with the potential range set to -0.9 to 0.5 V and the scan rate at 50 mV / s, deposition was carried out for 5 cycles to prepare AuNPs / PANI / GCE.

[0039] Finally, lasso peptide and water were mixed, with the lasso peptide concentration being 1 μm / L. 50 μL of the 1 μm / L lasso peptide solution was dropped onto the working surface of AuNPs / PANI / GCE and then incubated in a refrigerator at 4°C for 6-12 h to obtain the Lasso / AuNPs / PANI / GCE electrochemical biosensor.

[0040] like Figure 2 As shown, a scanning electron microscope (SEM) image of the electrodeposited gold nanoparticles in this embodiment is provided. It can be seen that aniline is uniformly aggregated on the glassy carbon electrode, while the gold nanoparticles are densely attached to the electrode surface. The adhesion of the polyaniline coating and gold nanoparticles effectively increases the electroactive area and enhances electron transfer and ion transport capabilities. The water contact angle of the lasso-modified electrochemical biosensor surface is shown in the figure. Figure 3As shown, its static water contact angle is 20.5°, indicating that the modified interface has high hydrophilicity. The zeta potential data for the lazopeptide are shown below. Figure 4 As shown, the zeta potential of the lasso peptide is around -7 mV, indicating excellent electroneutrality of the material. Furthermore, the circular dichroism chromatographic data of the lasso peptide are as follows: Figure 5 As shown in the figure, there is a large negative peak at 200 nm, which indicates that the tau peptide has formed an α-helix secondary structure, which is in line with the design expectation.

[0041] To further verify the antifouling capability of the electrochemical biosensor provided in this embodiment, the DPV data of Bare GCE, PANI / GCE, AuNPs / PANI / GCE, Lasso / AuNPs / PANI / GCE, and immunoglobulin G / Lasso / AuNPs / PANI / GCE of the same electrode were first measured to obtain... Figure 6 The feasibility analysis diagram is shown below. From the feasibility analysis, the decrease in DPV signal after polyaniline deposition on the bare electrode is because, although polyaniline is a conductive polymer, its conductivity is not as good as that of the bare electrode. The decrease in electrical signal after gold nanoparticle deposition is because, during the deposition of gold nanoparticles, a large number of gold nanoparticles were deposited to provide more binding sites for the lauryl peptide, thus affecting electron transfer. The reasons for the decrease in signal peaks generated by self-assembled peptides and recognition of immunoglobulin G are consistent with those for the decrease after polyaniline deposition. Therefore, from the feasibility analysis diagram, it can be seen that the electrochemical biosensor provided in this embodiment has excellent anti-fouling capabilities.

[0042] In addition, the electrochemical biosensor constructed in this embodiment was immersed in fetal bovine serum buffer solutions of different concentrations, and the current signal before and after immersion in the fetal bovine serum buffer solutions of different concentrations for half an hour was tested.

[0043] The different concentrations of fetal bovine serum (FBS) buffer were achieved by adding different volumes of FBS to water to dilute the FBS to different concentrations: 1%, 10%, 20%, 50%, and 100% (pure FBS, anhydrous). FBS contains abundant proteins and cells, and can simulate 40% of the human serum environment. The test results are as follows... Figure 7 As shown, the electrochemical biosensor provided in this embodiment exhibits minimal signal changes after immersion in fetal bovine serum buffer solutions of different concentrations, effectively enhancing analytical accuracy and avoiding false positive results.

[0044] Furthermore, to further confirm the high anti-fouling performance of the electrochemical biosensor provided in this embodiment, electrodes modified with PEG and branched peptides were used as controls. The signal change rate of the three electrodes was measured after immersion in 10%, 20%, and 100% FBS for half an hour, respectively. The final results are as follows: Figure 8 , Figure 9 , Figure 10 As shown, from Figures 8 to 10 The results showed that the electrodes modified with lasso peptides exhibited smaller signal change rates when exposed to different concentrations of FBS. In high-concentration FBS solutions, the electrodes modified with lasso peptides performed significantly better than those modified with branched peptides and PEG. This indicates that lasso peptides have superior antifouling capabilities compared to branched peptides and PEG.

[0045] The changes in current signal detected by the electrochemical biosensor provided in this embodiment for different concentrations (0.1 ng / mL - 1 mg / mL) of the target immunoglobulin G are as follows: Figure 11 As shown in the figure, the detection range of the lassotide-modified electrode for immunoglobulin G is 0.1 ng / mL to 1 mg / mL, with linear equations of ΔI = 6.67 LgC + 81.70 (red line) and ΔI = 7.24 LgC + 85.60 (blue line), and a limit of detection of 0.03 ng / mL. To ensure the capability of the lassotide-modified electrode, a branched peptide-modified electrode was used as a reference, and its detection capability for different concentrations of immunoglobulin G was measured. The results are shown below. Figure 12 As shown, the red line ΔI = 6.85LgC + 77.94, and the blue line ΔI = 5.20LgC + 59.07. From... Figure 11 and Figure 12 The data shows that the electrochemical biosensor provided in this embodiment has high sensitivity for the detection of immunoglobulin G, which is attributed to the PANI coating on the electrode substrate and the good conductivity of AuNPs, which improves the conductivity.

[0046] Therefore, the electrochemical biosensor for detecting IgG in blood described above, and its construction method, utilize the overall hydrophilicity and electroneutrality of the polypeptide to effectively resist non-specific protein adsorption, improve detection sensitivity and accuracy, and achieve ultra-low pollution immunoglobulin G detection in blood.

[0047] This document uses specific examples to illustrate the principles and implementation methods of the present invention. The descriptions of the above embodiments are only for the purpose of helping to understand the method and core ideas of the present invention. Furthermore, those skilled in the art will recognize that, based on the ideas of the present invention, there will be changes in the specific implementation methods and application scope. Therefore, the content of this specification should not be construed as a limitation of the present invention.

Claims

1. A method of constructing an electrochemical biosensor for detecting IgG in blood, characterized by, The method comprises the following steps: S1, mixing aniline, perchloric acid and water, and adding polystyrene sulfonic acid into the mixture to obtain an electrodeposition solution, then immersing a glassy carbon electrode into the electrodeposition solution, and performing electrodeposition by chronopotentiometry to obtain a polyaniline / glassy carbon electrode; S2, mixing chloroauric acid trihydrate and sodium chloride with water to obtain a mixed aqueous solution, and immersing the polyaniline / glassy carbon electrode into the mixed aqueous solution, and performing electrodeposition by cyclic voltammetry to obtain a gold nanoparticle / polyaniline / glassy carbon electrode; S3, dissolving a lariat peptide in water to prepare a lariat peptide solution, and adding the lariat peptide solution dropwise to a working surface of the gold nanoparticle / polyaniline / glassy carbon electrode for incubation to obtain a lariat peptide / gold nanoparticle / polyaniline / glassy carbon electrode, i.e. to construct an electrochemical biosensor for detecting IgG in blood.

2. The method of constructing an electrochemical biosensor for detecting IgG in blood according to claim 1, wherein, In S1, the concentration of aniline in the electrodeposition solution is 0.1 M, the concentration of perchloric acid is 0.03 M; the current density of the chronopotentiometry is 10 μA·cm -2 , and the electrodeposition time is 1 h.

3. The method of constructing an electrochemical biosensor for detecting IgG in blood according to claim 1, wherein, In S2, the concentration of chloroauric acid trihydrate in the mixed aqueous solution is 0.1-5.0 mM, and the concentration of sodium chloride is 0.1-1.0 M; in the cyclic voltammetry, the low potential for electrodeposition is-0.1-0.5 V, the high potential is 0.1-1.0 V, the scanning speed is 10-100 mV / s, and the scanning number is 2-10.

4. The method of constructing an electrochemical biosensor for detecting IgG in blood according to claim 1, wherein, In S3, the concentration of the lariat peptide solution is 1 μM.

5. The method of constructing an electrochemical biosensor for detecting IgG in blood according to claim 1, wherein, In S3, the volume of the lariat peptide solution added dropwise is 10-60 μL, and the incubation time is 6-12 h.

6. The method of constructing an electrochemical biosensor for detecting IgG in blood according to claim 1, wherein, The amino acid sequence of the lariat peptide is C(KEKEKE)EK(AVWGRWH)ECPPPPC, and the following structural features are met: the first amino acid-cysteine to the seventh amino acid-glutamic acid form a ring structure; the cysteine in the ring structure extends a branch chain, and the amino acid sequence of the branch chain is EKECPPPPC, which extends downward from above the ring through the internal space of the ring; the ninth amino acid-lysine is connected with a recognition branch chain with an amino acid sequence of AVWGRWH; the first amino acid-cysteine and the eleventh amino acid-cysteine are connected by a disulfide bond; all the amino acids of the lariat peptide are D-type amino acids, and the overall configuration is lariat-shaped.

7. An electrochemical biosensor for detecting IgG in blood, characterized by, The method is prepared by using the method for constructing an electrochemical biosensor for detecting IgG in blood according to any one of claims 1-6.

8. A detection method of the electrochemical biosensor for detecting IgG in blood according to claim 7, characterized by, The method comprises the following steps: a three-electrode system is formed by taking the electrochemical biosensor as a working electrode, a platinum wire as a counter electrode, and a silver / silver chloride electrode as a reference electrode; a standard immunoglobulin G solution is added dropwise to the surface of the working electrode, then the three-electrode system is immersed in a buffer solution with a pH of 7-8, the change of the current signal is measured by an electrochemical workstation, a standard working curve is drawn according to the corresponding relationship between the concentration of the standard immunoglobulin G and the current signal; the same volume of a blood sample to be detected is added dropwise to the surface of the working electrode, a three-electrode system is formed under the same conditions as above and the change of the current signal is measured, and the concentration of immunoglobulin G in the blood sample to be detected is calculated according to the standard working curve.

9. The detection method of the electrochemical biosensor for detecting IgG in blood according to claim 8, characterized in that, The detection of target immunoglobulin G adopts a ratio-type electrochemical signal mode, and the hydrophilicity and electrical neutrality of the lariat peptide are utilized to realize ultra-low pollution detection in blood.

10. The detection method of the electrochemical biosensor for detecting IgG in blood according to claim 8, characterized in that, The pH 7-8 buffer solution is a 0.2 M phosphate buffer solution.

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