CN102735728B - Electrochemical immunosensor, preparation method and use of electrochemical immunosensor - Google Patents
Electrochemical immunosensor, preparation method and use of electrochemical immunosensor Download PDFInfo
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- CN102735728B CN102735728B CN 201210230340 CN201210230340A CN102735728B CN 102735728 B CN102735728 B CN 102735728B CN 201210230340 CN201210230340 CN 201210230340 CN 201210230340 A CN201210230340 A CN 201210230340A CN 102735728 B CN102735728 B CN 102735728B
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Abstract
The invention relates to an electrochemical immunosensor, and a preparation method and use of the electrochemical immunosensor. The preparation method comprises the following steps: firstly, introducing a metal nanoparticles-mesoporous material compound (Pd hybrid SBA-15) into preparation of the electrochemical immunosensor of human chorionic gonadotropin (hCG); dispensing the compound of sulfated grapheme and thionine on the surface of a glassy carbon electrode to form a composite membrane with high surface energy and good redox-activity; taking thionine as an electronic mediator, taking glutaraldehyde as a cross-linking agent, covalently bonding Pd hybrid SBA-15 mesoporous material with amino, and preparing the hCG electrochemical immunosensor based on Pd hybrid SBA-15, drawing a work curve under the optimum condition and achieving measurement of hCG in a serum sample. The electrochemical immunosensor disclosed by the invention has the advantages of high sensitivity, good reproducibility and simple and fast operation in detection of hCG, and provides a new approach for detection of other tumor markers.
Description
Technical field
The present invention relates to a kind of electrochemical immunosensor, Its Preparation Method And Use, more specifically, the present invention relates to a kind of electrochemical immunosensor, its preparation method based on Pd hydridization SBA-15 (mesoporous silica molecular sieve), and the purposes of the electrochemical immunosensor prepared by the method in measuring human chorionic gonadotrophin.
Background technology
Human chorionic gonadotrophin (hCG) is a kind of ptyalose albumen of the main secretion of the trophocyte by placenta tissue.Under normal circumstances, the concentration in healthy unpregnancy human body is very low, may can't detect.And ectopic pregnancy, many pernicious non-trophoblastic tumors and non-gonadal tumor can produce dystopy hCG, so the detection of hCG becomes forecast early pregnancy and miscarriage, the conventional means that helps ectopic pregnancy and relevant disease thereof to diagnose.The detection method of hCG mainly contains enzyme-linked immunosorbent assay, radioimmunoassay, time resolution immunofluorescence technique etc. at present.But these methods having some limitations property all aspect sensitivity, selectivity, stability and operability.In recent years, electrochemical immunosensor has been subject to paying close attention to widely, and it is divided into sandwich type electrochemical immunosensor and unmarked type electrochemical immunosensor.With the electrochemical immunosensor of sandwich type, compare, unmarked type electrochemical immunosensor operation is more easy, and it does not need to carry out two anti-fixing.
SBA-15, a kind of mesoporous silica molecular sieve, have biocompatibility and the adjustable advantages such as aperture preferably, therefore is widely used in the structure of biology sensor.Up to the present, SBA-15 has been used to the fixing etc. of glucose oxidase, cromoci, antibody.But compare with SBA-15, the Pd nano particle is incorporated into the advantage that the Pd hydridization SBA-15 obtained in the duct of SBA-15 has not only kept the biocompatibility that SBA-15 is good, and show again the good catalytic activity of Pd, further improve the transmission efficiency of electronics, thus can be better fixing for hCG antibody.
Graphene has fabulous crystallinity and outstanding electronics, thermodynamics and mechanical property, is therefore the focus of people's extensive concern.In the research and application of Graphene, in order to give full play to its advantageous property, must carry out functionalization to Graphene.Namely, on grapheme material advantage basis, further material is carried out to property modification and optimization, the sulfonated Graphene (HSO3-GS) that synthesizing water-solubility is good, thus reach the better fixedly purpose of decorative material.
The present invention is dripped the compound of sulfonated Graphene and thionine to be coated onto the glass-carbon electrode surface, forms the composite membrane with high surface energy and good oxidation reducing activity.Thionine is as electron mediator, do the Pd hydridization SBA-15 mesoporous material of crosslinking chemical covalent bond crossed belt amino with glutaraldehyde, prepared the unmarked type electrochemical immunosensor based on Pd hydridization SBA-15, test result shows, the sensitivity of electrochemical immunosensor prepared by said method, selectivity and stability are high, based on above-mentioned discovery, the inventor has completed the present invention.
Summary of the invention
An object of the present invention is to provide a kind of electrochemical immunosensor, described electrochemical immunosensor comprises: working electrode, contrast electrode and to electrode, the basal electrode of described working electrode is glass-carbon electrode, and its surface is modified compound, the glutaraldehyde of sulfonated Graphene and thionine, hCG antibody-solutions, bovine serum albumin and the hCG antigen of Pd hydridization SBA-15 hatching successively.
A further object of the present invention is to provide a kind of preparation method of electrochemical immunosensor, transducer sensitivity prepared by described method is high, favorable reproducibility, easy and simple to handle, the shortcoming such as avoided the instrument and equipment of traditional detection method complicated, operating process is loaded down with trivial details.
Another object of the present invention is to provide the purposes of described electrochemical immunosensor in measuring HCG.
In order to solve the problems of the technologies described above, the present invention realizes by following measures.
Electrochemical immunosensor of the present invention comprises: working electrode, contrast electrode and to electrode, the basal electrode of described working electrode is glass-carbon electrode, and its surface is modified compound, the glutaraldehyde of sulfonated Graphene and thionine, hCG antibody-solutions, bovine serum albumin and the hCG antigen of Pd hydridization SBA-15 hatching successively.Described working electrode and contrast electrode, electrode is formed to electrochemical immunosensor detect hCG antigen.Described contrast electrode is saturated calomel electrode, to electrode, is platinum electrode.
The preparation method of electrochemical immunosensor of the present invention comprises the following steps:
A, synthetic sulfonated Graphene;
The hCG antibody-solutions of b, preparation Pd hydridization SBA-15 hatching;
C, prepare the electrochemical immunosensor working electrode;
D, making electrochemical immunosensor working curve.
Wherein, the synthetic sulfonated Graphene of step a specifically comprises the following steps:
1. prereduction: use sodium borohydride at 80 ℃ of prereduction graphene oxide 1h, to remove most oxygen-containing functional groups;
2. sulfonated: with the aryl diazonium salts ice bath 2h of sulfanilic acid;
3. thoroughly the reduction: with hydrazine go again the reduction (100 ℃, 24h), in order to remove remaining oxygen-containing functional group.
The hCG antibody-solutions of the preparation Pd hydridization SBA-15 hatching of step b specifically comprises the following steps:
1. SBA-15 is joined to back flow reaction 20h in the dry toluene that contains the 3-aminopropyl triethoxysilane, obtain the SBA-15 of amino functional;
2. take PdCl
2, add wherein the HCl of 0.2mol/L, dry and obtain H with infrared lamp
2pdCl
4;
3. by the SBA-15 of amino functional and H
2pdCl
4mix at 1: 1 and be diluted with water to solution with mass ratio, potpourri is at room temperature stirred to 24h;
4. centrifugal rear water dilutes again, with the pH of rare HCl regulator solution, is less than 2 and stir at least 30min.After having stirred, by the NaBH of 50mmol/L
4solution joining in mixed liquor dropwise, until all H
2pdCl
4all be reduced into metal Pd;
5. above-mentioned mixed liquor is centrifugal and vacuum drying, obtain required Pd hydridization SBA-15;
6. the hCG antibody-solutions is joined in the Pd hydridization SBA-15 solution of 6～8mg/mL ultrasonicly, obtain the hCG antibody-solutions of finely dispersed Pd hydridization SBA-15 hatching.
The method for preparing the electrochemical immunosensor working electrode of step c specifically comprises the following steps:
1. polishing is carried out in the glass-carbon electrode surface and make its any surface finish;
2. the compound (volume ratio is 2: 1～1: 1) of getting certain density sulfonated Graphene and thionine evenly drips and is coated in above-mentioned glass-carbon electrode surface, under room temperature, dries;
3. drip the hCG antibody that is coated with Pd hydridization SBA-15 hatching, cleaning is dried;
4. the electrode of above-mentioned processing is placed on to incubation 1h in 1% BSA solution, for the non-specific avtive spot on enclosed-electrode, finally modifies upper hCG antigen, electrode dries and cleans.The immunoelectrode prepared is placed in to refrigerator, saves backup under 4.0 ℃, obtain the working electrode of electrochemical immunosensor.
The making electrochemical immunosensor working curve of steps d, step is as follows:
1. by contrast electrode-saturated calomel electrode, to the working electrode exact connect ion of electrode-platinum electrode and above-mentioned preparation on electrochemical workstation;
2. the PBS buffer solution of pH 7.4 (containing 0.1mol/L KCl as supporting electrolyte) adopts cyclic voltammetry to measure respectively current value under different hCG antigen concentrations as end liquid; Draw human chorionic gonadotrophin working sensor curve.
The purposes of the electrochemical immunosensor that another aspect of the present invention provides said method to prepare in measuring HCG.
Useful achievement of the present invention:
(1) the present inventor is incorporated into metal nanoparticle-mesoporous material compound (Pd hydridization SBA-15) in the middle of the preparation of electrochemical immunosensor of HCG first, utilize the synergistic effect of nano material and the immobilized effect of mesoporous material excellence, make the electrode of made that higher sensitivity and wider sensing range be arranged.
(2) in preparation method of the present invention, hCG antibody and Pd hydridization SBA-15 are directly hatched, reduced the modification step of electrode, improved reappearance and the stability of sensor.
(3) electrochemical immunosensor prepared by method of the present invention, simple to operate, detection speed is fast, can realize at short notice the mensuration of a large amount of samples.
(4) electrochemical immunosensor that uses method of the present invention to prepare, because in detecting liquid, having added electron mediator, make electrochemical immunosensor have higher sensitivity.
Electrochemical immunosensor of the present invention has shown good accuracy, stability, reappearance and high sensitivity, and immunoassay detects rapidly, convenient, can be used for clinical analysis.
The accompanying drawing explanation
Below in conjunction with accompanying drawing explanation and specific embodiment, the present invention is described in further detail.
The working curve that Fig. 1 is electrochemical immunosensor of the present invention.
Embodiment
The human chorionic gonadotrophin (hCG) that the present invention uses and hCG antibodies are purchased from Zhengzhou City rich match biotechnology research institute, PdCl
2, sodium carbonate and sodium nitrite be purchased from Chemical Reagent Co., Ltd., Sinopharm Group, sodium borohydride and sulfanilic acid are purchased from Chinese Shanghai reagent one factory.Cleaning with ultrapure water except glass-carbon electrode, what whole experimentation was used is all intermediate waters.
The CHI760D electrochemical workstation is purchased from Shanghai Chen Hua Instrument Ltd..
Embodiment 1
(1) sulfonated Graphene is synthetic
The graphite oxide of 37.5mg is distributed in the water of 37.5g.Then obtain brown graphene oxide suspending liquid after ultrasonic 1h.
Prereduction: the pH that first with the sodium carbonate liquor of 5wt%, regulates above-mentioned graphene oxide suspending liquid is 9, by 300mg NaBH
4be dissolved in 7.5g water, and then by NaBH
4solution is added in graphene oxide suspending liquid.Potpourri is constantly stirred under 80 ℃ of conditions to 1h.This reduction process is carried out under nitrogen atmosphere.Suspending liquid becomes black by coffee color.See gathering after reduction reaction finishes, water is centrifugal and clean after 2～3 times, through ultrasonic, the graphene oxide of partial reduction is dispersed in the water of 37.5g, obtains suspending liquid;
Sulfonated: 23mg sulfanilic acid and 9mg sodium nitrite react in 5g water and 0.25g 1mol/LHCl, form diazo salt in ice bath.Under the condition of ice bath and constantly stirring, add above-mentioned diazonium salt solution in the graphene oxide suspending liquid of partial reduction, and keep ice bath 2h.Reaction mixture passes into nitrogen and drives bubble away.After after reaction finishes, centrifugal and water cleans 2～3 times, Sulfonated graphene oxide is distributed in 37.5g water, obtains suspending liquid;
Thoroughly reduction: after reduction step in, drip to the hydrazine of 1g (50%) in 2.5g water and join in the suspending liquid that sulfonated step obtains, and reaction mixture 100 ℃ of maintenances under the condition constantly stirred, 24h.In potpourri, add the sodium carbonate liquor of 5wt% in order to precipitate Sulfonated Graphene afterwards.After the thorough cleaning of water and vacuum drying, obtain Sulfonated Graphene, by ultrasonic, it is dispersed in water to be mixed with the sulfonated graphene solution of 2mg/mL.
(2) prepare the hCG antibody-solutions of Pd hydridization SBA-15 hatching
1g SBA-15 is joined to back flow reaction 20h in the 80mL dry toluene that contains 1mL 3-aminopropyl triethoxysilane, obtain the SBA-15 of amino functional;
Take the PdCl of 0.0887g
2, add wherein the HCl of 6mL 0.2mol/L, dry and obtain H with infrared lamp
2pdCl
4;
By the SBA-15 of amino functional and H
2pdCl
4mix at 1: 1 and be diluted with water to solution with mass ratio, potpourri is at room temperature stirred to 24h;
Centrifugal rear water dilutes again, with the pH of rare HCl regulator solution, is less than 2 and stir at least 30min.After having stirred, by the NaBH of 50mmol/L
4solution joining in mixed liquor dropwise, until all H
2pdCl
4all be reduced into metal Pd;
Centrifugal and the vacuum drying by above-mentioned mixed liquor, obtain required Pd hydridization SBA-15;
The Pd hydridization SBA-15 solid that takes above-mentioned preparation is dissolved in it in water and is mixed with the Pd hydridization SBA-15 solution that concentration is 6mg/mL;
By 5mL concentration be 12ng/mL hCG antibody-solutions to join 5mL concentration be in 6mg/mL Pd hydridization SBA-15 solution, shake 24h in 4 ℃ of constant temperature oscillation casees, obtain the hCG antibody-solutions of finely dispersed Pd hydridization SBA-15 hatching;
(3) prepare the electrochemical immunosensor working electrode
Alundum (Al2O3) burnishing powder polishing by the glass-carbon electrode of 4mm diameter with 1.0,0.3,0.05 μ m, the ethanol ultrasonic cleaning, with ultrapure water, rinse well again, then electrode is placed in to the 0.05mol/L potassium ferricyanide solution, at-0.2～0.6V, scan, make the poor 110mV of being less than of spike potential, with ultrapure water cleaning electrode surface, dry up;
The sulfonated Graphene that fixedly prepared by above-mentioned steps (1) and the concentration of thionine solution are 2mg/mL, and the sulfonated Graphene that is 2: 1 by volume ratio and thionine mixed solution evenly drip the glass-carbon electrode surface that is coated in above-mentioned processing, under room temperature, dry;
Then drip the hCG antibody-solutions of the Pd hydridization SBA-15 hatching that is coated with preparation in above-mentioned step (2), cleaning is dried;
The electrode of above-mentioned processing is placed on to incubation 1h in 1%BSA solution, for the non-specific avtive spot on enclosed-electrode, finally modifies upper hCG, electrode dries and cleans.The immunoelectrode prepared is placed in to refrigerator, saves backup under 4.0 ℃, obtain the working electrode of electrochemical immunosensor.
(4) make the electrochemical immunosensor working curve
Open electrochemical workstation, by contrast electrode-saturated calomel electrode, to the working electrode exact connect ion of electrode-platinum electrode and above-mentioned preparation on electrochemical workstation;
The PBS buffer solution of 10mL pH 7.4, as end liquid, is measured respectively the current value of oxidation peak under different hCG antigen concentrations;
Linear according to gained oxidation peak current value and hCG antigen concentration, the drawing curve, as shown in Figure 1.Measurement result shows, hCG concentration is linear in 0.01～16.00ng/mL scope, and linearly dependent coefficient is 0.9991.Detect and be limited to 8.60pg/mL.
Embodiment 2
(1) sulfonated Graphene is synthetic
The graphite oxide of 37.5mg is distributed in the water of 37.5g.Then obtain brown graphene oxide suspending liquid after ultrasonic 1h.
Prereduction: the pH that first with the sodium carbonate of 5wt%, regulates above-mentioned graphene oxide suspending liquid is 10, by 300mg NaBH
4be dissolved in 7.5g water, and then by NaBH
4solution is added in above-mentioned graphene oxide suspending liquid.Potpourri is constantly stirred under 80 ℃ of conditions to 1h.This reduction process is carried out under nitrogen atmosphere.Suspending liquid becomes black by coffee color.See gathering after reduction reaction finishes, water is centrifugal and clean after 2～3 times, through ultrasonic, the graphene oxide of partial reduction is dispersed in the water of 37.5g, obtains suspending liquid;
Sulfonated: 23mg sulfanilic acid and 9mg sodium nitrite react in 5g water and 0.25g 1mol/LHCl, under ice bath, form diazo salt.Under the condition of ice bath and constantly stirring, add above-mentioned diazonium salt solution in the graphene oxide suspending liquid of partial reduction, and keep ice bath 2h.Reaction mixture passes into nitrogen and drives bubble away.After after reaction finishes, centrifugal and water cleans 2～3 times, Sulfonated graphene oxide is distributed in 37.5g water, obtains suspending liquid;
Thoroughly reduction: after reduction step in, drip to the hydrazine of 1g (50%) in 2.5g water and join in the suspending liquid that sulfonated step obtains, and reaction mixture 100 ℃ of maintenances under the condition constantly stirred, 24h.In potpourri, add the sodium carbonate liquor of 5wt% in order to precipitate Sulfonated Graphene afterwards.After the thorough cleaning of water and vacuum drying, obtain Sulfonated Graphene, by ultrasonic, it is dispersed in water to be mixed with the sulfonated graphene solution of 2mg/mL.
(2) prepare the hCG antibody-solutions of Pd hydridization SBA-15 hatching
1g SBA-15 is joined to back flow reaction 20h in the 80mL dry toluene that contains 1mL 3-aminopropyl triethoxysilane, obtain the SBA-15 of amino functional;
Take the PdCl of 0.0887g
2, add wherein the HCl of 6mL 0.2mol/L, dry and obtain H with infrared lamp
2pdCl
4;
By the SBA-15 of amino functional and H
2pdCl
4mix at 1: 1 and be diluted with water to solution with mass ratio, potpourri is at room temperature stirred to 24h;
Centrifugal rear water dilutes again, with the pH of rare HCl regulator solution, is less than 2 and stir at least 30min.After having stirred, by the NaBH of 50mmol/L
4solution joining in mixed liquor dropwise, until all H
2pdCl
4all be reduced into metal Pd;
Centrifugal and the vacuum drying by above-mentioned mixed liquor, obtain required Pd hydridization SBA-15;
Taking a certain amount of Pd hydridization SBA-15 solid is dissolved in it in water and is mixed with the Pd hydridization SBA-15 solution that concentration is 8mg/mL;
By 5mL concentration be 12ng/mL hCG antibody-solutions to join 5mL concentration be in 8mg/mL Pd hydridization SBA-15 solution, shake 24h in 4 ℃ of constant temperature oscillation casees, obtain the hCG antibody-solutions of finely dispersed Pd hydridization SBA-15 hatching;
(3) prepare the electrochemical immunosensor working electrode
Alundum (Al2O3) burnishing powder polishing by the glass-carbon electrode of 4mm diameter with 1.0,0.3,0.05 μ m, the ethanol ultrasonic cleaning, with ultrapure water, rinse well again, then electrode is placed in to the 0.05mol/L potassium ferricyanide solution, at-0.2～0.6V, scan, make the poor 110mV of being less than of spike potential, with ultrapure water cleaning electrode surface, dry up;
The sulfonated Graphene of fixing above-mentioned preparation and the concentration of thionine solution are 2mg/mL, and the sulfonated Graphene that is 1: 1 by volume ratio and thionine mixed solution evenly drip and are coated in above-mentioned glass-carbon electrode surface, under room temperature, dry;
Then drip the hCG antibody-solutions that is coated with above-mentioned Pd hydridization SBA-15 hatching, cleaning is dried;
The glass-carbon electrode of above-mentioned processing is placed on to incubation 1h in 1wt% BSA solution, for the non-specific avtive spot on enclosed-electrode, finally modifies upper hCG, electrode dries and cleans.The immunoelectrode prepared is placed in to refrigerator, saves backup under 4.0 ℃, obtain the working electrode of electrochemical immunosensor.
(4) make the electrochemical immunosensor working curve
Open electrochemical workstation, by contrast electrode-saturated calomel electrode, to the working electrode exact connect ion of electrode-platinum electrode and above-mentioned preparation on electrochemical workstation;
HCG in human serum is measured, according to embodiment 1 gained working curve, calculate the content of hCG in serum.Five times measurement result shows, five measurement results of Human Blood are respectively 1.54,1.56,1.52,1.55,1.51ng/mL, and average hCG content is 1.54ng/mL, and its result is basically identical with the 1.57ng/mL as a result that adopts the standard enzyme linked immunosorbent assay to obtain.Adopt standard addition method, the addition of hCG is 5.00ng/mL simultaneously, and the average recovery rate obtained is 101%, and relative standard deviation is 1.4%.Illustrate that the electrochemical immunosensor that preparation method of the present invention obtains has higher accuracy and precision.
Claims (1)
1. a preparation method who measures the electrochemical immunosensor of HCG (hCG), described electrochemical immunosensor comprises: working electrode, contrast electrode and to electrode, the basal electrode of described working electrode is glass-carbon electrode, and its surface is modified compound, the glutaraldehyde of sulfonated Graphene and thionine, hCG antibody-solutions, bovine serum albumin and the hCG antigen of Pd hydridization SBA-15 hatching successively;
It is characterized in that, described preparation method comprises the following steps:
A, synthetic sulfonated Graphene;
The hCG antibody-solutions of b, preparation Pd hydridization SBA-15 hatching;
C, prepare the working electrode of electrochemical immunosensor;
The working curve of d, making electrochemical immunosensor;
Wherein, the concrete steps of the described synthetic sulfonated Graphene of step a are as follows:
(1) prereduction: use sodium borohydride 80
oc prereduction graphene oxide 1 h, to remove most oxygen-containing functional groups;
(2) sulfonated: with aryl diazonium salts ice bath 2 h of sulfanilic acid;
(3) thoroughly reduction: 100
oc removes reductase 12 4 h again with hydrazine, in order to remove remaining oxygen-containing functional group;
The concrete preparation process of the hCG antibody-solutions of the described preparation of step b Pd hydridization SBA-15 hatching is as follows:
(1) SBA-15 is joined to back flow reaction 20 h in the dry toluene that contains the 3-aminopropyl triethoxysilane, obtain the SBA-15 of amino functional;
(2) take PdCl
2, add wherein the HCl of 0.2 mol/L, dry and obtain H with infrared lamp
2pdCl
4;
(3) by the SBA-15 of amino functional and H
2pdCl
4mix at 1: 1 and be diluted with water to solution with mass ratio, potpourri is at room temperature stirred to 24 h;
(4) centrifugal rear water dilutes again, with the pH of rare HCl regulator solution, is less than 2 and stir at least 30 min; After having stirred, by the NaBH of 50 mmol/L
4solution joining in mixed liquor dropwise, until all H
2pdCl
4all be reduced into metal Pd;
(5) above-mentioned mixed liquor is centrifugal and vacuum drying, obtain required Pd hydridization SBA-15;
(6) solution of hCG antibody is joined in the Pd hydridization SBA-15 solution of 6 ~ 8 mg/mL ultrasonicly, obtain the hCG antibody-solutions of finely dispersed Pd hydridization SBA-15 hatching;
Step c is described, and to prepare the concrete steps of electrochemical immunosensor working electrode as follows:
(1) polishing is carried out in the glass-carbon electrode surface and make its any surface finish;
(2) take volume ratio gets the synthetic sulfonated Graphene of step a and the compound of thionine as 2:1 ~ 1:1, evenly drips and is coated in described glass-carbon electrode surface, under room temperature, dries;
(3) drip the hCG antibody that is coated with Pd hydridization SBA-15 hatching prepared by step b, cleaning is dried;
(4) glass-carbon electrode of above-mentioned processing is placed on to incubation 1 h in 1 wt% BSA solution, for the non-specific avtive spot on enclosed-electrode, finally modifies upper hCG antigen, electrode dries and cleans, and the immunoelectrode prepared is placed in to refrigerator, 4.0
?under C, save backup;
The concrete steps of the described making electrochemical immunosensor of steps d working curve are as follows:
(1) by contrast electrode, to the working electrode exact connect ion of electrode and above-mentioned preparation on electrochemical workstation, described contrast electrode is saturated calomel electrode, described is platinum electrode to electrode;
(2) pH 7.4 containing 0.1 mol/L KCl as the PBS buffer solution of supporting electrolyte as end liquid, adopt cyclic voltammetry to measure respectively current value under different hCG antigen concentrations; Draw human chorionic gonadotrophin electrochemical immunosensor working curve.
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