CN109521006B - Preparation method and application of double-quenching competitive electrochemiluminescence sensor based on Au @ NiFe MOFs - Google Patents
Preparation method and application of double-quenching competitive electrochemiluminescence sensor based on Au @ NiFe MOFs Download PDFInfo
- Publication number
- CN109521006B CN109521006B CN201811578328.6A CN201811578328A CN109521006B CN 109521006 B CN109521006 B CN 109521006B CN 201811578328 A CN201811578328 A CN 201811578328A CN 109521006 B CN109521006 B CN 109521006B
- Authority
- CN
- China
- Prior art keywords
- mofs
- nife
- solution
- electrode
- dispersing
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/75—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
- G01N21/76—Chemiluminescence; Bioluminescence
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N27/00—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means
- G01N27/26—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means by investigating electrochemical variables; by using electrolysis or electrophoresis
- G01N27/28—Electrolytic cell components
- G01N27/30—Electrodes, e.g. test electrodes; Half-cells
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N27/00—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means
- G01N27/26—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means by investigating electrochemical variables; by using electrolysis or electrophoresis
- G01N27/28—Electrolytic cell components
- G01N27/30—Electrodes, e.g. test electrodes; Half-cells
- G01N27/327—Biochemical electrodes, e.g. electrical or mechanical details for in vitro measurements
- G01N27/3275—Sensing specific biomolecules, e.g. nucleic acid strands, based on an electrode surface reaction
- G01N27/3278—Sensing specific biomolecules, e.g. nucleic acid strands, based on an electrode surface reaction involving nanosized elements, e.g. nanogaps or nanoparticles
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/531—Production of immunochemical test materials
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Immunology (AREA)
- Physics & Mathematics (AREA)
- Molecular Biology (AREA)
- Engineering & Computer Science (AREA)
- Pathology (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Hematology (AREA)
- Biomedical Technology (AREA)
- Electrochemistry (AREA)
- Urology & Nephrology (AREA)
- Nanotechnology (AREA)
- Spectroscopy & Molecular Physics (AREA)
- Plasma & Fusion (AREA)
- Biotechnology (AREA)
- Cell Biology (AREA)
- Microbiology (AREA)
- Food Science & Technology (AREA)
- Medicinal Chemistry (AREA)
- Investigating Or Analysing Materials By The Use Of Chemical Reactions (AREA)
Abstract
The invention relates to a preparation method and application of a double-quenching competitive electrochemiluminescence immunosensor based on novel nano-materials Au @ NiFe MOFs, belongs to the field of electrochemiluminescence sensors, and firstly takes nano-porous Au @ NiFe MOFs as a double-quenching marker marking antigen and utilizes Ru (bpy)3 2+the/Zr-MOFs is used as a luminophore to construct a double quenching competitive sensor. Due to the double quenching effect of the nano-porous Au @ NiFe MOFs material, the sensitivity of the sensor is greatly improved. According to different electrochemiluminescence signal intensities caused by standard solutions with different concentrations, the ultrasensitive detection of the estrogen diethylstilbestrol is realized.
Description
Technical Field
The invention relates to a preparation method and application of a double quenching competitive electrochemiluminescence immunosensor based on Au @ NiFe MOFs. In particular to a nano material Ru (bpy)3 2+Preparation of/Zr-MOFs and double quenchers Au @ NiFe MOFs and application of the two quenchers Au @ NiFe MOFs in an electrochemiluminescence sensor. By usingAu @ NiFe MOFs have excellent biocompatibility, large specific surface area and high porosity to increase antigen immobilization capacity, and can double quench Ru (bpy)3 2+The luminescence effect of the/Zr-MOFs further achieves the purpose of enhancing the sensitivity of the sensor. The invention belongs to the technical field of electrochemical luminescence detection.
Background
Diethylstilbestrol (DES) is a synthetic non-steroidal estrogenic substance that produces all the same pharmacological and therapeutic effects as natural estradiol. In 1971, Herbst et al formally proposed a relationship between DES and the rare vaginal adenocarcinoma (CCA) of young women in the New England journal of medicine, and thus DES is extremely harmful to humans. If the DES is taken during the pregnancy of the mother, the abortion is easy when the female fetus develops to the adult pregnancy, mainly because the DES causes the uterus to be deformed; for a male fetus, the symptoms caused by DES mainly include: cryptorchidism, testicular cancer, epididymal cyst, infertility, etc. DES is mainly present in animal food such as pork, mutton and fish as an estrogen capable of promoting the synthesis of animal protein, so that the publication No. 235 of Ministry of agriculture of the people's republic of China in the revised highest residual amount of veterinary drugs in animal food stipulates that the highest content of DES in all food animals and all edible tissues is not detected. Therefore, the method for detecting the DES is simple, convenient, rapid and sensitive and has important significance. The electrochemical luminescence method used in the invention has the advantages of low consumption, easy control, high sensitivity and low detection limit, has the advantages of electrochemical and chemiluminescence methods, and is an environment-friendly method. Therefore, the invention designs a competitive electrochemiluminescence immunoassay method for detecting the diethylstilbestrol.
In the present invention, a novel nanomaterial Ru (bpy) is used3 2+The Zr-MOFs is used as an electrochemical luminescent material. The Zr-MOFs can load more Ru (bpy) due to the larger specific surface area of the porous Zr-MOFs3 2+And further enhance the luminous intensity of the sensor. And a novel nano porous material Au @ NiFe MOFs is used as a quencher for the first time and is introduced into the field of electrochemiluminescence. Wherein the Au nano-particles and the NiFe MOFs can both quench Ru (bpy)3 2+The luminescent effect of the/Zr-MOFs achieves the effect of double quenching. And Au @ NiFe MOFs can load more diethylstilbestrol antigen because of the larger specific surface area of the porosity of the Au @ NiFe MOFs. The sensitivity of the sensor can be greatly enhanced as described above. The principle of the invention is that based on the fact that the binding capacity of the diethylstilbestrol standard solution and the diethylstilbestrol antibody is greatly stronger than that of the Au @ NiFe MOFs labeled diethylstilbestrol-BSA coupled antigen, as the concentration of the diethylstilbestrol standard solution is higher, the Au @ NiFe MOFs labeled diethylstilbestrol-BSA coupled antigen combined with the diethylstilbestrol antibody is less, the double quenching effect is weakened, and the luminous intensity is enhanced. In addition, the competitive immunosensor designed by the invention also provides a new method for detecting other estrogen analytes. At present, a method for detecting diethylstilbestrol by constructing a dual-quenching competitive immunosensor based on Au @ NiFe MOFs is not reported.
Disclosure of Invention
The invention aims to provide a simpler and more reliable preparation method and application of a double-quenching competitive electrochemiluminescence immunosensor based on Au @ NiFe MOFs, and the method can be used for realizing the rapid, sensitive, specific and efficient detection of estrogen.
The technical scheme of the invention is as follows:
in order to achieve the purpose, the technical scheme adopted by the invention is as follows:
1. nano material Ru (bpy)3 2+Preparation of/Zr-MOFs labeled antibody, Au @ NiFe MOFs labeled diethylstilbestrol-BSA coupled antigen hatching solution
（1）Ru(bpy)3 2+Preparation of/Zr-MOFs
Dissolving 30-45 mg of 2, 2-bipyridine-5, 5-dicarboxylic acid (BPDC) in 12 mL of N, N-Dimethylformamide (DMF), adding 120-360 muL of triethylamine, then adding 1-5 mL of acetic acid, stirring for 10-15 min to obtain an organic precursor solution, and adding 30-50 mg of ZrCl4Dispersing in 16 mL of N, N-dimethylformamide, stirring for 10-15 min to obtain a metal solution precursor, mixing the two precursor solutions in a 50 mL polytetrafluoroethylene high-pressure reaction kettle, reacting at 70-90 ℃ for 24 h, centrifuging, and performing precipitation with N, N-dimethylformamide,Washing with methanol and ethanol for several times respectively, and vacuum drying at 35 ℃ for 12h to obtain white solid Zr-MOFs; dispersing 18 mg-24 mg Zr-MOFs in 10 mL ethanol solution, stirring for 5 min, and adding 10 mL Ru (bpy)3Cl2·6H2Mixing O (1 mg-3 mg) in N, N-dimethylformamide solution, stirring at 80-100 ℃ for 12h, respectively washing N, N-dimethylformamide and ethanol for several times, and vacuum drying at 35 ℃ for 12h to obtain orange yellow solid Ru (bpy)3 2+/Zr-MOFs；
（2）Ru(bpy)3 2+Preparation of solution of incubation of/Zr-MOFs labeled antibody
10 mg of Ru (bpy)3 2+dispersing/Zr-MOFs in 1 mL of EDC/NHS solution, carrying out shaking reaction at room temperature for 4-8 h, washing off excessive EDC/NHS by using ultrapure water, dispersing the solution in 1 mL of PBS with pH = 7.5, adding 4-6 μ L of 6 mg/mL antibody, carrying out shaking incubation in a 4 ℃ constant temperature shaking incubator for 12-24 h, carrying out centrifugal separation at 4 ℃, and finally dispersing the solution in a PBS buffer solution with pH = 7.5 to prepare 0.5-5 mg/mL Ru (bpy)3 2+The Zr-MOFs labeled conjugate antibody is stored at 4 ℃ for standby;
(3) preparation of nano porous material Au @ NiFe MOFs
Dispersing 140-160 mg of nickel chloride hexahydrate and 260-280 mg of trisodium citrate dihydrate in 20 mL of ultrapure water, magnetically stirring for 5 min, similarly dispersing 130-150 mg of potassium ferricyanide in 20 mL of ultrapure water, magnetically stirring for 5 min, mixing the two solutions, continuously stirring for 8-12 h at room temperature, centrifugally separating suspended matters, respectively washing for 2 times by using ultrapure water and ethanol, and performing vacuum drying at 35 ℃ to obtain yellowish-brown nanoporous NiFe MOFs;
dispersing 30-50 mg of nanoporous NiFe MOFs in 50 mL of ultrapure water, adding 1-3 mL of 1-3% chloroauric acid solution, stirring for 5 min at room temperature, adding 4-6 mg of polyvinylpyrrolidone (PVP) to inhibit agglomeration of gold nanoparticles, adding a reducing agent, namely trisodium citrate (70-90 mg), stirring for 8-12 h at room temperature with sodium borohydride (0.5-1 mg), slowly turning the solution into purple black, centrifugally separating suspended matters, washing with ultrapure water until the solution is colorless, and performing vacuum drying for 12h at 35 ℃ to obtain purple black solid Au @ NiFe MOFs;
(4) preparation of Au @ NiFe MOFs labeled antigen hatching solution
Dispersing Au @ NiFe MOFs of 8 mg-12 mg into 1 mL PBS with pH = 7.5, adding 4 muL-6 muL and 6 mg/mL antigens, carrying out oscillation incubation in a constant-temperature oscillation incubator at 4 ℃ for 12 h-24 h, carrying out centrifugal separation at 4 ℃, adding 1 mL of 0.5% -0.9% bovine serum albumin solution, carrying out oscillation incubation in a constant-temperature oscillation incubator at 4 ℃ for 2h to seal the nonspecific active site on the Au @ NiFe MOFs, carrying out centrifugal separation at 4 ℃, finally dispersing in PBS buffer solution with pH = 7.5, preparing Au @ NiFe MOFs labeled antigens of 0.5 mg/mL-5 mg/mL, and storing the Au @ NiFe MOFs labeled antigens at 4 ℃ for later use.
2. A preparation method of a double quenching competitive electrochemiluminescence sensor of Au @ NiFe MOFs comprises the following steps:
(1) polishing glassy carbon electrodes with the diameter of 4 mm by using alumina polishing powder with the diameter of 1.0 mm, 0.3 mm and 0.05 mm respectively, and washing the glassy carbon electrodes with ultrapure water;
(2) 6 mu L of 0.5 mg/mL to 5 mg/mL Ru (bpy)3 2+Dripping the Zr-MOFs labeled coupled antibody on the surface of the electrode, and airing at 4 ℃;
(3) putting 3 μ L of bovine serum albumin BSA solution with a volume fraction of 1% on the electrode surface to block non-specific active sites on the electrode surface, washing with PBS buffer solution with pH = 7.5, and air-drying at 4 ℃;
(4) dripping 6 μ L of standard solution of the substance to be detected with a certain concentration on the surface of the electrode, washing with PBS buffer solution with pH = 7.5, and air drying at 4 deg.C
(5) Dripping 6 mu L of Au @ NiFe MOFs marked antigen incubation solution of the object to be detected, which is 0.5 mg/mL-5 mg/mL, on the surface of an electrode, airing at 4 ℃, and washing with PBS (phosphate buffer solution) with pH = 7.5 to prepare the double-quenching competitive electrochemiluminescence immunosensor based on Au @ NiFe MOFs.
3. The electrochemiluminescence sensor is used for electrochemiluminescence detection of a sample to be detected:
(1) the method comprises the following steps of (1) testing by using a three-electrode system of an electrochemical workstation, taking an Ag/AgCl electrode as a reference electrode, taking a platinum wire electrode as a counter electrode, taking a prepared glassy carbon electrode modified by an electrochemiluminescence sensor as a working electrode, connecting the electrochemical workstation and a chemiluminescence detector together, setting the high voltage of a photomultiplier to be 500V, setting the cyclic voltammetry scanning potential range to be 0-1.4V, and setting the scanning rate to be 0.1V/s;
(2) in 10 mL of PBS (phosphate buffer solution) with pH of 6.0-8.5 and tripropylamine concentration of 1-12 mmol/L, detecting the intensity of an electrochemiluminescence signal generated by diethylstilbestrol standard solutions with different concentrations through an electrochemiluminescence system, and drawing a working curve;
(3) and (4) replacing the standard solution with the sample solution to be tested for determination.
Advantageous results of the invention
(1) The nano-porous material Au @ NiFe MOFs is used as a quencher for labeling the antigen to carry out double quenching on the luminous effect of the luminophor for the first time, and the nano-porous material Au @ NiFe MOFs has the advantages of excellent biocompatibility, larger specific surface area and the like, and effectively increases the immobilized amount of the antigen, thereby greatly improving the sensitivity of the sensor;
(2) with nano-material Ru (bpy)3 2+the/Zr-MOFs is a luminescent material used in the construction of an electrochemiluminescence sensor, and Ru (bpy) is utilized3 2+The output of the optical signal of the sensor is improved by the high and stable luminous efficiency of the/Zr-MOFs, so that higher sensitivity is obtained;
(3) the invention firstly uses nano-porous Au @ NiFe MOFs and Ru (bpy)3 2+The construction method of the electrochemiluminescence sensor based on the Zr-MOFs is used for constructing the electrochemiluminescence sensor, the sensor based on the construction method can be applied to clinical detection of estrogen, and has the advantages of being simple to operate, rapid in detection, wide in signal linear range (0.1 pg/mL-50 ng/mL) and low in detection limit (0.039 pg/mL).
Example 1
Nano material Ru (bpy)3 2+Preparation of/Zr-MOFs labeled antibody, Au @ NiFe MOFs labeled diethylstilbestrol-BSA coupled antigen hatching solution
（1）Ru(bpy)3 2+Preparation of/Zr-MOFs
Dissolving 30 mg of 2, 2-bipyridine-5, 5-dicarboxylic acid (BPDC) in 12 mL of N, N-Dimethylformamide (DMF), adding 120 mu L of triethylamine, then adding 1 mL-5 mL of acetic acid, stirring for 10 min to obtain an organic precursor solution, and adding 30 mg of ZrCl4Dispersing in 16 mL of N, N-dimethylformamide, stirring for 10 min to obtain a metal solution precursor, mixing two precursor solutions in a 50 mL polytetrafluoroethylene high-pressure reaction kettle, reacting for 24 h at 70 ℃, performing centrifugal separation, washing for several times with N, N-dimethylformamide, methanol and ethanol respectively, and performing vacuum drying for 12h at 35 ℃ to obtain white solid Zr-MOFs; 18 mg of Zr-MOFs were dispersed in 10 mL of an ethanol solution, stirred for 5 min, and 10 mL of a solution containing Ru (bpy)3Cl2·6H2O (1 mg) in N, N-dimethylformamide, stirring at 80 ℃ for 12 hours, washing N, N-dimethylformamide and ethanol respectively several times, vacuum-drying at 35 ℃ for 12 hours to obtain orange-yellow solid Ru (bpy)3 2+/Zr-MOFs；
（2）Ru(bpy)3 2+Preparation of solution of incubation of/Zr-MOFs labeled antibody
10 mg of Ru (bpy)3 2+dispersing/Zr-MOFs in 1 mL of EDC/NHS solution, shaking for 4 h at room temperature, washing off excessive EDC/NHS with ultrapure water, dispersing in 1 mL of PBS with pH = 7.5, adding 4. mu.L of 6 mg/mL antibody, incubating for 12h at 4 ℃ under constant temperature shaking incubator, centrifuging at 4 ℃, and finally dispersing in PBS buffer solution with pH = 7.5 to obtain 0.5 mg/mL Ru (bpy)3 2+The Zr-MOFs labeled conjugate antibody is stored at 4 ℃ for standby;
(3) preparation of nano porous material Au @ NiFe MOFs
Dispersing 140 mg of nickel chloride hexahydrate and 260 mg of trisodium citrate dihydrate in 20 mL of ultrapure water, magnetically stirring for 5 min, similarly dispersing 130 mg of potassium ferricyanide in 20 mL of ultrapure water, magnetically stirring for 5 min, mixing the two solutions, continuously stirring for 8 h at room temperature, centrifugally separating suspended matters, respectively washing for 2 times by using the ultrapure water and ethanol, and performing vacuum drying at 35 ℃ to obtain khaki nanoporous NiFe MOFs;
dispersing 30 mg of nano-porous NiFe MOFs in 50 mL of ultrapure water, adding 1 mL of 1% chloroauric acid solution, stirring for 5 min at room temperature, adding 4 mg of polyvinylpyrrolidone (PVP) to inhibit the agglomeration of gold nanoparticles, then adding a reducing agent, namely trisodium citrate (70 mg) and sodium borohydride (0.5 mg), stirring for 8 h at room temperature, slowly changing the solution into purple black, centrifugally separating suspended matters, washing ultrapure water until the supernatant is colorless, and performing vacuum drying for 12h at 35 ℃ to obtain purple black solid Au @ NiFe MOFs;
(4) preparation of Au @ NiFe MOFs labeled antigen hatching solution
Dispersing 8 mg of Au @ NiFe MOFs into 1 mL of PBS with pH = 7.5, adding 4 muL and 6 mg/mL of antigen, carrying out oscillation incubation for 12h in a 4 ℃ constant-temperature oscillation incubator, carrying out centrifugal separation at 4 ℃, adding 1 mL of 0.5% bovine serum albumin solution, carrying out oscillation incubation for 2h in the 4 ℃ constant-temperature oscillation incubator to seal the nonspecific active site on the Au @ NiFe MOFs, carrying out centrifugal separation at 4 ℃, finally dispersing into a PBS buffer solution with pH = 7.5 to prepare 0.5 mg/mL of Au @ NiFe MOFs labeled antigen, and storing at 4 ℃ for later use.
Example 2
Nano material Ru (bpy)3 2+Preparation of/Zr-MOFs labeled antibody, Au @ NiFe MOFs labeled diethylstilbestrol-BSA coupled antigen hatching solution
（1）Ru(bpy)3 2+Preparation of/Zr-MOFs
Dissolving 37.5 mg of 2, 2-bipyridine-5, 5-dicarboxylic acid (BPDC) in 12 mL of N, N-Dimethylformamide (DMF), adding 240. mu.L of triethylamine, then adding 3 mL of acetic acid, stirring for 12.5 min to obtain an organic precursor solution, and adding 40 mg of ZrCl4Dispersing in 16 mL of N, N-dimethylformamide, stirring for 12.5 min to obtain a metal solution precursor, mixing two precursor solutions in a 50 mL polytetrafluoroethylene high-pressure reaction kettle, reacting for 24 h at 80 ℃, performing centrifugal separation, washing for several times by using N, N-dimethylformamide, methanol and ethanol respectively, and performing vacuum drying for 12h at 35 ℃ to obtain white solid Zr-MOFs; 21 mg of Zr-MOFs were dispersed in 10 mL of an ethanol solution, stirred for 5 min, and 10 mL of a solution containing Ru (bpy)3Cl2·6H2Mixing O (2 mg) in N, N-dimethylformamide, stirring at 90 deg.C for 12 hr, and washing with N, N-dimethylformamide and ethanol respectivelyVacuum drying at 35 deg.C for 12h to obtain orange solid Ru (bpy)3 2+/Zr-MOFs；
（2）Ru(bpy)3 2+Preparation of solution of incubation of/Zr-MOFs labeled antibody
10 mg of Ru (bpy)3 2+dispersing/Zr-MOFs in 1 mL of EDC/NHS solution, shaking for 6 h at room temperature, washing off excessive EDC/NHS with ultrapure water, dispersing in 1 mL of PBS with pH = 7.5, adding 5. mu.L of 6 mg/mL antibody, shaking and incubating in a 4 ℃ constant temperature shaking incubator for 18 h, centrifuging at 4 ℃, and finally dispersing in PBS buffer solution with pH = 7.5 to obtain 2.5 mg/mL Ru (bpy)3 2+The Zr-MOFs labeled conjugate antibody is stored at 4 ℃ for standby;
(3) preparation of nano porous material Au @ NiFe MOFs
Dispersing 150 mg of nickel chloride hexahydrate and 170 mg of trisodium citrate dihydrate in 20 mL of ultrapure water, magnetically stirring for 5 min, similarly dispersing 140 mg of potassium ferricyanide in 20 mL of ultrapure water, magnetically stirring for 5 min, mixing the two solutions, continuously stirring for 10 h at room temperature, centrifugally separating suspended matters, respectively washing for 2 times by using the ultrapure water and ethanol, and performing vacuum drying at 35 ℃ to obtain khaki nanoporous NiFe MOFs;
dispersing 40 mg of nano-porous NiFe MOFs in 50 mL of ultrapure water, adding 2 mL of 2% chloroauric acid solution, stirring for 5 min at room temperature, adding 5 mg of polyvinylpyrrolidone (PVP) to inhibit the agglomeration of gold nanoparticles, then adding a reducing agent, namely trisodium citrate (80 mg) and sodium borohydride (0.75 mg), stirring for 10 h at room temperature, slowly changing the solution into purple black, centrifugally separating suspended matters, washing ultrapure water until the supernatant is colorless, and performing vacuum drying for 12h at 35 ℃ to obtain purple black solid Au @ NiFe MOFs;
(4) preparation of Au @ NiFe MOFs labeled antigen hatching solution
Dispersing 10 mg of Au @ NiFe MOFs into 1 mL of PBS with pH = 7.5, adding 5 muL and 6 mg/mL of antigen, carrying out oscillation incubation for 18 h in a 4 ℃ constant-temperature oscillation incubator, carrying out centrifugal separation at 4 ℃, adding 1 mL of 0.7% bovine serum albumin solution, carrying out oscillation incubation for 2h in the 4 ℃ constant-temperature oscillation incubator to seal the nonspecific active site on the Au @ NiFe MOFs, carrying out centrifugal separation at 4 ℃, finally dispersing in a PBS buffer solution with pH = 7.5 to prepare 2.5 mg/mL of Au @ NiFe MOFs labeled antigen, and storing the antigen at 4 ℃ for later use.
Example 3
Nano material Ru (bpy)3 2+Preparation of/Zr-MOFs labeled antibody, Au @ NiFe MOFs labeled diethylstilbestrol-BSA coupled antigen hatching solution
（1）Ru(bpy)3 2+Preparation of/Zr-MOFs
Dissolving 45 mg of 2, 2-bipyridine-5, 5-dicarboxylic acid (BPDC) in 12 mL of N, N-Dimethylformamide (DMF), adding 360 μ L of triethylamine, then adding 5 mL of acetic acid, stirring for 15 min to obtain an organic precursor solution, and adding 50 mg of ZrCl4Dispersing in 16 mL of N, N-dimethylformamide, stirring for 15 min to obtain a metal solution precursor, mixing two precursor solutions in a 50 mL polytetrafluoroethylene high-pressure reaction kettle, reacting for 24 h at 90 ℃, performing centrifugal separation, washing for several times with N, N-dimethylformamide, methanol and ethanol respectively, and performing vacuum drying for 12h at 35 ℃ to obtain white solid Zr-MOFs; 24 mg of Zr-MOFs were dispersed in 10 mL of an ethanol solution, stirred for 5 min, and then stirred with 10 mL of a solution containing (Ru (bpy))3 2+）Cl2·6H2O (3 mg) in N, N-dimethylformamide, stirring at 100 ℃ for 12 hours, washing N, N-dimethylformamide and ethanol respectively several times, vacuum-drying at 35 ℃ for 12 hours to obtain orange-yellow solid Ru (bpy)3 2+/Zr-MOFs；
（2）Ru(bpy)3 2+Preparation of solution of incubation of/Zr-MOFs labeled antibody
10 mg of Ru (bpy)3 2+dispersing/Zr-MOFs in 1 mL of EDC/NHS solution, shaking for 8 h at room temperature, washing off excessive EDC/NHS with ultrapure water, dispersing in 1 mL of PBS with pH = 7.5, adding 6 uL of 6 mg/mL antibody, shaking and incubating in a 4 ℃ constant temperature shaking incubator for 24 h, centrifuging at 4 ℃, and finally dispersing in PBS buffer solution with pH = 7.5 to obtain 5 mg/mL Ru (bpy)3 2+The Zr-MOFs labeled conjugate antibody is stored at 4 ℃ for standby;
(3) preparation of nano porous material Au @ NiFe MOFs
Dispersing 160 mg of nickel chloride hexahydrate and 280 mg of trisodium citrate dihydrate in 20 mL of ultrapure water, magnetically stirring for 5 min, similarly dispersing 150 mg of potassium ferricyanide in 20 mL of ultrapure water, magnetically stirring for 5 min, mixing the two solutions, continuously stirring at room temperature for 12h, centrifugally separating suspended matters, washing for 2 times by using the ultrapure water and ethanol respectively, and performing vacuum drying at 35 ℃ to obtain khaki nanoporous NiFe MOFs;
dispersing 50 mg of nano-porous NiFe MOFs in 50 mL of ultrapure water, adding 3 mL of 3% chloroauric acid solution, stirring for 5 min at room temperature, adding 6 mg of polyvinylpyrrolidone (PVP) to inhibit the agglomeration of gold nanoparticles, then adding a reducing agent, namely trisodium citrate (90 mg), stirring for 12h at room temperature by using sodium borohydride (1 mg), slowly changing the solution into purple black, centrifugally separating suspended matters, washing the ultrapure water until the supernatant is colorless, and performing vacuum drying for 12h at 35 ℃ to obtain a purple black solid Au @ NiFe MOFs;
(4) preparation of Au @ NiFe MOFs labeled antigen hatching solution
Dispersing 12 mg of Au @ NiFe MOFs into 1 mL of PBS with pH = 7.5, adding 6 muL and 6 mg/mL of antigen, carrying out oscillation incubation for 24 h in a 4 ℃ constant-temperature oscillation incubator, carrying out centrifugal separation at 4 ℃, adding 1 mL of 0.9% bovine serum albumin solution, carrying out oscillation incubation for 2h in the 4 ℃ constant-temperature oscillation incubator to seal the nonspecific active site on the Au @ NiFe MOFs, carrying out centrifugal separation at 4 ℃, finally dispersing into a PBS buffer solution with pH = 7.5 to prepare 5 mg/mL of labeled antigen, and storing at 4 ℃ for later use.
Example 4
A preparation method of a double-quenching competitive electrochemiluminescence sensor based on Au @ NiFe MOFs comprises the following steps:
(1) polishing glassy carbon electrodes with the diameter of 4 mm by using alumina polishing powder with the diameter of 1.0 mm, 0.3 mm and 0.05 mm respectively, and washing the glassy carbon electrodes with ultrapure water;
(2) 6 μ L0.5 mg/mL Ru (bpy)3 2+Dripping the Zr-MOFs labeled coupled antibody on the surface of the electrode, and airing at 4 ℃;
(3) putting 3 μ L of bovine serum albumin BSA solution with a volume fraction of 1% on the electrode surface to block non-specific active sites on the electrode surface, washing with PBS buffer solution with pH = 7.5, and air-drying at 4 ℃;
(4) dripping 6 μ L of standard solution of the substance to be detected with a certain concentration on the surface of the electrode, washing with PBS buffer solution with pH = 7.5, and air drying at 4 deg.C
(5) And dripping 6 mu L of 0.5 mg/mL of Au @ NiFe MOFs labeled antigen incubation solution of the object to be detected on the surface of the electrode, drying at 4 ℃, and washing with PBS (phosphate buffer solution) with pH = 7.5 to prepare the double-quenching competitive electrochemiluminescence immunosensor based on the Au @ NiFe MOFs.
Example 5
A preparation method of a double-quenching competitive electrochemiluminescence sensor based on Au @ NiFe MOFs comprises the following steps:
(1) polishing glassy carbon electrodes with the diameter of 4 mm by using alumina polishing powder with the diameter of 1.0 mm, 0.3 mm and 0.05 mm respectively, and washing the glassy carbon electrodes with ultrapure water;
(2) 6 μ L2.5 mg/mL Ru (bpy)3 2+Dripping the Zr-MOFs labeled coupled antibody on the surface of the electrode, and airing at 4 ℃;
(3) putting 3 μ L of bovine serum albumin BSA solution with a volume fraction of 1% on the electrode surface to block non-specific active sites on the electrode surface, washing with PBS buffer solution with pH = 7.5, and air-drying at 4 ℃;
(4) dripping 6 μ L of standard solution of the substance to be detected with a certain concentration on the surface of the electrode, washing with PBS buffer solution with pH = 7.5, and air drying at 4 deg.C
(5) And dripping 6 mu L of 2.5 mg/mL of Au @ NiFe MOFs labeled antigen incubation solution of the object to be detected on the surface of the electrode, drying at 4 ℃, and washing with PBS (phosphate buffer solution) with pH = 7.5 to prepare the double-quenching competitive electrochemiluminescence immunosensor based on the Au @ NiFe MOFs.
Example 6
A preparation method of a double-quenching competitive electrochemiluminescence sensor based on Au @ NiFe MOFs comprises the following steps:
(1) polishing glassy carbon electrodes with the diameter of 4 mm by using alumina polishing powder with the diameter of 1.0 mm, 0.3 mm and 0.05 mm respectively, and washing the glassy carbon electrodes with ultrapure water;
(2) 6 μ L5 mg/mL Ru (bpy)3 2+Dripping the Zr-MOFs labeled coupled antibody on the surface of the electrode, and airing at 4 ℃;
(3) putting 3 μ L of bovine serum albumin BSA solution with a volume fraction of 1% on the electrode surface to block non-specific active sites on the electrode surface, washing with PBS buffer solution with pH = 7.5, and air-drying at 4 ℃;
(4) dripping 6 μ L of standard solution of the substance to be detected with a certain concentration on the surface of the electrode, washing with PBS buffer solution with pH = 7.5, and air drying at 4 deg.C
(5) Dripping 6 mu L of 5 mg/mL of Au @ NiFe MOFs labeled antigen incubation solution of the object to be detected on the surface of the electrode, drying at 4 ℃, and washing with PBS (phosphate buffer solution) with pH = 7.5 to prepare the Au @ NiFe MOFs-based double-quenching competitive electrochemiluminescence immunosensor.
Example 7
The electrochemiluminescence sensor is used for electrochemiluminescence detection of diethylstilbestrol:
(1) the method comprises the following steps of (1) testing by using a three-electrode system of an electrochemical workstation, taking an Ag/AgCl electrode as a reference electrode, taking a platinum wire electrode as a counter electrode, taking a prepared glassy carbon electrode modified by an electrochemiluminescence sensor as a working electrode, connecting the electrochemical workstation and a chemiluminescence detector together, setting the high voltage of a photomultiplier to be 500V, setting the cyclic voltammetry scanning potential range to be 0-1.4V, and setting the scanning rate to be 0.1V/s;
(2) detecting the intensity of an electrochemiluminescence signal generated to diethylstilbestrol antigen with different concentrations in 10 mL of PBS (phosphate buffer solution) with pH 7.5 and tripropylamine concentration of 10 mmol/L by an electrochemiluminescence system, and drawing a working curve;
(3) and (4) replacing the standard solution with the sample solution to be tested for determination.
Example 8
Detection of estrogen diethylstilbestrol in fish extract
1 mL of fish extract, diethylstilbestrol standard solutions with different concentrations are added into the fish extract, and the average recovery rate of diethylstilbestrol in the samples is determined by adopting a standard addition method, and the results are shown in Table 1.
TABLE 1 results of detection of diethylstilbestrol in samples
The detection results in the table 1 show that the recovery rate of the detection result of the diethylstilbestrol in the fish meat extract is 95.0-105%, which indicates that the method can be used for detecting actual biological samples, the precision of the method is high, and the result is accurate and reliable.
Claims (2)
1. A preparation method of a double-quenching electrochemiluminescence immunosensor based on Au @ NiFe MOFs is characterized by comprising the following steps:
（1）Ru(bpy)3 2+preparation of/Zr-MOFs
Dissolving 30-45 mg of 2, 2-bipyridine-5, 5-dicarboxylic acid in 12 mL of N, N-dimethylformamide, adding 120-360 muL of triethylamine, then adding 1-5 mL of acetic acid, stirring for 10-15 min to obtain an organic precursor solution, and adding 30-50 mg of ZrCl4Dispersing in 16 mL of N, N-dimethylformamide, stirring for 10-15 min to obtain a metal solution precursor, mixing two precursor solutions in a 50 mL polytetrafluoroethylene high-pressure reaction kettle, reacting for 24 h at 70-90 ℃, performing centrifugal separation, washing for several times by using N, N-dimethylformamide, methanol and ethanol respectively, and performing vacuum drying for 12h at 35 ℃ to obtain white solid Zr-MOFs; dispersing 18 mg-24 mg of Zr-MOFs in 10 mL of ethanol solution, stirring for 5 min, and mixing with 10 mL of 1 mg-3 mg Ru (bpy)3Cl2·6H2Mixing O in N, N-dimethylformamide solution, stirring at 80-100 ℃ for 12h, respectively washing N, N-dimethylformamide and ethanol for several times, vacuum drying at 35 ℃ for 12h to obtain orange yellow solid Ru (bpy)3 2+/Zr-MOFs；
（2）Ru(bpy)3 2+Preparation of solution of incubation of/Zr-MOFs labeled antibody
10 mg of Ru (bpy)3 2+The Zr-MOFs are dispersed inShaking and reacting for 4-8 h at room temperature in 1 mL of EDC/NHS solution, washing off excessive EDC/NHS by ultrapure water, dispersing into 1 mL of PBS with pH = 7.5, adding 4-6 μ L of 6 mg/mL antibody, shaking and incubating for 12-24 h in a 4 ℃ constant temperature shaking incubator, centrifugally separating at 4 ℃, and finally dispersing into PBS buffer solution with pH = 7.5 to prepare 0.5-5 mg/mL Ru (bpy)3 2+The Zr-MOFs labeled conjugate antibody is stored at 4 ℃ for standby;
(3) preparation of nano porous material Au @ NiFe MOFs
Dispersing 140-160 m of nickel chloride hexahydrate and 260-280 mg of trisodium citrate dihydrate into 20 mL of ultrapure water, magnetically stirring for 5 min, similarly dispersing 130-150 mg of potassium ferricyanide into 20 mL of ultrapure water, magnetically stirring for 5 min, mixing the two solutions, continuously stirring for 8-12 h at room temperature, centrifugally separating suspended matters, respectively washing for 2 times by using ultrapure water and ethanol, and performing vacuum drying at 35 ℃ to obtain yellowish-brown nanoporous NiFe MOFs;
dispersing 30 mg-50 mg of nanoporous NiFe MOFs in 50 mL of ultrapure water, adding 1 mL-3 mL of 1% -3% chloroauric acid solution, stirring for 5 min at room temperature, adding 4 mg-6 mg of polyvinylpyrrolidone to inhibit agglomeration of gold nanoparticles, adding 70 mg-90 mg of reducing agent trisodium citrate and 0.5 mg-1 mg of sodium borohydride, stirring for 8 h-12 h at room temperature, slowly turning the solution into purple black, centrifugally separating suspended matters, washing ultrapure water to obtain a supernatant, and performing vacuum drying for 12h at 35 ℃ to obtain purple black solid Au @ NiFe MOFs;
(4) preparation of Au @ NiFe MOFs labeled antigen hatching solution
Dispersing Au @ NiFe MOFs of 8 mg-12 mg into 1 mL PBS with pH = 7.5, adding antigens of 4 muL-6 muL and 6 mg/mL, carrying out oscillation incubation in a constant-temperature oscillation incubator at 4 ℃ for 12 h-24 h, carrying out centrifugal separation at 4 ℃, adding a bovine serum albumin solution of 1 mL 0.5% -0.9%, carrying out oscillation incubation in a constant-temperature oscillation incubator at 4 ℃ for 2h to seal the nonspecific active site on the Au @ NiFe MOFs, carrying out centrifugal separation at 4 ℃, finally dispersing in a PBS buffer solution with pH = 7.5, preparing Au @ NiFe MOFs labeled antigens of 0.5 mg/mL-5 mg/mL, and storing the Au @ NiFe MOFs labeled antigens at 4 ℃ for later use;
(5) polishing glassy carbon electrodes with the diameter of 4 mm by using alumina polishing powder with the diameter of 1.0 mm, 0.3 mm and 0.05 mm respectively, and washing the glassy carbon electrodes with ultrapure water;
(6) 6 mu L of 0.5 mg/mL to 5 mg/mL Ru (bpy)3 2+Dripping the Zr-MOFs labeled coupled antibody on the surface of the electrode, and airing at 4 ℃;
(7) putting 3 μ L of bovine serum albumin BSA solution with a volume fraction of 1% on the electrode surface to block non-specific active sites on the electrode surface, washing with PBS buffer solution with pH = 7.5, and air-drying at 4 ℃;
(8) dripping 6 μ L of standard solution of the substance to be detected with a certain concentration on the surface of the electrode, washing with PBS buffer solution with pH = 7.5, and air drying at 4 deg.C
(9) Dripping 6 mu L of Au @ NiFe MOFs marked antigen incubation solution of the object to be detected, which is 0.5 mg/mL-5 mg/mL, on the surface of an electrode, airing at 4 ℃, and washing with PBS (phosphate buffer solution) with pH = 7.5 to prepare the double-quenching competitive electrochemiluminescence immunosensor based on Au @ NiFe MOFs.
2. The double-quenching competitive electrochemiluminescence immunosensor prepared by the preparation method of claim 1 and based on Au @ NiFe MOFs is used for detecting diethylstilbestrol, and is characterized in that the detection steps are as follows:
(1) the method comprises the following steps of (1) testing by using a three-electrode system of an electrochemical workstation, taking an Ag/AgCl electrode as a reference electrode, taking a platinum wire electrode as a counter electrode, taking a prepared glassy carbon electrode modified by an electrochemiluminescence immunosensor as a working electrode, connecting the electrochemical workstation and a chemiluminescence detector together, setting the high voltage of a photomultiplier to be 500V, setting the cyclic voltammetry scanning potential range to be 0-1.4V, and setting the scanning rate to be 0.1V/s;
(2) in 10 mL of PBS (phosphate buffer solution) with pH of 6.0-8.5 and tripropylamine concentration of 1-12 mmol/L, detecting the intensity of an electrochemiluminescence signal generated by diethylstilbestrol standard solutions with different concentrations by an electrochemiluminescence method, and drawing a working curve;
(3) and (4) replacing the diethylstilbestrol standard solution with the diethylstilbestrol sample solution to be detected for determination.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201811578328.6A CN109521006B (en) | 2018-12-24 | 2018-12-24 | Preparation method and application of double-quenching competitive electrochemiluminescence sensor based on Au @ NiFe MOFs |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201811578328.6A CN109521006B (en) | 2018-12-24 | 2018-12-24 | Preparation method and application of double-quenching competitive electrochemiluminescence sensor based on Au @ NiFe MOFs |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN109521006A CN109521006A (en) | 2019-03-26 |
| CN109521006B true CN109521006B (en) | 2021-03-19 |
Family
ID=65795486
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201811578328.6A Active CN109521006B (en) | 2018-12-24 | 2018-12-24 | Preparation method and application of double-quenching competitive electrochemiluminescence sensor based on Au @ NiFe MOFs |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN109521006B (en) |
Families Citing this family (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110835091B (en) * | 2019-11-25 | 2021-05-14 | 浙江蓝能燃气设备有限公司 | PAN-based porous carbon-magnesium alloy loaded Zr-based MOFs hydrogen storage material and preparation method thereof |
| CN110988067B (en) * | 2019-12-13 | 2022-05-17 | 常州大学 | Electrochemical luminescence method for detecting diethylstilbestrol |
| CN111208178B (en) * | 2020-01-09 | 2022-12-06 | 济南大学 | Method for constructing electrochemical luminescence sensor based on double amplification of perylene tetracarboxylic acid signal by cobalt-based metal organic framework |
| CN111721825B (en) * | 2020-05-18 | 2022-08-09 | 济南大学 | Preparation method of lanthanide metal self-luminous Au @ Gd-MOFs-based signal enhancement type immunosensor |
| CN111721826B (en) * | 2020-05-18 | 2022-08-09 | 济南大学 | Preparation method of competitive immunosensor based on lanthanide metal self-luminescence Au @ Eu-MOFs |
| CN111721824B (en) * | 2020-05-18 | 2022-07-08 | 济南大学 | Preparation method of lanthanide metal self-luminous Au @ Tb-MOFs-based signal enhancement type immunosensor |
| CN111499880B (en) * | 2020-05-22 | 2022-03-18 | 中国农业科学院农业质量标准与检测技术研究所 | Composite nano material for dioxin enrichment and detection and preparation method thereof |
| CN113663646A (en) * | 2021-07-19 | 2021-11-19 | 中国计量大学 | Preparation method and application of wide-spectrum specific antibody modified magnetic metal organic framework material |
| CN114956161A (en) * | 2022-05-20 | 2022-08-30 | 郑州轻工业大学 | La doped In 2 O 3 Hexagonal prism-shaped nano material, preparation method thereof and application of hexagonal prism-shaped nano material as TEA sensor |
| CN115096963B (en) * | 2022-07-03 | 2023-06-23 | 济南大学 | Preparation method of electroluminescent sensor for detecting CD44 |
Citations (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102262125A (en) * | 2011-07-28 | 2011-11-30 | 南京师范大学 | Electrochemical immune sensor for detecting diethylstilbestrol and preparation method and application of sensor |
| CN104865242A (en) * | 2015-03-02 | 2015-08-26 | 济南大学 | Preparation method and application of mycotoxin and hormone electrogenerated chemiluminescence sensor constructed based on NPCo/Co3O4-Au/RuSi@Ru(bpy)3<2+> |
| CN105837610A (en) * | 2016-04-20 | 2016-08-10 | 浙江大学 | Preparation method of triazole functionalized Zr-MOFs material |
| WO2016161255A1 (en) * | 2015-04-01 | 2016-10-06 | The Board Of Trustees Of The University Of Illinois | Analyte sensing for eye injuries and conditions |
| CN106248657A (en) * | 2016-09-05 | 2016-12-21 | 济南大学 | A kind of based on dendroid Fe2o3the preparation method and application of the Electrochemiluminescsensor sensor of@Au |
| WO2017015669A1 (en) * | 2015-07-23 | 2017-01-26 | Adarza Biosystems, Inc. | Competitive small molecule detection assays using arrayed imaging reflectometry |
| CN106404756A (en) * | 2016-09-05 | 2017-02-15 | 济南大学 | Preparation method and application of electrochemiluminescence sensor based on graphene/Fe3O4@Au/CeO2/TiO2 |
| CN108918627A (en) * | 2018-08-01 | 2018-11-30 | 济南大学 | A kind of preparation method and application based on MOFs material competition type electrochemical immunosensor |
-
2018
- 2018-12-24 CN CN201811578328.6A patent/CN109521006B/en active Active
Patent Citations (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102262125A (en) * | 2011-07-28 | 2011-11-30 | 南京师范大学 | Electrochemical immune sensor for detecting diethylstilbestrol and preparation method and application of sensor |
| CN104865242A (en) * | 2015-03-02 | 2015-08-26 | 济南大学 | Preparation method and application of mycotoxin and hormone electrogenerated chemiluminescence sensor constructed based on NPCo/Co3O4-Au/RuSi@Ru(bpy)3<2+> |
| WO2016161255A1 (en) * | 2015-04-01 | 2016-10-06 | The Board Of Trustees Of The University Of Illinois | Analyte sensing for eye injuries and conditions |
| WO2017015669A1 (en) * | 2015-07-23 | 2017-01-26 | Adarza Biosystems, Inc. | Competitive small molecule detection assays using arrayed imaging reflectometry |
| CN105837610A (en) * | 2016-04-20 | 2016-08-10 | 浙江大学 | Preparation method of triazole functionalized Zr-MOFs material |
| CN106248657A (en) * | 2016-09-05 | 2016-12-21 | 济南大学 | A kind of based on dendroid Fe2o3the preparation method and application of the Electrochemiluminescsensor sensor of@Au |
| CN106404756A (en) * | 2016-09-05 | 2017-02-15 | 济南大学 | Preparation method and application of electrochemiluminescence sensor based on graphene/Fe3O4@Au/CeO2/TiO2 |
| CN108918627A (en) * | 2018-08-01 | 2018-11-30 | 济南大学 | A kind of preparation method and application based on MOFs material competition type electrochemical immunosensor |
Non-Patent Citations (2)
| Title |
|---|
| Quenching Electrochemiluminescence Immunosensor Based on Resonance Energy Transfer between Ruthenium (II) Complex Incorporated in the UiO-67 Metal−Organic Framework and Gold Nanoparticles for Insulin Detection;Guanhui Zhao 等;《ACS Appl. Mater. Interfaces》;20180619;全文 * |
| Ultrasensitive competitive method-based electrochemiluminescence immunosensor for diethylstilbestrol detection based on Ru(bpy)32+ as luminophor encapsulated in metal–organic frameworks UiO-67;Xue Dong 等;《Biosensors and Bioelectronics》;20180330;全文 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN109521006A (en) | 2019-03-26 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN109521006B (en) | Preparation method and application of double-quenching competitive electrochemiluminescence sensor based on Au @ NiFe MOFs | |
| CN108802133B (en) | A kind of preparation method and application detecting stomach neoplasms tumor markers interlayer type immunosensor | |
| KR101097882B1 (en) | Mesoporous Silica Conjugate Integrating Magnetic Nano Particles Having Peroxidase Activity and Enzymes and Method for Manufacturing the Same | |
| CN109444403B (en) | Ru (bpy) -based32+Preparation method of/Zn-oxalate immunosensor | |
| CN108663424B (en) | Preparation method and application of immunosensor based on sea urchin-shaped core-shell gold @ palladium nanospheres | |
| CN109839501B (en) | Electrochemiluminescence immunosensor for measuring circulating tumor cells and preparation method and application thereof | |
| CN109959691B (en) | Method for detecting nucleic acid based on cascade photoelectric active material and triple helix molecular switch | |
| CN107328930B (en) | A kind of preparation and application based on dual signal response ratio type screen printing electrode immunosensor | |
| CN104297479B (en) | The preparation of detection tumor markers electrochemiluminescimmunosensor immunosensor and application | |
| CN110455786B (en) | Based on CeO2@SnS2Preparation method of luminol-promoted electrochemiluminescence sensor | |
| CN107045010B (en) | The preparation method of optical electro-chemistry sensor based on the mesoporous carbonitride of stannic disulfide- | |
| CN109613244B (en) | Preparation method and application of Ag @ Pt-CuS labeled immunosensor | |
| CN108226252B (en) | Preparation method and application of current type immunosensor for detecting breast cancer | |
| CN106442994A (en) | Preparation method and application of electrochemical immunosensor based on Ag@Au nanocomposite | |
| CN106582848A (en) | Preparing method and application of mimic enzyme with double catalysis functions based on hemin mediation gold mineralization path | |
| CN110687175A (en) | Construction method of electrochemical luminescence sensor based on cerium dioxide and nano-silver dual-enhanced perylene tetracarboxylic acid luminescence | |
| CN110133252A (en) | For detecting kit and detection method and its application of carcinomebryonic antigen | |
| CN109946289A (en) | A kind of preparation method without mark type Electrochemiluminescsensor sensor based on self-luminescent material Ru@MOF-5 detection estradiol | |
| CN110927226A (en) | Construction method of electrochemical luminescence sensor based on cerium dioxide and nano platinum double-enhancement luminol luminescence | |
| CN108445213B (en) | Nano composite probe, composition and fluorescent quantitative kit for high-sensitivity fluorescent quantitative detection of serum tumor marker | |
| CN112710709A (en) | Cadmium sulfide quantum dot glassy carbon electrode for target DNA detection, preparation method thereof, electrochemical luminescence sensor system and application | |
| CN106645094B (en) | Electrochemical luminescence sensor for detecting concanavalin A by Fe3O4 immobilized Ru (bpy)32+ | |
| CN109406602B (en) | Preparation method and application of immunosensor based on echinoid hollow silver platinum palladium trimetal nanoparticles | |
| CN114034864B (en) | Method for detecting alpha fetoprotein by using aptamer fluorescent sensor constructed based on gold nanocomposite and polypeptide | |
| CN108132287B (en) | Preparation method and application of current type immunosensor based on polypyrrole nanosheet composite material |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |