CN106841617B - A kind of preparation method and application of electrochemical luminescence immunosensor sensing interface - Google Patents
A kind of preparation method and application of electrochemical luminescence immunosensor sensing interface Download PDFInfo
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- CN106841617B CN106841617B CN201710090224.XA CN201710090224A CN106841617B CN 106841617 B CN106841617 B CN 106841617B CN 201710090224 A CN201710090224 A CN 201710090224A CN 106841617 B CN106841617 B CN 106841617B
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Abstract
The invention discloses a kind of preparation method and applications at the electrochemical luminescence immune sensing interface based on conducting supramolecular gel, are related to the fields such as nano science, biological immune technology, electrochemical sensing.The present invention is carrier using tannic acid and the conducting supramolecular gel that transition metal ions is formed, and realizes electrochemical luminescence semiconductor nano material in the simple fixation of electrode interface and High Efficiency Luminescence.Antibody can be fixed on electrode surface by biomolecule tannic acid with the good biocompatibility of metal titanium ion and supermolecular gel to the reactivity of large biological molecule simultaneously.Not only step is simple for this method, and operation is easy, and with higher electrochemical luminescence performance, solves nano material and the problems such as biomolecule lacks simple and effective electrode fixing means.This method can be adapted for the preparation of a variety of biomarker immunosensor electrodes, be with a wide range of applications in scientific research and clinic.
Description
Technical field
The present invention relates to the fields such as nano science, biological immune technology, electrochemical sensing, and in particular to a kind of electrochemistry hair
The preparation method and application of light immunosensor sensing interface.
Background technology
Highly sensitive, highly selective chemistry and biosensor has in fields such as environmental monitoring, food security, clinical analysis
Have broad application prospects, in recent years the research hotspot of always domestic and international analytical chemistry worker.Wherein it is based on electrode process
Electrochemical sensor, electrochemical luminescence sensor and optical electro-chemistry sensor etc. due to high sensitivity, being easy to minimize
The features such as show huge advantage in practical applications, and the electrode sensing interface of stable response is this kind of electrochemistry of structure
The key of sensor.Nano material because it is with special structure and resulting unique physics, chemical property, by
It is widely used for the structure of electrochemical sensor.Although nano structural material is in signal transduction, signal amplification, artificial mimic enzyme etc.
Aspect has an obvious effect, but there are the problems such as substrate selective is poor, low biometric identification capabilities in itself for nano material.Therefore in structure
Nano material is combined, but nano material equally exists biocompatibility with bio-identification unit mostly when building sensing interface
Difference couples the problems such as difficulty with biomolecule.With the rise of bionics concept, biomimetic chemistry gradually with nano science, life
The fields such as life science, medicine, pharmacy Cross slot interference.Biomimetic chemistry is combined with nano structural material, structure nano bionic passes
Feel interface, nano material problems present in sensor application can be solved, be development and the wound of electrochemical sensor
New thinking is newly provided, there is highly important research significance.
The electrochemical immunosensor of early stage usually detects signal, but enzyme mark by carrying out enzyme label to antigen or antibody
During operating cost, and enzyme is higher to environmental requirement, is particularly easily inactivated after enzyme is attached to solid carrier surface.Therefore it is based on
Nano material structure with enzymatic activity causes the great research interest of people without enzyme electrochemical immunosensor.And
Wherein unmarked type electrochemical immunosensor has that step is simple, fast and easy, high repeatability and other advantages again, has good
Application prospect.But the problems such as still remaining difficult nano material synthesis, shortage simple and effective fixing means.To working electrode
Modification and the preparation of nano material be the key that the unmarked type electrochemical luminescence immunosensor of structure.
Supermolecular gel has gradually developed into a kind of intelligence/functional nanomaterials with broad prospect of application.At present
What most of supermolecular gel of research utilized be hydrogen bond or packing of molecules effect, and coordinate bond this in supramolecular chemistry
The application acted in gel synthesis of no less important is then much insufficient.Since after introducing metal ion light can be carried out to gel band
The new performances such as electricity, catalysis, redox, therefore the research of super-molecule metal gel is also heated up in recent years.Based on super
Molecular metal gel structure electrochemical luminescence immunosensor sensing interface has not been reported.
The present invention, as carrier, realizes electricity using the super-molecule metal gel of tannic acid and transition metal ions formation
Chemiluminescence semiconductor nano material is in the simple fixation of electrode interface and High Efficiency Luminescence.Biomolecule tannic acid and metal simultaneously
Antibody can be fixed on electrode by the good biocompatibility of titanium ion and supermolecular gel to the reactivity of large biological molecule
Surface.This method can be adapted for the preparation at a variety of biomarker electrochemical luminescence bio-sensing interfaces, in scientific research and clinic
In be with a wide range of applications.
Invention content
An object of the present invention is to provide simple and practicable unmarked electrochemical luminescence immunosensor sensing interface
The problems such as preparation method, solution electrode modification step is complicated, poor reproducibility.
The second object of the present invention is to provide to provide quick, low cost, general biological marker object detecting method, for electrification
It learns application of the electrochemiluminescent immunoassay sensor in clinic and technical foundation is provided.
Technical scheme is as follows：
1. a kind of preparation method of electrochemical luminescence immunosensor sensing interface, which is characterized in that including following step
Suddenly：
（1）By 100 μ L10mgmL-1Graphene oxide or carbon nano-tube aqueous solutions, 100 μ L10mgmL-1Nitrogen
It is 30 mgmL to change carbon nanosheet aqueous solution and 200 μ L mass fractions-1Tannic acid aqueous solution mixing, ultrasonic vibration 5 divides
Clock, with a concentration of 1 molL of 50 μ L-1Sodium hydroxide solution pH is adjusted to 7；
（2）Above-mentioned mixed solution and 300 μ L mass fractions are closed for 40% two (2 hydroxy propanoic acid) two hydroxide diammonium
The aqueous solution mixing of titanium, be vortexed concussion 1 minute, obtains the colloidal sol of nano material doping；
（3）By 10 μ L steps（2）Colloidal sol obtained drops in processing, the glassy carbon electrode surface of a diameter of 4 mm is got well in activation, and 30
The glass-carbon electrode of Signa Gel modification is obtained after minute；
（4）5 μ L biomarkers are captured into antibody-solutions with liquid-transfering gun（10 µg·mL-1）Drop coating is in step（3）It is made
Electrode surface, dry under the conditions of 4 DEG C；
（5）With the multiple rinsing step of ultra-pure water（4）Electrode obtained, with liquid-transfering gun by 6 μ L mass fractions be 1% ox blood
Pure protein solution drop coating is in step（4）Electrode surface obtained is dried under the conditions of 4 DEG C, and ultrapure water obtains immune sensing
Interface.
The useful achievement of the present invention
Not only step is simple for the 1 sensing interface preparation method, and operation is easy, of low cost, and with higher electrification
Luminescent properties are learned, solve the problems such as nano material and biomolecule lack simple and effective electrode fixing means.
The good biocompatibility of 2 tannic acid supermolecular gels and reactivity are convenient for the fixation of antibody, avoid
The antibody fixation procedure of complexity and thus caused deactivation prob.
3 this method can be adapted for the preparation of a variety of biomarker immunosensor sensing interfaces, in scientific research and clinic
In be with a wide range of applications.
Specific embodiment
With reference to specific embodiment, the present invention is further explained.It should be understood that these embodiments are merely to illustrate the present invention
Rather than it limits the scope of the invention.
Embodiment 1（1）By 100 μ L10mgmL-1Graphene oxide water solution, 100 μ L10mgmL-1Nitridation
Carbon nanosheet aqueous solution and 200 μ L mass fractions are 30 mgmL-1Tannic acid aqueous solution mixing, ultrasonic vibration 5 minutes,
With a concentration of 1 molL of 50 μ L-1Sodium hydroxide solution pH is adjusted to 7；
（2）Above-mentioned mixed solution and 300 μ L mass fractions are closed for 40% two (2 hydroxy propanoic acid) two hydroxide diammonium
The aqueous solution mixing of titanium, be vortexed concussion 1 minute, obtains the colloidal sol of nano material doping；
（3）By 10 μ L steps（2）Colloidal sol obtained drops in processing, the glassy carbon electrode surface of a diameter of 4 mm is got well in activation, and 30
The glass-carbon electrode of Signa Gel modification is obtained after minute；
（4）5 μ L prostate-specific antigen are captured into antibody-solutions with liquid-transfering gun（10 µg·mL-1）Drop coating is in step
（3）Electrode surface obtained is dried under the conditions of 4 DEG C；
（5）With the multiple rinsing step of ultra-pure water（4）Electrode obtained, with liquid-transfering gun by 6 μ L mass fractions be 1% ox blood
Pure protein solution drop coating is in step（4）Electrode surface obtained is dried under the conditions of 4 DEG C, ultrapure water；
（6）With the multiple rinsing step of ultra-pure water（5）The electrochemical luminescence immunosensor sensing interface of structure, uses liquid-transfering gun
5 μ L prostate-specific antigen standard solution of drop coating or unknown sample solution, as working electrode after drying under the conditions of 4 DEG C,
As reference electrode, Pt electrodes are used as to electrode Ag/AgCl electrodes, with the 20 of the PBS buffer preparations that pH is 7.4
mmol·mL-1K2S2O8Solution measures -1.2~0 V on electrochemical luminescence work station as bottom liquid with cyclic voltammetric pattern
Luminous signal under potential realizes the detection to prostate-specific antigen.
Embodiment 2（1）By 100 μ L10mgmL-1Carbon nano-tube aqueous solutions, 100 μ L10mgmL-1Carbonitride
Nanometer sheet aqueous solution and 200 μ L mass fractions are 30 mgmL-1Tannic acid aqueous solution mixing, ultrasonic vibration 5 minutes, use
A concentration of 1 molL of 50 μ L-1Sodium hydroxide solution pH is adjusted to 7；
（2）Above-mentioned mixed solution and 300 μ L mass fractions are closed for 40% two (2 hydroxy propanoic acid) two hydroxide diammonium
The aqueous solution mixing of titanium, be vortexed concussion 1 minute, obtains the colloidal sol of nano material doping；
（3）By 10 μ L steps（2）Colloidal sol obtained drops in processing, the glassy carbon electrode surface of a diameter of 4 mm is got well in activation, and 30
The glass-carbon electrode of Signa Gel modification is obtained after minute；
（4）5 μ L breast cancer susceptibility genes are captured into antibody-solutions with liquid-transfering gun（10 µg·mL-1）Drop coating is in step（3）
Electrode surface obtained is dried under the conditions of 4 DEG C；
（5）With the multiple rinsing step of ultra-pure water（4）Electrode obtained, with liquid-transfering gun by 6 μ L mass fractions be 1% ox blood
Pure protein solution drop coating is in step（4）Electrode surface obtained is dried under the conditions of 4 DEG C, ultrapure water；
（6）With the multiple rinsing step of ultra-pure water（5）The electrochemical luminescence immunosensor sensing interface of structure, uses liquid-transfering gun
5 μ L breast cancer susceptibility genes standard solution of drop coating or unknown sample solution, as working electrode, Ag/ after drying under the conditions of 4 DEG C
As reference electrode, Pt electrodes are used as to electrode AgCl electrodes, with 20 mmol of the PBS buffer preparations that pH is 7.4
mL-1K2S2O8Solution is measured with cyclic voltammetric pattern under -1.2~0 V potentials as bottom liquid on electrochemical luminescence work station
Luminous signal, realize the detection to breast cancer susceptibility gene.
Claims (3)
1. a kind of preparation method of electrochemical luminescence immunosensor sensing interface, which is characterized in that include the following steps：
（1）By 100 μ L10mgmL-1Graphene oxide or carbon nano-tube aqueous solutions, 100 μ L10mgmL-1Carbonitride
Nanometer sheet aqueous solution and 200 μ L mass fractions are 30 mgmL-1Tannic acid aqueous solution mixing, ultrasonic vibration 5 minutes, use
A concentration of 1 molL of 50 μ L-1Sodium hydroxide solution pH is adjusted to 7；
（2）Above-mentioned mixed solution and 300 μ L mass fractions are closed into titanium for 40% two (2 hydroxy propanoic acid) two hydroxide diammonium
Aqueous solution mixes, and be vortexed concussion 1 minute, obtains the colloidal sol of nano material doping；
（3）By 10 μ L steps（2）Colloidal sol obtained drops in processing, the glassy carbon electrode surface of a diameter of 4 mm is got well in activation, 30 minutes
The glass-carbon electrode of Signa Gel modification is obtained afterwards；
（4）With liquid-transfering gun by a concentration of 10 μ gmL of 5 μ L-1Biomarker captures antibody-solutions drop coating in step（3）It is made
Electrode surface, dry under the conditions of 4 DEG C；
（5）With the multiple rinsing step of ultra-pure water（4）Electrode obtained, it is with liquid-transfering gun that the ox blood that 6 μ L mass fractions are 1% is pure
Protein solution drop coating is in step（4）Electrode surface obtained is dried under the conditions of 4 DEG C, and ultrapure water obtains immune sensing circle
Face.
2. a kind of preparation method of electrochemical luminescence immunosensor sensing interface according to claim 1, feature exist
In the biomarker is carcinomebryonic antigen, prostate-specific antigen, glycoprotein antigen, alpha-fetoprotein, breast cancer susceptibility
One kind in gene.
3. a kind of preparation method of electrochemical luminescence immunosensor sensing interface according to claim 1, the biography of preparation
Feel interface and be used for biological marker analyte detection, which is characterized in that step is as follows：With the multiple rinsing step of ultra-pure water（5）The electricity of structure
Chemiluminescence immunoassay sensor senses interface, with 5 μ L biomarkers standard solution of liquid-transfering gun drop coating or unknown sample solution, 4
As working electrode after being dried under the conditions of DEG C, as reference electrode, Pt electrodes are used as to electrode Ag/AgCl electrodes, are 7.4 with pH
PBS buffer preparations 20 mmolmL-1K2S2O8Solution is as bottom liquid, with cycle on electrochemical luminescence work station
Volt-ampere pattern measures the luminous signal under -1.2~0 V potentials, realizes the detection to biomarker.
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| CN107764880A (en) * | 2017-10-11 | 2018-03-06 | 浙江海洋大学 | A kind of biosensor thin layer electrode of quick detection miocardial infarction protein marker |
| CN110702749B (en) * | 2019-11-04 | 2021-11-09 | 太原理工大学 | Method for constructing electrochemical immunosensing interface based on conductive gel with active sites |
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| CN102507953A (en) * | 2011-10-20 | 2012-06-20 | 济南大学 | Preparation method of electrochemistry immunosensor for determining alpha fetoprotein |
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| CN103245656A (en) * | 2013-04-25 | 2013-08-14 | 济南大学 | Preparation and application of alpha fetoprotein and carcino-embryonic antigen electrochemiluminescence sensor |
Family Cites Families (1)
| Publication number | Priority date | Publication date | Assignee | Title |
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| US6737279B1 (en) * | 2001-03-28 | 2004-05-18 | The Regents Of The University Of California | Tuning the properties of conjugated polyelectrolytes and application in a biosensor platform |
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| KR20070014292A (en) * | 2005-07-28 | 2007-02-01 | 주식회사 아이센스 | Electrochemical determination system of glycated proteins |
| CN102507953A (en) * | 2011-10-20 | 2012-06-20 | 济南大学 | Preparation method of electrochemistry immunosensor for determining alpha fetoprotein |
| CN102778453A (en) * | 2012-08-08 | 2012-11-14 | 济南大学 | Manufacture method and application of silver hybridization SBA-15 electrochemical luminescence immunosensor |
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