CN106841617A - 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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- G01N27/3275—Sensing specific biomolecules, e.g. nucleic acid strands, based on an electrode surface reaction
- G01N27/3277—Sensing specific biomolecules, e.g. nucleic acid strands, based on an electrode surface reaction being a redox reaction, e.g. detection by cyclic voltammetry
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- G01N33/57484—Immunoassay; Biospecific binding assay; Materials therefor for cancer involving compounds serving as markers for tumor, cancer, neoplasia, e.g. cellular determinants, receptors, heat shock/stress proteins, A-protein, oligosaccharides, metabolites
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Abstract
The invention discloses a kind of preparation method and application at the electrochemical luminescence immune sensing interface based on conducting supramolecular gel, it is related to the fields such as nano science, biological immune technology, electrochemical sensing.The present invention is carrier using the conducting supramolecular gel that tannic acid and transition metal ions are formed, and realizes simple fixed and High Efficiency Luminescence of the electrochemical luminescence semiconductor nano material in electrode interface.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 the method, and operation is easy, and with electrochemical luminescence performance higher, solves nano material and the problems such as biomolecule lacks simple and effective electrode fixing means.The method goes for the preparation of various biomarker immunosensor electrodes, is with a wide range of applications in scientific research and clinic.
Description
Technical field
The present invention relates to fields such as nano science, biological immune technology, electrochemical sensings, and in particular to a kind of electrochemistry hair
The preparation method and application of light immunosensor sensing interface.
Background technology
Highly sensitive, high selectivity chemistry and biology sensor have in fields such as environmental monitoring, food security, clinical analysis
Have broad application prospects, in recent years the study 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 sensitivity it is high, be easy to miniaturization
The features such as show huge advantage in actual applications, and the electrode sensing interface of stable response is to build this kind of electrochemistry
The key of sensor.Nano material has special structure and resulting unique physics, chemical property because of it, 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 obvious effect, but nano material has that substrate selective is poor, low in itself.Therefore in structure
Mostly nano material is combined with bio-identification unit, but nano material equally exists biocompatibility when building sensing interface
The problems such as difference couples difficulty with biomolecule.With the rise of bionics concept, biomimetic chemistry gradually with nano science, life
The field such as life science, medical science, pharmacy Cross slot interference.Biomimetic chemistry is combined with nano structural material, nano bionic is built and is passed
Sense interface, can solve nano material problems present in sensor application, be development and the wound of electrochemical sensor
New thinking is newly provided, with highly important Research Significance.
The electrochemical immunosensor of early stage is generally marked come detection signal, but enzyme mark by carrying out enzyme 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
What the nano material with enzymatic activity built 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, with good
Application prospect.But still there are problems that nano material synthesis is difficult, lack simply and effective.To working electrode
Modification and the preparation of nano material be the key for building unmarked type electrochemical luminescence immunosensor.
Supermolecular gel has gradually developed into a class has the intelligence/functional nanomaterials of broad prospect of application.At present
What most of supermolecular gel of research was 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 not enough.Can carry out light to gel band after due to introducing metal ion
The new performances such as electricity, catalysis, redox, therefore the research of super-molecule metal gel also heated up in recent years.Based on super
Molecular metal gel builds electrochemical luminescence immunosensor sensing interface and has not been reported.
The present invention realizes electricity by the use of tannic acid and the super-molecule metal gel of transition metal ions formation as being carrier
Simple fixed and High Efficiency Luminescence of the chemiluminescence semiconductor nano material in electrode interface.While biomolecule tannic acid and metal
Antibody can be fixed on electrode by the good biocompatibility of titanium ion and supermolecular gel to the reactivity of large biological molecule
Surface.The method goes for the preparation at various biomarker electrochemical luminescence bio-sensing interfaces, in scientific research and clinic
In be with a wide range of applications.
The content of the invention
An object of the present invention is to provide unmarked electrochemical luminescence immunosensor sensing interface simple and easy to apply
The problems such as preparation method, solution electrode modification step complexity, poor reproducibility.
The second object of the present invention is to provide and provides quick, inexpensive, general biological marker object detecting method, is electrification
Learn 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, it is characterised in that comprise the following steps：
（1）By 100 μ L10mgmL-1Graphene oxide or carbon nano-tube aqueous solutions, 100 μ L10mgmL-1Carbonitride
The nanometer sheet aqueous solution and 200 μ L mass fractions are 30 mgmL-1Tannin aqueous acid mixing, ultrasonic vibration 5 minutes, use
50 μ L concentration are 1 molL-1Sodium hydroxide solution pH is adjusted to 7；
（2）Above-mentioned mixed solution and the ammonium of two (2 hydroxy propanoic acid) two hydroxide two that 300 μ L mass fractions are 40% are closed into titanium
The aqueous solution mixes, and be vortexed concussion 1 minute, obtains the colloidal sol of nano material doping；
（3）By 10 μ L steps（2）Obtained colloidal sol drops in treatment, the glassy carbon electrode surface of good a diameter of 4 mm of activation, 30 minutes
The glass-carbon electrode of Signa Gel modification is obtained afterwards；
（4）5 μ L biomarkers are captured into antibody-solutions with liquid-transfering gun（10 µg·mL-1）Drop coating is in step（3）Obtained electricity
Pole surface, dries under the conditions of 4 DEG C；
（5）With the multiple rinsing step of ultra-pure water（4）Obtained electrode, 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）Obtained electrode surface, dries under the conditions of 4 DEG C, and ultrapure water obtains immune sensing circle
Face.
Useful achievement of the invention
Not only step is simple for the 1 sensing interface preparation method, and operation is easy, with low cost, and is sent out with electrochemistry higher
Optical property, the problems such as solving nano material and biomolecule and 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, it is to avoid
Complicated antibody fixation procedure and the deactivation prob for thus causing.
3 the method go for the preparation of various 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 expanded on further.It should be understood that these embodiments are merely to illustrate the present invention
Rather than limitation the scope of the present invention.
Embodiment 1（1）By 100 μ L10mgmL-1Graphene oxide water solution, 100 μ L10mgmL-1Nitridation
The carbon nanosheet aqueous solution and 200 μ L mass fractions are 30 mgmL-1Tannin aqueous acid mixing, ultrasonic vibration 5 minutes,
It is 1 molL with 50 μ L concentration-1Sodium hydroxide solution pH is adjusted to 7；
（2）Above-mentioned mixed solution and the ammonium of two (2 hydroxy propanoic acid) two hydroxide two that 300 μ L mass fractions are 40% are closed into titanium
The aqueous solution mixes, and be vortexed concussion 1 minute, obtains the colloidal sol of nano material doping；
（3）By 10 μ L steps（2）Obtained colloidal sol drops in treatment, the glassy carbon electrode surface of good a diameter of 4 mm of activation, 30 minutes
The glass-carbon electrode of Signa Gel modification is obtained afterwards；
（4）5 μ L PSAs are captured into antibody-solutions with liquid-transfering gun（10 µg·mL-1）Drop coating is in step（3）System
The electrode surface for obtaining, dries under the conditions of 4 DEG C；
（5）With the multiple rinsing step of ultra-pure water（4）Obtained electrode, 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）Obtained electrode surface, dries, ultrapure water under the conditions of 4 DEG C；
（6）With the multiple rinsing step of ultra-pure water（5）The electrochemical luminescence immunosensor sensing interface of structure, uses liquid-transfering gun drop coating
5 μ L PSAs standard liquids or unknown sample solution, as working electrode after being dried under the conditions of 4 DEG C, Ag/
AgCl electrodes as reference electrode, Pt electrodes as to electrode, with 20 mmol of the PBS buffer preparations that pH is 7.4
mL-1K2S2O8Solution is determined under -1.2~0 V potentials on electrochemical luminescence work station as bottom liquid with cyclic voltammetric pattern
Luminous signal, realize detection to PSA.
Embodiment 2（1）By 100 μ L10mgmL-1Carbon nano-tube aqueous solutions, 100 μ L10mgmL-1Carbonitride
The nanometer sheet aqueous solution and 200 μ L mass fractions are 30 mgmL-1Tannin aqueous acid mixing, ultrasonic vibration 5 minutes, use
50 μ L concentration are 1 molL-1Sodium hydroxide solution pH is adjusted to 7；
（2）Above-mentioned mixed solution and the ammonium of two (2 hydroxy propanoic acid) two hydroxide two that 300 μ L mass fractions are 40% are closed into titanium
The aqueous solution mixes, and be vortexed concussion 1 minute, obtains the colloidal sol of nano material doping；
（3）By 10 μ L steps（2）Obtained colloidal sol drops in treatment, the glassy carbon electrode surface of good a diameter of 4 mm of activation, 30 minutes
The glass-carbon electrode of Signa Gel modification is obtained afterwards；
（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）It is obtained
Electrode surface, dried under the conditions of 4 DEG C；
（5）With the multiple rinsing step of ultra-pure water（4）Obtained electrode, 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）Obtained electrode surface, dries, ultrapure water under the conditions of 4 DEG C；
（6）With the multiple rinsing step of ultra-pure water（5）The electrochemical luminescence immunosensor sensing interface of structure, uses liquid-transfering gun drop coating
5 μ L breast cancer susceptibility genes standard liquids or unknown sample solution, as working electrode after being dried under the conditions of 4 DEG C, Ag/AgCl
Electrode as reference electrode, Pt electrodes as to electrode, with 20 mmolmL of the PBS buffer preparations that pH is 7.4-1's
K2S2O8Solution determines luminous under -1.2~0 V potentials on electrochemical luminescence work station as bottom liquid with cyclic voltammetric pattern
Signal, realizes the detection to breast cancer susceptibility gene.
Claims (3)
1. a kind of preparation method of electrochemical luminescence immunosensor sensing interface, it is characterised in that comprise the following steps：
（1）By 100 μ L10mgmL-1Graphene oxide or carbon nano-tube aqueous solutions, 100 μ L10mgmL-1Carbonitride
The nanometer sheet aqueous solution and 200 μ L mass fractions are 30 mgmL-1Tannin aqueous acid mixing, ultrasonic vibration 5 minutes, use
50 μ L concentration are 1 molL-1Sodium hydroxide solution pH is adjusted to 7；
（2）Above-mentioned mixed solution and the ammonium of two (2 hydroxy propanoic acid) two hydroxide two that 300 μ L mass fractions are 40% are closed into titanium
The aqueous solution mixes, and be vortexed concussion 1 minute, obtains the colloidal sol of nano material doping；
（3）By 10 μ L steps（2）Obtained colloidal sol drops in treatment, the glassy carbon electrode surface of good a diameter of 4 mm of activation, 30 minutes
The glass-carbon electrode of Signa Gel modification is obtained afterwards；
（4）5 μ L biomarkers are captured into antibody-solutions with liquid-transfering gun（10 µg·mL-1）Drop coating is in step（3）Obtained electrode
Surface, dries under the conditions of 4 DEG C；
（5）With the multiple rinsing step of ultra-pure water（4）Obtained electrode, 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）Obtained electrode surface, dries 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, its feature exists
In described biomarker is carcinomebryonic antigen, PSA, 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
Sense interface is used for biological marker analyte detection, it is characterised 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 the μ L biomarkers standard liquids of liquid-transfering gun drop coating 5 or unknown sample solution, 4
As working electrode after being dried under the conditions of DEG C, Ag/AgCl electrodes, as to electrode, are 7.4 with pH as reference electrode, Pt electrodes
PBS buffer preparations 20 mmolmL-1K2S2O8Solution as bottom liquid, on electrochemical luminescence work station with circulate
Volt-ampere pattern determines the luminous signal under -1.2~0 V potentials, realizes the detection to biomarker.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| 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 |
| CN110702749A (en) * | 2019-11-04 | 2020-01-17 | 太原理工大学 | Method for constructing electrochemical immunosensing interface based on conductive gel with active sites |
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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 |
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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 |
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| US20040214220A1 (en) * | 2001-03-28 | 2004-10-28 | Liaohai Chen | Tuning the properties of conjugated polyelectrolytes and application in a biosensor platform |
| 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 |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| 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 |
| CN110702749A (en) * | 2019-11-04 | 2020-01-17 | 太原理工大学 | Method for constructing electrochemical immunosensing interface based on conductive gel with active sites |
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