CN102253094B - Electromagnetic control electrode and preparation method thereof, and detection method of biological sample - Google Patents
Electromagnetic control electrode and preparation method thereof, and detection method of biological sample Download PDFInfo
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- CN102253094B CN102253094B CN 201110116774 CN201110116774A CN102253094B CN 102253094 B CN102253094 B CN 102253094B CN 201110116774 CN201110116774 CN 201110116774 CN 201110116774 A CN201110116774 A CN 201110116774A CN 102253094 B CN102253094 B CN 102253094B
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Abstract
The invention discloses an electromagnetic control electrode, which is in a multilayer hollow tubular structure and comprises, from inside to outside, a PVDF (Polyvinylidene Fluoride) layer, a copper foil layer, an insulating layer, a PTME (polytetramethylene ether glycol dicarboxylate) layer and an outer cavity layer made of a PTFM material. One end face of the electromagnetic control electrode is used as an electrode working surface, and the other end is enclosed; and the electrode working surface has a gold sheet structure and is led out by leads and connected with an external power supply terminal. The electromagnetic control electrode simultaneously utilizes a layered structure to built an electromagnetic structure inside thereof; functionalized magnetic particles are absorbed on the working electrode surface to change a complex modification process of a traditional electrode, shorten detection time substantially and realize an electrochemical biological detection in a reagent approach and renewable and repetitive usage. The invention also discloses a preparation method of the electromagnetic control electrode and a biological sample detection method utilizing the electromagnetic control electrode.
Description
Technical field
The present invention relates to the biosensor technology field, be particularly related to a kind of biology sensor that is specially adapted to the sub-sessile antibody of novel functional Fe_3 O_4 magnetic particles (or antigen), also relate to the preparation method of this electromagnetic control electrode, and utilize electromagnetic control electrode to carry out the method for biology sample detection.
Background technology
Cyclic voltammetry is a kind of electrochemical research method of great use, can be used for the research of character, mechanism and the Kinetics parameters of electrode processes of electrode reaction.At present, the working electrode that electrochemical cyclic voltammetry detects in the three-electrode system used adopts gold electrode, platinum electrode, glass-carbon electrode etc. substantially, its complicated process of preparation, when at these electrode face finish magnetic materials, need to use traditional modification method, the reproducibility of its process complexity and electrode is poor, need the electrode surface of polishing after detection, and carry out loaded down with trivial details processing, length consuming time, can not meet the requirement of modern clinic fast detecting, can not well give prominence to the superiority of magnetic material simultaneously.
Summary of the invention
In view of this, one of purpose of the present invention has been to provide a kind of electromagnetic control electrode, its workplace adopts the proof gold material, utilize hierarchy section's structure electromagnetic structure within it simultaneously, by functionalized magnetic particles is adsorbed on to working electrode surface, changed the modification of traditional electrode complexity, greatly shorten detection time, realize the reagent that electrochemica biological detects, and renewable reusing; Two of purpose of the present invention has been to provide the preparation method of this electromagnetic control electrode; Three of purpose of the present invention has been to provide a kind of method of utilizing electromagnetic control electrode to carry out biology sample detection.
One of purpose of the present invention is achieved through the following technical solutions:
This electromagnetic control electrode is the multilayer hollow tubular structures, comprises respectively from inside to outside PVDF layer I, copper foil layer, insulation course, PVDF layer II and external cavity layer, and described external cavity layer adopts the PVDF material to make;
One end end face of described electromagnetic control electrode is as electrode face, and the other end seals, and described electrode face is piece structure, and described electrode face utilizes wire to draw, and with the external power source binding post, is connected;
Further, the wall thickness of described electromagnetic control electrode is 0.5 ~ 2 mm, and external diameter is 8 ~ 10mm, is highly 80 ~ 110mm;
Further, the external diameter of described piece structure is 3 ~ 4 mm, and thickness is 0.5 ~ 0.6 mm;
Further, total wall thickness of the sandwich construction of described electromagnetic control electrode is less than 2.15 mm.
Two of purpose of the present invention is achieved through the following technical solutions:
The preparation method of this electromagnetic control electrode comprises the following steps:
(1) adopt the PVDF material to prepare the external cavity layer of electrode;
(2) an end end face of external cavity layer adopts golden material to be prepared with electrode face, adopts wire to draw, and it is connected with the external power source binding post;
(3) interior laminate layer of cavity layer is assembled each layer structure outside, is followed successively by from inside to outside PVDF layer I, copper foil layer, insulation course and PVDF layer II, and gross thickness is less than 2.15 mm;
Further, the wall thickness of described electromagnetic control electrode is 1.5 ~ 2 mm, and external diameter is 8 ~ 10mm, is highly 80 ~ 110mm;
Further, the external diameter of described electrode face is 3 ~ 4 mm, and thickness is 0.5 ~ 0.6mm.
Three of purpose of the present invention is achieved through the following technical solutions:
Utilize foregoing electromagnetic control electrode to carry out the method for biology sample detection, comprise the following steps:
(1) nano combined magnetic bead preparation: Prussian blue, nm of gold are modified successively in the magnetic particle sub-surface, and then fixing alpha-fetoprotein specific antibody, form nano combined magnetic bead;
(2) cyclic voltammetric detects: by the electromagnetic control electrode energising, under the attraction of externally-applied magnetic field, in conjunction with nano combined magnetic bead be absorbed and fixed at the magnetic control electrode surface, electrode is placed in to pH 6.5 PBS solution and carries out the cyclic voltammetric detection, obtain electric current I
1;
(3) hatch: the AFP standard items are diluted to respectively to 0.5 ng/mL, 1.0 ng/mL, 2.0 ng/mL, 5.0 ng/mL, 10 ng/mL, 25 ng/mL, 50 ng/mL, 75 ng/mL, 100 ng/mL, 150 ng/mL, 200 ng/mL, 250 ng/mL and 300ng/mL, the AFP standard items that nano combined magnetic bead are placed in successively to above-mentioned variable concentrations are hatched, and incubation time is 15 ~ 30min;
(4) detect: by the electromagnetic control electrode energising, and the electrode front end is placed in and hatches solution, after absorption, electromagnetic control electrode is moved to and detects in solution, and the solution of pH 6.5 PBS, carry out the cyclic voltammetry detection, the amount of its peak current and adsorption antigen is inversely proportional to, and wherein, peak point current is designated as I
2, the current value I=I descended with oxidation peak current before and after immune response
2-I
1as quantitative basis, to each concentration standard antigen duplicate detection three times, using its mean value current value corresponding as this antigen concentration, and the current value corresponding with it carries out linear regression analysis to antigen concentration;
Further, also comprise step (5),, after detecting, outage disappears the magnetic of electromagnetic control electrode, and the PBS flushing is carried out in the surface of electromagnetic control electrode, and nano combined magnetic bead can break away from from the electromagnetic control electrode surface under the PBS washing action;
Further, the incubation time in described step (3) is 20 min.
The invention has the beneficial effects as follows:
1. electromagnetic control electrode of the present invention, its workplace adopts the proof gold material, in electrode interior, adopt hierarchy to build calutron, by functionalized magnetic particles by being adsorbed on working electrode surface, changed the modification of traditional electrode complexity, greatly shorten detection time, realize the reagent that electrochemica biological detects, and can exempt from regeneration and use;
2. to take alpha-fetoprotein (AFP) be model to detection method of the present invention, utilize the specificity affinity interaction of Ag-Ab to catch the determined antigen in serum specimen, the curent change that functionalized magnetic particles that adopts cyclic voltammetry to detect the immune response front and back produces, realized the quantitative detection of AFP in serum, the method is simple to operate, it is quick, highly sensitive to detect, testing cost is low, exempt from regeneration, be easy to promote, and can be used as clinically by detecting a kind of quick, the effective detection method of AFP content in serum.
Other advantages of the present invention, target and feature will be set forth to a certain extent in the following description, and to a certain extent, based on will be apparent to those skilled in the art to investigating hereinafter, or can be instructed from the practice of the present invention.Target of the present invention and other advantages can realize and obtain by following instructions and claims.
The accompanying drawing explanation
In order to make the purpose, technical solutions and advantages of the present invention clearer, below in conjunction with accompanying drawing, the present invention is described in further detail, wherein:
The cross-sectional view that Fig. 1 is electromagnetic control electrode of the present invention;
The external structure schematic diagram that Fig. 2 is electromagnetic control electrode of the present invention;
Fig. 3 is that Fig. 3 magnetic control electrochemical immunosensor detects schematic diagram; (wherein I is the detected peaks current value)
Fig. 4 detects linear matched curve figure for being applied to AFP.
Embodiment
Hereinafter with reference to accompanying drawing, the preferred embodiments of the present invention are described in detail.Should be appreciated that preferred embodiment is only for the present invention is described, rather than in order to limit the scope of the invention.
As shown in Figure 1, 2, electromagnetic control electrode of the present invention adopts the multilayer hollow tubular structures, comprises respectively from inside to outside PVDF layer I 1, copper foil layer 2, insulation course 3, PVDF layer II 4 and external cavity layer 5, and described external cavity layer 5 adopts the PVDF material to make; In the present embodiment, PVDF layer II 4 is preferably the PTME material and makes, and the external cavity layer is preferably the PTFM material and makes.
The PVDF resin mainly refers to the multipolymer of vinylidene fluoride homopolymer or vinylidene and other a small amount of fluorine-containing vinyl monomers, the PVDF resin has the characteristic of fluororesin and resins for universal use concurrently, except thering is good chemical resistance, heat-resisting quantity, oxidative resistance, weatherability, outside anti-x radiation x performance, also there is piezoelectricity, dielectricity, the properties such as pyroelectricity, it is the large product that in current fluoro-containing plastic, output is at the second place, application mainly concentrates on petrochemical complex, electric and the large field of fluorocarbon coating three, due to the good chemical resistance of PVDF, processability and antifatigue and creep properties, the perforated membrane that adopts in recent years the PVDF resin to make, gel, barrier film etc., be widely used.PTFM and PTME material are all belong to the PVDF resin a kind of, are the new materials with excellent physical and chemical property of abroad developing.
One end end face of electromagnetic control electrode is as electrode face 6, and the other end seals, and described electrode face 6 is piece structure, and described electrode face 6 utilizes wire to draw, and with the external power source binding post, is connected.
In the present embodiment, the wall thickness of the external cavity layer of described electromagnetic control electrode is 1 mm, and external diameter is 10mm, is highly 80 mm.; The external diameter of described piece structure is 4mm, and thickness is 2 mm.Total wall thickness of the sandwich construction of described electromagnetic control electrode is less than 2.15 mm.
The modification of this electromagnetic control electrode is by the electromagnetic control electrode energising, under the attraction of externally-applied magnetic field, makes the nano combined magnetic bead of combination be absorbed and fixed at the magnetic control electrode surface; The making of nano combined magnetic bead is that Prussian blue, nm of gold are modified successively in the magnetic particle sub-surface, fixing alpha-fetoprotein specific antibody then, thus form nano combined magnetic bead.
The preparation method of this electromagnetic control electrode mainly comprises the following steps:
(1) adopt the PVDF material to prepare the external cavity layer of electrode;
(2) an end end face of external cavity layer adopts golden material to be prepared with electrode face, adopts wire to draw, and it is connected with the external power source binding post;
(3) interior laminate layer of cavity layer is assembled each layer structure outside, is followed successively by from inside to outside PVDF layer I, copper foil layer, insulation course and PVDF layer II, and gross thickness is less than 2.15 mm.
Wherein, PVDF layer II 4 is preferably the PTME material and makes, and the external cavity layer is preferably the PTFM material and makes.
As shown in Figure 3, utilize this electromagnetic control electrode to carry out the method for biology sample detection, mainly comprise the following steps:
(1) nano combined magnetic bead preparation: Prussian blue, nm of gold are modified successively in the magnetic particle sub-surface, and then fixing alpha-fetoprotein specific antibody, form nano combined magnetic bead;
(2) cyclic voltammetric detects: by electromagnetic control electrode energising of the present invention, under the attraction of externally-applied magnetic field, in conjunction with nano combined magnetic bead be absorbed and fixed at the magnetic control electrode surface, electrode is placed in to pH 6.5 PBS solution and carries out the cyclic voltammetric detection, obtain electric current I
1;
(3) hatch: the AFP standard items are diluted to respectively to 0.5 ng/mL, 1.0 ng/mL, 2.0 ng/mL, 5.0 ng/mL, 10 ng/mL, 25 ng/mL, 50 ng/mL, 75 ng/mL, 100 ng/mL, 150 ng/mL, 200 ng/mL, 250 ng/mL and 300ng/mL, the AFP standard items that nano combined magnetic bead are placed in successively to above-mentioned variable concentrations are hatched, and incubation time is 20 min;
(4) detect: electromagnetic control electrode is switched on, and the electrode front end is placed in and hatches solution, after absorption, electromagnetic control electrode is moved to and detects in solution, be the solution of pH 6.5 PBS, adopt MEC-12B multifunctional microcomputer electrochemical analyser to carry out the cyclic voltammetry detection, the amount of its peak current and adsorption antigen is inversely proportional to, wherein, peak point current is designated as I
2, the current value I=I descended with oxidation peak current before and after immune response
2-I
1as quantitative basis, to each concentration standard antigen duplicate detection three times, using its mean value current value corresponding as this antigen concentration, and the current value corresponding with it carries out linear regression analysis to antigen concentration.
(5) rinse:, after detecting, outage disappears the magnetic of electromagnetic control electrode, and the PBS flushing is carried out in the surface of electromagnetic control electrode, and nano combined magnetic bead can break away from from the electromagnetic control electrode surface under the PBS washing action.Afterwards, can proceed the detection of sample next time.
Its testing result is shown in Fig. 4, and result shows, AFP standard items concentration is in 1.0 ng/mL ~ 300 ng/mL scopes, and magnetic control electrochemical immunosensor oxidation peak current drop-out current value and antigen concentration are good linear relationship.
At present, the results showed, this technology can be effectively applied to the detection of target molecule, key technical problem of the present invention be by magnetic particle functional modification and magnetic control electrode realized electrochemical immunosensor convenient, effectively prepare and regenerate, realized reusing of electrochemical immunosensor.
Finally explanation is, above embodiment is only unrestricted in order to technical scheme of the present invention to be described, although with reference to preferred embodiment, the present invention is had been described in detail, those of ordinary skill in the art is to be understood that, can modify or be equal to replacement technical scheme of the present invention, and not breaking away from aim and the scope of the technical program, it all should be encompassed in the middle of claim scope of the present invention.
Claims (10)
1. electromagnetic control electrode, it is characterized in that: described electromagnetic control electrode is the multilayer hollow tubular structures, comprises respectively from inside to outside PVDF layer I, copper foil layer, insulation course, PVDF layer II and external cavity layer, described external cavity layer adopts the PVDF material to make;
One end end face of described electromagnetic control electrode is as electrode face, and the other end seals, and described electrode face is piece structure, and described electrode face utilizes wire to draw, and with the external power source binding post, is connected.
2. electromagnetic control electrode as claimed in claim 1, it is characterized in that: the wall thickness of the external cavity layer of described electromagnetic control electrode is 1 ~ 2 mm, external diameter is 8 ~ 10 mm, is highly 80 ~ 110mm.
3. electromagnetic control electrode according to claim 1, it is characterized in that: the external diameter of described piece structure is 3 ~ 4 mm, thickness is 0.5 ~ 2mm.
4. electromagnetic control electrode according to claim 1, it is characterized in that: total wall thickness of the sandwich construction of described electromagnetic control electrode is less than 2.15 mm.
5. the preparation method of electromagnetic control electrode according to claim 1 is characterized in that: comprise the following steps:
(1) adopt the PVDF material to prepare the external cavity layer of electrode;
(2) an end end face of external cavity layer adopts golden material to be prepared with electrode face, adopts wire to draw, and it is connected with the external power source binding post;
(3) interior laminate layer of cavity layer is assembled each layer structure outside, is followed successively by from inside to outside PVDF layer I, copper foil layer, insulation course and PVDF layer II, and gross thickness is less than 2.15 mm.
6. the preparation method of electromagnetic control electrode according to claim 5, it is characterized in that: the wall thickness of described electromagnetic control electrode is 1.5 ~ 2 mm, external diameter is 8 ~ 10mm, is highly 80 ~ 110 mm.
7. according to the preparation method of claim 5 or 6 described electromagnetic control electrodes, it is characterized in that: the external diameter of described electrode face is 3 ~ 4mm, and thickness is 0.5 ~ 0.6mm.
8. utilize described electromagnetic control electrode as arbitrary as claims 1 to 3 to carry out the method for biology sample detection, it is characterized in that: comprise the following steps:
(1) nano combined magnetic bead preparation: Prussian blue, nm of gold are modified successively in the magnetic particle sub-surface, and then fixing alpha-fetoprotein specific antibody, form nano combined magnetic bead;
(2) cyclic voltammetric detects: by the electromagnetic control electrode energising, under the attraction of externally-applied magnetic field, in conjunction with nano combined magnetic bead be absorbed and fixed at the magnetic control electrode surface, electrode is placed in to pH 6.5 PBS solution and carries out the cyclic voltammetric detection, obtain electric current I
1;
(3) hatch: the AFP standard items are diluted to respectively to 0.5 ng/mL, 1.0 ng/mL, 2.0 ng/mL, 5.0 ng/mL, 10 ng/mL, 25 ng/mL, 50 ng/mL, 75 ng/mL, 100 ng/mL, 150 ng/mL, 200 ng/mL, 250 ng/mL and 300 ng/mL, the AFP standard items that nano combined magnetic bead are placed in successively to above-mentioned variable concentrations are hatched, and incubation time is 15 ~ 30 min;
(4) detect: by the electromagnetic control electrode energising, and the electrode front end is placed in and hatches solution, after absorption, electromagnetic control electrode is moved to and detects in solution, and the solution of pH 6.5 PBS, carry out the cyclic voltammetry detection, the amount of its peak current and adsorption antigen is inversely proportional to, and wherein, peak point current is designated as I
2, the current value I=I descended with oxidation peak current before and after immune response
2-I
1as quantitative basis, to each concentration standard antigen duplicate detection three times, using its mean value current value corresponding as this antigen concentration, and the current value corresponding with it carries out linear regression analysis to antigen concentration.
9. biology sample detection method according to claim 8, it is characterized in that: also comprise step (5), after detecting, outage disappears the magnetic of electromagnetic control electrode, the PBS flushing is carried out on surface to electromagnetic control electrode, and nano combined magnetic bead can break away from from the electromagnetic control electrode surface under the PBS washing action.
10. biology sample detection method according to claim 8, it is characterized in that: the incubation time in described step (3) is 20 min.
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN1262440A (en) * | 2000-01-12 | 2000-08-09 | 上海本草生物医学工程研究所 | Semi-quantitative quick-diagnosing reagent for detecting alpha-fetoglobulin and its preparing process |
| CN1392194A (en) * | 2002-07-02 | 2003-01-22 | 武汉理工大学 | Poly meta-fluoroethylene/lead zirconate titanate/terbium dysprosium iron composite material sheet with magnetoelectric property and its preparation |
| CN101776636A (en) * | 2010-01-22 | 2010-07-14 | 福州大学 | Magnetic working electrode and preparation method thereof |
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| US8993346B2 (en) * | 2009-08-07 | 2015-03-31 | Nanomix, Inc. | Magnetic carbon nanotube based biodetection |
| CN202166651U (en) * | 2011-05-06 | 2012-03-14 | 中国人民解放军第三军医大学 | Electromagnetic control immunosensor |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN1262440A (en) * | 2000-01-12 | 2000-08-09 | 上海本草生物医学工程研究所 | Semi-quantitative quick-diagnosing reagent for detecting alpha-fetoglobulin and its preparing process |
| CN1392194A (en) * | 2002-07-02 | 2003-01-22 | 武汉理工大学 | Poly meta-fluoroethylene/lead zirconate titanate/terbium dysprosium iron composite material sheet with magnetoelectric property and its preparation |
| CN101776636A (en) * | 2010-01-22 | 2010-07-14 | 福州大学 | Magnetic working electrode and preparation method thereof |
Non-Patent Citations (2)
| Title |
|---|
| "纳米微粒固载抗体蛋白构建高灵敏电流型免疫传感器的研究";洪成林;《中国博士学位论文全文数据库 信息科技辑》;20100115;第37-46页 * |
| 洪成林."纳米微粒固载抗体蛋白构建高灵敏电流型免疫传感器的研究".《中国博士学位论文全文数据库 信息科技辑》.2010,第37-46页. |
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Inventor after: Li Yan Inventor after: Zheng Junsong Inventor after: Fang Lichao Inventor after: He Juan Inventor after: Deng Jun Inventor after: Huang Hui Inventor after: Jiang Lili Inventor before: Li Yan Inventor before: Zheng Junsong Inventor before: Fang Lichao Inventor before: He Juan Inventor before: Deng Jun Inventor before: Huang Hui Inventor before: Jiang Lili |
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Free format text: CORRECT: INVENTOR; FROM: YAN GUOHUA QIN TAIGUI YAN GUOHUA ZHANG GUANGYU YING GUOZHEN ZHANG YONGZHONG ZHANG YONGZHONG TO: YAN GUOHUA SHI WEIKE YAN GUOHUA ZHANG GUANGYU YING GUOZHEN ZHANG YONGZHONG ZHANG YONGZHONG |
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