CN105717180B - A kind of preparation method and application of the optical electro-chemistry aflatoxin biology sensor based on two-dimensional nano composite - Google Patents
A kind of preparation method and application of the optical electro-chemistry aflatoxin biology sensor based on two-dimensional nano composite Download PDFInfo
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- 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
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- 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
- G01N33/543—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
- G01N33/54366—Apparatus specially adapted for solid-phase testing
- G01N33/54373—Apparatus specially adapted for solid-phase testing involving physiochemical end-point determination, e.g. wave-guides, FETS, gratings
- G01N33/5438—Electrodes
Abstract
The invention discloses a kind of preparation method of optical electro-chemistry aflatoxin biology sensor.Belong to Nano-function thin films and biosensor technology field.The method comprises the steps of firstly, preparing a kind of New Two Dimensional nano composite material Mn TiO2/g‑C3N4, good biocompatibility and big specific surface area using the material, aflatoxin antibody in load, then alkaline phosphatase is fixed by the crosslinked action of glutaraldehyde, when being detected, because alkaline phosphatase can be catalyzed, the tricresyl phosphate sodium salt AAP of L ascorbic acid 2 is in situ to produce L ascorbic acid AA, and and then provide electron donor for Photoelectric Detection, antibody is recycled to be combined the influence to electron transport ability with the specific quantification of antigen, so that photo-current intensity accordingly reduces, it is low that cost finally has been made, high sensitivity, specificity is good, detection is quick, prepare the Photoelectrochemistrbiosensor biosensor of simple detection aflatoxin.
Description
Technical field
The present invention relates to a kind of preparation method of optical electro-chemistry aflatoxin biology sensor.Belong to novel nanometer functional
Material and biosensor technology field.
Background technology
Aflatoxin is the similar compound of a kind of chemical constitution, is the derivative of dihydrofuran cumarin.Huang Qu
Mould toxin is present in soil, animals and plants, various nuts, particularly easily pollutes the grains such as peanut, corn, rice, soybean, wheat
Oil product, it is a kind of mycotoxin that mycotoxicosis is maximum, extremely protrudes human health risk.Aflatoxin master
There are Aflatoxins M1, aflatoxin B1, aflatoxin B 2, aflatoxin G 1, aflatoxin G 2 etc. several.It is yellow
It is 1 class carcinogenic substance that aspertoxin delimited by the World Health Organization, and toxicity is bigger than arsenic 68 times, is only second to meat poisoning mycin, is current
Known mycetism is most strong.The harmfulness of aflatoxin is there is destruction to people and animal's liver tissue, seriously
When liver cancer can be caused even dead, Homemade fermented food can also detect aflatoxin, especially hot and humid regional
Grain and oil and product kind recall rate are higher.
At present, the method for detecting aflatoxin mainly has chromatography, mass spectrography etc..Such method instrument is valuable, operation
Complexity, laboratory personnel could be detected after needing professional training.Therefore, R＆D costs are low, detection is fast, high sensitivity, special
The strong aflatoxin sensor of property is significant.
Optical electro-chemistry sensor due to high sensitivity, testing cost is low the features such as, in recent years by increasing researcher
It is of interest.Optical electro-chemistry sensor is to cause electron-hole pair to be separated based on additional light source activation Electrophotosensitivmaterial material,
Under suitable potential condition partially, quick transmission of the electronics on electrode, semiconductor and trim and analyte is realized, and form light
Electric current.In optimal conditions, the change of analyte concentration can directly affect the size of photoelectric current, recycle biological immune to combine,
Can realizes the qualitative and quantitative analysis to analyte according to the change of photoelectric current.
Optical electro-chemistry sensor most critical technology is exactly the raising to performances such as the size of photoelectric current and stability.Titanium dioxide
Titanium is a kind of photochemical catalyst and the light induced electron host material being most widely used, because sheet-like titanium dioxide nanomaterial can
The more high miller index surfaces of exposure, have higher photocatalytic activity, and titanium dioxide nanoplate has than nano-particle preferably
Application prospect, the research for titanium dioxide nanoplate also receive much concern.And the photoproduction electricity of single titanium dioxide nano material
Son-hole is to easily compound, and so as to cause the decrease of photosignal, and titanium dioxide poorly conductive also limit by single dioxy
It is universal not high to change the sensitivity of the optical electro-chemistry sensor of titanium nano material structure, is unfavorable for practical application.But in semiconductor
Modification or compound special nano material in nano material, the valid density of photo-generated carrier pair can be effectively improved, improves light
Photoelectric transformation efficiency, and greatly improve detection sensitivity.Therefore, design, prepare titanium dioxide nanoplate efficiently, stable and its repair
Jewelry is the key technology for preparing optical electro-chemistry sensor.
The content of the invention
Prepare that simple, high sensitivity, detection be quick, optical electro-chemistry of high specificity it is an object of the invention to provide a kind of
The preparation method of aflatoxin biology sensor, prepared sensor, quick, the sensitive inspection available for aflatoxin
Survey.Based on this purpose, the method comprises the steps of firstly, preparing a kind of New Two Dimensional nano composite material, i.e. additive Mn titanium dioxide nanoplate
In-situ reaction carbonitride two-dimensional nano composite Mn-TiO2/g-C3N4, utilize the good biocompatibility of the material and big
Specific surface area, aflatoxin antibody in load, alkaline phosphatase is then fixed by the crosslinked action of glutaraldehyde, carried out
During detection, because alkaline phosphatase can be catalyzed, L-AA -2- tricresyl phosphate sodium salts AAP is in situ to produce L-AA AA,
And and then provide electron donor for Photoelectric Detection, recycle antibody to be combined with the specific quantification of antigen to electron transport ability
Influence so that photo-current intensity accordingly reduces, and finally realizes using unmarked PhotoelectrochemicalMethod Method detection aflatoxin
Biology sensor structure.
The technical solution adopted by the present invention is as follows：
1. a kind of preparation method of the optical electro-chemistry aflatoxin biology sensor based on two-dimensional nano composite,
Described two-dimensional nano composite is additive Mn titanium dioxide nanoplate In-situ reaction carbonitride two-dimensional nano composite Mn-
TiO2/g-C3N4, described optical electro-chemistry aflatoxin biology sensor is by working electrode, Mn-TiO2/g-C3N4, aspergillus flavus
Toxin antibody, bovine serum albumin(BSA), glutaraldehyde, alkaline phosphatase composition；
Characterized in that, described preparation method includes following preparation process：
a. Mn-TiO2/g-C3N4Preparation；
B. the preparation of optical electro-chemistry aflatoxin biology sensor；
Wherein, step a prepares Mn-TiO2/g-C3N4Concretely comprise the following steps：
First, 0.8 ~ 1.2 mmol manganese salts are taken to be added in 5 mL butyl titanates, in whipping process, it is slowly added to 0.5 ~
0.8 mL hydrofluoric acid, react in a kettle 18 ~ 24 hours, after being cooled to room temperature at 160 ~ 200 DEG C, with ultra-pure water and anhydrous
Ethanol centrifuge washing three times after, be dried in vacuo at 50 DEG C；Secondly, the dried solids of 150 ~ 250 mg and 400mg melamines are taken
Amine mixes, and grind into powder；Then, the powder of grinding is put into Muffle furnace, programming rate is 1 ~ 3 DEG C/min, 480
Calcined 0.5 ~ 5 hour at ~ 560 DEG C；Finally, the powder after calcining is cooled to room temperature, that is, Mn-TiO is made2/g-C3N4；
Described manganese salt is selected from one of following：Manganese sulfate, manganese chloride, manganese nitrate；
Step b prepares concretely comprising the following steps for optical electro-chemistry aflatoxin biology sensor：
（1）Using ITO electro-conductive glass as working electrode, in the uL of electrode surface drop coating 8 ~ 12 Mn-TiO2/g-C3N4Colloidal sol,
Dry at room temperature；
（2）By step（1）In obtained electrode cushioning liquid PBS, continue in the μ L of electrode surface drop coating 8 ~ 12
10 μ g/mL aflatoxin antibody-solutions, preserve in 4 DEG C of refrigerators and dry；
（3）By step（2）In obtained electrode PBS, it is 100 to continue in the μ L concentration of electrode surface drop coating 8 ~ 12
μ g/mL bovine serum albumin solution, preserve in 4 DEG C of refrigerators and dry；
（4）By step（3）In obtained electrode PBS, the glutaraldehyde continued in the μ L of electrode surface drop coating 2 ~ 4 is molten
Liquid, preserve in 4 DEG C of refrigerators and dry；
（5）By step（4）In obtained electrode PBS, it is 20 μ to continue in the μ L concentration of electrode surface drop coating 6 ~ 10
G/mL alkaline phosphatase enzyme solutions, preserve in 4 DEG C of refrigerators and dry；
（6）By step（5）In obtained electrode PBS, preserved in 4 DEG C of refrigerators after drying, that is, photoelectricity be made
Chemical aflatoxin biology sensor；
Described Mn-TiO2/g-C3N4Colloidal sol is by 50 mg Mn-TiO2/g-C3N4Powder is dissolved in 10 mL ultra-pure waters
In, and the obtained hydrosol after 30 min of ultrasound；
Described PBS is 10mmol/L phosphate buffer solution, and the pH value of described phosphate buffer solution is 7.4；
Described glutaraldehyde solution is the glutaraldehyde water solution that volume ratio is 2.5%.
2. the system of the optical electro-chemistry aflatoxin biology sensor of the present invention based on two-dimensional nano composite
Preparation Method, it is characterised in that the aflatoxin is selected from one of following：Aflatoxins M1, aflatoxin B1, aspergillus flavus
Toxin B2, aflatoxin G 1, aflatoxin G 2.
3. the application of the optical electro-chemistry aflatoxin biology sensor prepared by preparation method of the present invention, its
It is characterised by, including following applying step：
A. standard liquid is prepared：The aflatoxin standard for preparing one group of various concentrations including blank standard specimen is molten
Liquid；
B. working electrode is modified：Optical electro-chemistry aflatoxin prepared by preparation method as described in the present invention is given birth to
Thing sensor is working electrode, and the aflatoxin standard liquid for the various concentrations prepared in step b is distinguished into drop coating to work
Electrode surface, preserve in 4 DEG C of refrigerators；
C. working curve is drawn：Using saturated calomel electrode as reference electrode, platinum electrode is as auxiliary electrode, with step
The working electrode composition three-electrode system that rapid b has been modified, is connected on optical electro-chemistry detection device；Successively add in a cell
Enter 15mL pH=9.6 Tris-HCl cushioning liquid and the mmol/L of 5 mL 10 L-AA -2- tricresyl phosphate sodium salts AAP it is molten
Liquid；Using i-t means of testing, according to the relation between the photocurrent values of gained and aflatoxin concentration of standard solution, draw
Working curve；
D. the detection of aflatoxin：The aflatoxin standard liquid in step a is replaced with testing sample, according to step
Method in rapid b and c is detected, and according to the photocurrent values and working curve of gained, obtains aflatoxin in testing sample
Content.
The useful achievement of the present invention
（1）Optical electro-chemistry aflatoxin biology sensor of the present invention prepare it is simple, it is easy to operate, realize pair
The quick, sensitive of sample, high selectivity detection, and cost is low, can be applied to portable inspectiont, has market development prospect；
（2）The present invention is prepared for new light-sensitive material Mn-TiO first2/g-C3N4, because manganese is on titanium dioxide nanoplate
Growth in situ and fully contacted with titanium dioxide nanoplate, using manganese metal surface plasma body act on, effectively prevent
Photo-generate electron-hole pair it is compound, although it is good to solve titanium dioxide nanoplate photocatalysis effect, photoelectric current produces smaller
Technical problem；Simultaneously because carbonitride g-C3N4Good electric conductivity and titanium dioxide nanoplate it is fully dispersed thereon,
Greatly increase the photocatalytic activity of titanium dioxide nanoplate and solve titanium dioxide nanoplate poorly conductive and be unfavorable for
The technical problem of optical electro-chemistry sensor is prepared, therefore, effective preparation of the material, there is important scientific meaning and using valency
Value；
（3）The present invention is first by Mn-TiO2/g-C3N4Applied in the preparation of Photoelectrochemistrbiosensor biosensor, significantly improve
The valid density of photo-generated carrier, substantially increase the detection sensitivity of optical electro-chemistry sensor so that optical electro-chemistry biology
Sensor realizes the application in real work；The application of the material, also it is associated biomolecule sensor, such as electrogenerated chemiluminescence
Sensor, electrochemical sensor etc. provide Technical Reference, have extensive potential use value.
Embodiment
The Mn-TiO of embodiment 12/g-C3N4Preparation
First, take 0.8 mmol manganese salts to be added in 5 mL butyl titanates, in whipping process, be slowly added to 0.5 mL hydrogen
Fluoric acid, react in a kettle 24 hours, after being cooled to room temperature at 160 DEG C, with ultra-pure water and absolute ethyl alcohol centrifuge washing three times
Afterwards, it is dried in vacuo at 50 DEG C；Secondly, the dried solids of 150 mg are taken to be mixed with 400 mg melamines, and pulverize
End；Then, the powder of grinding is put into Muffle furnace, programming rate is 1 DEG C/min, is calcined 5 hours at 480 DEG C；Finally,
Powder after calcining is cooled to room temperature, that is, Mn-TiO is made2/g-C3N4；
Described manganese salt is manganese sulfate.
The Mn-TiO of embodiment 22/g-C3N4Preparation
First, take 1.0 mmol manganese salts to be added in 5 mL butyl titanates, in whipping process, be slowly added to 0.6 mL hydrogen
Fluoric acid, react in a kettle 21 hours, after being cooled to room temperature at 180 DEG C, with ultra-pure water and absolute ethyl alcohol centrifuge washing three times
Afterwards, it is dried in vacuo at 50 DEG C；Secondly, the dried solids of 200 mg are taken to be mixed with 400 mg melamines, and pulverize
End；Then, the powder of grinding is put into Muffle furnace, programming rate is 2 DEG C/min, is calcined 2 hours at 520 DEG C；Finally,
Powder after calcining is cooled to room temperature, that is, Mn-TiO is made2/g-C3N4；
Described manganese salt is manganese chloride.
The Mn-TiO of embodiment 32/g-C3N4Preparation
First, take 1.2 mmol manganese salts to be added in 5 mL butyl titanates, in whipping process, be slowly added to 0.8 mL hydrogen
Fluoric acid, react in a kettle 18 hours, after being cooled to room temperature at 200 DEG C, with ultra-pure water and absolute ethyl alcohol centrifuge washing three
After secondary, it is dried in vacuo at 50 DEG C；Secondly, the dried solids of 250 mg are taken to be mixed with 400 mg melamines, and pulverize
End；Then, the powder of grinding is put into Muffle furnace, programming rate is 3 DEG C/min, is calcined 0.5 hour at 560 DEG C；Most
Afterwards, the powder after calcining is cooled to room temperature, that is, Mn-TiO is made2/g-C3N4；
Described manganese salt is manganese nitrate.
The preparation method of the optical electro-chemistry aflatoxin biology sensor of embodiment 4
（1）Using a width of 1 cm, a length of 4 cm ITO electro-conductive glass as working electrode, the μ L's of electrode surface drop coating 8
Mn-TiO2/g-C3N4Colloidal sol, dry at room temperature；
（2）By step（1）In obtained electrode cushioning liquid PBS, continue in the μ of 8 μ L of electrode surface drop coating 10
G/mL aflatoxin antibody-solutions, preserve in 4 DEG C of refrigerators and dry；
（3）By step（2）In obtained electrode PBS, it is 100 μ to continue in the μ L concentration of electrode surface drop coating 8
G/mL bovine serum albumin solution, preserve in 4 DEG C of refrigerators and dry；
（4）By step（3）In obtained electrode PBS, the glutaraldehyde continued in the μ L of electrode surface drop coating 2 is molten
Liquid, preserve in 4 DEG C of refrigerators and dry；
（5）By step（4）In obtained electrode PBS, it is 20 μ g/ to continue in the μ L concentration of electrode surface drop coating 6
ML alkaline phosphatase enzyme solutions, preserve in 4 DEG C of refrigerators and dry；
（6）By step（5）In obtained electrode PBS, preserved in 4 DEG C of refrigerators after drying, that is, photoelectricity be made
Chemical aflatoxin biology sensor；
Described Mn-TiO2/g-C3N4Colloidal sol is by 50 mg Mn-TiO2/g-C3N4Powder is dissolved in 10 mL ultra-pure waters
In, and the obtained hydrosol after 30 min of ultrasound；
Described PBS is 10mmol/L phosphate buffer solution, and the pH value of described phosphate buffer solution is 7.4；
Described glutaraldehyde solution is the glutaraldehyde water solution that volume ratio is 2.5%.
The preparation method of the optical electro-chemistry aflatoxin biology sensor of embodiment 5
（1）Using a width of 1 cm, a length of 4 cm ITO electro-conductive glass as working electrode, the μ L's of electrode surface drop coating 10
Mn-TiO2/g-C3N4Colloidal sol, dry at room temperature；
（2）By step（1）In obtained electrode cushioning liquid PBS, continue in the μ L 10 of electrode surface drop coating 10
μ g/mL aflatoxin antibody-solutions, preserve in 4 DEG C of refrigerators and dry；
（3）By step（2）In obtained electrode PBS, it is 100 μ to continue in the μ L concentration of electrode surface drop coating 10
G/mL bovine serum albumin solution, preserve in 4 DEG C of refrigerators and dry；
（4）By step（3）In obtained electrode PBS, the glutaraldehyde continued in the μ L of electrode surface drop coating 3 is molten
Liquid, preserve in 4 DEG C of refrigerators and dry；
（5）By step（4）In obtained electrode PBS, it is 20 μ g/ to continue in the μ L concentration of electrode surface drop coating 8
ML alkaline phosphatase enzyme solutions, preserve in 4 DEG C of refrigerators and dry；
（6）By step（5）In obtained electrode PBS, preserved in 4 DEG C of refrigerators after drying, that is, photoelectricity be made
Chemical aflatoxin biology sensor；
Described Mn-TiO2/g-C3N4Colloidal sol is by 50 mg Mn-TiO2/g-C3N4Powder is dissolved in 10 mL ultra-pure waters
In, and the obtained hydrosol after 30 min of ultrasound；
Described PBS is 10 mmol/L phosphate buffer solution, and the pH value of described phosphate buffer solution is 7.4；
Described glutaraldehyde solution is the glutaraldehyde water solution that volume ratio is 2.5%.
The preparation method of the optical electro-chemistry aflatoxin biology sensor of embodiment 6
（1）Using a width of 1 cm, a length of 4 cm ITO electro-conductive glass as working electrode, the μ L's of electrode surface drop coating 12
Mn-TiO2/g-C3N4Colloidal sol, dry at room temperature；
（2）By step（1）In obtained electrode cushioning liquid PBS, continue in the μ L 10 of electrode surface drop coating 12
μ g/mL aflatoxin antibody-solutions, preserve in 4 DEG C of refrigerators and dry；
（3）By step（2）In obtained electrode PBS, it is 100 μ to continue in the μ L concentration of electrode surface drop coating 12
G/mL bovine serum albumin solution, preserve in 4 DEG C of refrigerators and dry；
（4）By step（3）In obtained electrode PBS, the glutaraldehyde continued in the μ L of electrode surface drop coating 4 is molten
Liquid, preserve in 4 DEG C of refrigerators and dry；
（5）By step（4）In obtained electrode PBS, it is 20 μ g/ to continue in the μ L concentration of electrode surface drop coating 10
ML alkaline phosphatase enzyme solutions, preserve in 4 DEG C of refrigerators and dry；
（6）By step（5）In obtained electrode PBS, preserved in 4 DEG C of refrigerators after drying, that is, photoelectricity be made
Chemical aflatoxin biology sensor；
Described Mn-TiO2/g-C3N4Colloidal sol is by 50 mg Mn-TiO2/g-C3N4Powder is dissolved in 10 mL ultra-pure waters
In, and the obtained hydrosol after 30 min of ultrasound；
Described PBS is 10 mmol/L phosphate buffer solution, and the pH value of described phosphate buffer solution is 7.4；
Described glutaraldehyde solution is the glutaraldehyde water solution that volume ratio is 2.5%.
Optical electro-chemistry aflatoxin biology sensor prepared by the embodiment 1 and 3 of embodiment 7, applied to aspergillus flavus poison
The detection of element, step are as follows：
（1）Standard liquid is prepared：The aflatoxin standard for preparing one group of various concentrations including blank standard specimen is molten
Liquid；
（2）Working electrode is modified：Optical electro-chemistry aflatoxin prepared by preparation method as described in the present invention is given birth to
Thing sensor is working electrode, by step（1）The aflatoxin standard liquid of the various concentrations of middle preparation distinguishes drop coating to work
Make electrode surface, preserved in 4 DEG C of refrigerators；
（3）Working curve is drawn：Using saturated calomel electrode as reference electrode, platinum electrode is as auxiliary electrode, with step
Suddenly（2）The working electrode composition three-electrode system modified, is connected on optical electro-chemistry detection device；In a cell successively
Add the Tris-HCl cushioning liquid of 15mL pH=9.6 and the mmol/L of 5 mL 10 L-AA -2- tricresyl phosphate sodium salts AAP
Solution；Using i-t means of testing, according to the relation between the photocurrent values of gained and aflatoxin concentration of standard solution, paint
Working curve processed；The linear detection range of aflatoxin is：0.002 ~ 200 ng/mL, detection are limited to：0.8 pg/mL；
（4）Actual sample detects：Step is replaced with testing sample（1）In aflatoxin standard liquid, according to step
（2）With（3）In method detected, according to the photocurrent values and working curve of gained, obtain aspergillus flavus poison in testing sample
The content of element；
Described aflatoxin is Aflatoxins M1.
Optical electro-chemistry aflatoxin biology sensor prepared by the embodiment 2 and 4 of embodiment 8, applied to aspergillus flavus poison
The detection of element, step are as follows：
（1）Standard liquid is prepared：The aflatoxin standard for preparing one group of various concentrations including blank standard specimen is molten
Liquid；
（2）Working electrode is modified：Optical electro-chemistry aflatoxin prepared by preparation method as described in the present invention is given birth to
Thing sensor is working electrode, by step（1）The aflatoxin standard liquid of the various concentrations of middle preparation distinguishes drop coating to work
Make electrode surface, preserved in 4 DEG C of refrigerators；
（3）Working curve is drawn：Using saturated calomel electrode as reference electrode, platinum electrode is as auxiliary electrode, with step
Suddenly（2）The working electrode composition three-electrode system modified, is connected on optical electro-chemistry detection device；In a cell successively
Add the Tris-HCl cushioning liquid of 15mL pH=9.6 and the mmol/L of 5 mL 10 L-AA -2- tricresyl phosphate sodium salts AAP
Solution；Using i-t means of testing, according to the relation between the photocurrent values of gained and aflatoxin concentration of standard solution, paint
Working curve processed；The linear detection range of aflatoxin is：0.002 ~ 200 ng/mL, detection are limited to：0.8 pg/mL；
（4）Actual sample detects：Step is replaced with testing sample（1）In aflatoxin standard liquid, according to step
（2）With（3）In method detected, according to the photocurrent values and working curve of gained, obtain aspergillus flavus poison in testing sample
The content of element；
Described aflatoxin is aflatoxin B1.
Optical electro-chemistry aflatoxin biology sensor prepared by the embodiment 3 and 6 of embodiment 9, applied to aspergillus flavus poison
The detection of element, step are as follows：
（1）Standard liquid is prepared：The aflatoxin standard for preparing one group of various concentrations including blank standard specimen is molten
Liquid；
（2）Working electrode is modified：Optical electro-chemistry aflatoxin prepared by preparation method as described in the present invention is given birth to
Thing sensor is working electrode, by step（1）The aflatoxin standard liquid of the various concentrations of middle preparation distinguishes drop coating to work
Make electrode surface, preserved in 4 DEG C of refrigerators；
（3）Working curve is drawn：Using saturated calomel electrode as reference electrode, platinum electrode is as auxiliary electrode, with step
Suddenly（2）The working electrode composition three-electrode system modified, is connected on optical electro-chemistry detection device；In a cell successively
Add the Tris-HCl cushioning liquid of 15mL pH=9.6 and the mmol/L of 5 mL 10 L-AA -2- tricresyl phosphate sodium salts AAP
Solution；Using i-t means of testing, according to the relation between the photocurrent values of gained and aflatoxin concentration of standard solution, paint
Working curve processed；The linear detection range of aflatoxin is：0.002 ~ 200 ng/mL, detection are limited to：0.8 pg/mL；
（4）Actual sample detects：Step is replaced with testing sample（1）In aflatoxin standard liquid, according to step
（2）With（3）In method detected, according to the photocurrent values and working curve of gained, obtain aspergillus flavus poison in testing sample
The content of element；
Described aflatoxin is aflatoxin G 1.
Claims (3)
1. a kind of preparation method of the optical electro-chemistry aflatoxin biology sensor based on two-dimensional nano composite, described
Two-dimensional nano composite is additive Mn titanium dioxide nanoplate In-situ reaction carbonitride two-dimensional nano composite Mn-TiO2/
g-C3N4, described optical electro-chemistry aflatoxin biology sensor is by working electrode, Mn-TiO2/g-C3N4, aflatoxin resists
Body, bovine serum albumin(BSA), glutaraldehyde, alkaline phosphatase composition；
Characterized in that, described preparation method includes following preparation process：
a. Mn-TiO2/g-C3N4Preparation；
B. the preparation of optical electro-chemistry aflatoxin biology sensor；
Wherein, step a prepares Mn-TiO2/g-C3N4Concretely comprise the following steps：
First, take 0.8 ~ 1.2 mmol manganese salts to be added in 5 mL butyl titanates, in whipping process, be slowly added to 0.5 ~ 0.8
ML hydrofluoric acid, react in a kettle 18 ~ 24 hours, after being cooled to room temperature at 160 ~ 200 DEG C, with ultra-pure water and absolute ethyl alcohol
Centrifuge washing three times after, be dried in vacuo at 50 DEG C；Secondly, the dried solids of 150 ~ 250 mg are taken to be mixed with 400mg melamines
Close, and grind into powder；Then, the powder of grinding is put into Muffle furnace, programming rate is 1 ~ 3 DEG C/min, 480 ~ 560
Calcined 0.5 ~ 5 hour at DEG C；Finally, the powder after calcining is cooled to room temperature, that is, Mn-TiO is made2/g-C3N4；
Described manganese salt is selected from one of following：Manganese sulfate, manganese chloride, manganese nitrate；
Step b prepares concretely comprising the following steps for optical electro-chemistry aflatoxin biology sensor：
（1）Using ITO electro-conductive glass as working electrode, in the μ L of electrode surface drop coating 8 ~ 12 Mn-TiO2/g-C3N4Colloidal sol, room temperature
Under dry；
（2）By step（1）In obtained electrode cushioning liquid PBS, continue in the μ of 8 ~ 12 μ L of electrode surface drop coating 10
G/mL aflatoxin antibody-solutions, preserve in 4 DEG C of refrigerators and dry；
（3）By step（2）In obtained electrode PBS, it is 100 μ g/ to continue in the μ L concentration of electrode surface drop coating 8 ~ 12
ML bovine serum albumin solution, preserve in 4 DEG C of refrigerators and dry；
（4）By step（3）In obtained electrode PBS, continue the glutaraldehyde solution in the μ L of electrode surface drop coating 2 ~ 4,4
Preserve and dry in DEG C refrigerator；
（5）By step（4）In obtained electrode PBS, it is 20 μ g/mL to continue in the μ L concentration of electrode surface drop coating 6 ~ 10
Alkaline phosphatase enzyme solutions, preserve in 4 DEG C of refrigerators and dry；
（6）By step（5）In obtained electrode PBS, preserved in 4 DEG C of refrigerators after drying, that is, optical electro-chemistry be made
Aflatoxin biology sensor；
Described Mn-TiO2/g-C3N4Colloidal sol is by 50 mg Mn-TiO2/g-C3N4Powder is dissolved in 10 mL ultra-pure waters, and
The obtained hydrosol after 30 min of ultrasound；
Described PBS is 10mmol/L phosphate buffer solution, and the pH value of described phosphate buffer solution is 7.4；
Described glutaraldehyde solution is the glutaraldehyde water solution that volume ratio is 2.5%.
2. the system of the optical electro-chemistry aflatoxin biology sensor based on two-dimensional nano composite as claimed in claim 1
Preparation Method, it is characterised in that the aflatoxin is selected from one of following：Aflatoxins M1, aflatoxin B1, aspergillus flavus
Toxin B2, aflatoxin G 1, aflatoxin G 2.
3. the application of the optical electro-chemistry aflatoxin biology sensor prepared by preparation method as claimed in claim 1, its
It is characterised by, including following applying step：
A. standard liquid is prepared：Prepare the aflatoxin standard liquid of one group of various concentrations including blank standard specimen；
B. working electrode is modified：By the optical electro-chemistry aflatoxin biology prepared by preparation method as claimed in claim 1
Sensor is working electrode, and the aflatoxin standard liquid for the various concentrations prepared in step a is distinguished into drop coating to work electricity
Pole surface, preserve in 4 DEG C of refrigerators；
C. working curve is drawn：Using saturated calomel electrode as reference electrode, platinum electrode is as auxiliary electrode, with step b institutes
The working electrode composition three-electrode system modified, is connected on optical electro-chemistry detection device；Successively add in a cell
Tris-HCl the cushioning liquid of 15mL pH=9.6 and the mmol/L of 5 mL 10 L-AA -2- tricresyl phosphate sodium salts AAP are molten
Liquid；Using i-t means of testing, according to the relation between the photocurrent values of gained and aflatoxin concentration of standard solution, draw
Working curve；
D. the detection of aflatoxin：The aflatoxin standard liquid in step a is replaced with testing sample, according to step b and
Method in c is detected, and according to the photocurrent values and working curve of gained, obtains containing for aflatoxin in testing sample
Amount.
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| CN108318553B (en) * | 2018-02-06 | 2019-11-12 | 红河学院 | AFB1Electrochemical immunosensor and preparation method thereof and its be used for aflatoxins B1Detection |
| CN110411951B (en) * | 2019-08-13 | 2021-11-30 | 信阳师范学院 | Preparation method of photoelectrochemical biosensor for simultaneously detecting double myocardial markers |
| CN111189895A (en) * | 2020-01-07 | 2020-05-22 | 江苏师范大学 | Method for photoelectrochemical detection of acid phosphatase based on electron transfer inhibition strategy |
| CN111220671A (en) * | 2020-03-30 | 2020-06-02 | 济南大学 | Detect aflatoxin B1Preparation method and application of electrochemiluminescence sensing electrode |
| CN112924662A (en) * | 2021-01-27 | 2021-06-08 | 齐鲁工业大学 | Preparation method of electrochemiluminescence immunosensor and application of electrochemiluminescence immunosensor in detection of aflatoxin B1 |
| CN113189158B (en) * | 2021-04-25 | 2023-03-31 | 武汉科技大学 | Hydrogen sulfide photoelectrochemical sensor and preparation method thereof |
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