CN104880456A - Preparation method and application of electrochemiluminescence immunosensor constructed on basis of GO/MWCNTs-COOH/Au @ CeO2 - Google Patents
Preparation method and application of electrochemiluminescence immunosensor constructed on basis of GO/MWCNTs-COOH/Au @ CeO2 Download PDFInfo
- Publication number
- CN104880456A CN104880456A CN201510262821.7A CN201510262821A CN104880456A CN 104880456 A CN104880456 A CN 104880456A CN 201510262821 A CN201510262821 A CN 201510262821A CN 104880456 A CN104880456 A CN 104880456A
- Authority
- CN
- China
- Prior art keywords
- mwcnts
- cooh
- electrode
- auceo
- preparation
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
- 125000003178 carboxy group Chemical group [H]OC(*)=O 0.000 title claims abstract description 35
- 238000002360 preparation method Methods 0.000 title claims abstract description 20
- 229910000422 cerium(IV) oxide Inorganic materials 0.000 title abstract 3
- 206010028980 Neoplasm Diseases 0.000 claims abstract description 27
- 239000000243 solution Substances 0.000 claims description 23
- 238000001514 detection method Methods 0.000 claims description 17
- 229910021642 ultra pure water Inorganic materials 0.000 claims description 17
- 239000012498 ultrapure water Substances 0.000 claims description 17
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 claims description 11
- 239000000427 antigen Substances 0.000 claims description 11
- 102000036639 antigens Human genes 0.000 claims description 11
- 108091007433 antigens Proteins 0.000 claims description 11
- 239000000843 powder Substances 0.000 claims description 9
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 claims description 8
- 238000013019 agitation Methods 0.000 claims description 8
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 6
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 claims description 6
- 238000004458 analytical method Methods 0.000 claims description 6
- 239000007853 buffer solution Substances 0.000 claims description 6
- 230000000694 effects Effects 0.000 claims description 6
- 229910021389 graphene Inorganic materials 0.000 claims description 6
- BASFCYQUMIYNBI-UHFFFAOYSA-N platinum Chemical compound [Pt] BASFCYQUMIYNBI-UHFFFAOYSA-N 0.000 claims description 6
- 238000001556 precipitation Methods 0.000 claims description 6
- 238000001291 vacuum drying Methods 0.000 claims description 6
- 238000005406 washing Methods 0.000 claims description 6
- 230000000890 antigenic effect Effects 0.000 claims description 5
- 229910052799 carbon Inorganic materials 0.000 claims description 5
- 239000002041 carbon nanotube Substances 0.000 claims description 5
- 229910021393 carbon nanotube Inorganic materials 0.000 claims description 5
- 239000007788 liquid Substances 0.000 claims description 5
- 210000002966 serum Anatomy 0.000 claims description 5
- 206010006187 Breast cancer Diseases 0.000 claims description 4
- 208000026310 Breast neoplasm Diseases 0.000 claims description 4
- 108090000288 Glycoproteins Proteins 0.000 claims description 4
- 102000003886 Glycoproteins Human genes 0.000 claims description 4
- 101000598160 Homo sapiens Nuclear mitotic apparatus protein 1 Proteins 0.000 claims description 4
- 101100043112 Homo sapiens SERPINB3 gene Proteins 0.000 claims description 4
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 claims description 4
- 102000008297 Nuclear Matrix-Associated Proteins Human genes 0.000 claims description 4
- 108010035916 Nuclear Matrix-Associated Proteins Proteins 0.000 claims description 4
- 102100036961 Nuclear mitotic apparatus protein 1 Human genes 0.000 claims description 4
- NBIIXXVUZAFLBC-UHFFFAOYSA-N Phosphoric acid Chemical compound OP(O)(O)=O NBIIXXVUZAFLBC-UHFFFAOYSA-N 0.000 claims description 4
- 108010072866 Prostate-Specific Antigen Proteins 0.000 claims description 4
- 102100038358 Prostate-specific antigen Human genes 0.000 claims description 4
- 102100036383 Serpin B3 Human genes 0.000 claims description 4
- QAOWNCQODCNURD-UHFFFAOYSA-N Sulfuric acid Chemical compound OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 claims description 4
- 210000004027 cell Anatomy 0.000 claims description 4
- 108090000623 proteins and genes Proteins 0.000 claims description 4
- 238000003756 stirring Methods 0.000 claims description 4
- 238000012360 testing method Methods 0.000 claims description 4
- 229910021607 Silver chloride Inorganic materials 0.000 claims description 3
- 238000004401 flow injection analysis Methods 0.000 claims description 3
- 229910052697 platinum Inorganic materials 0.000 claims description 3
- 239000000047 product Substances 0.000 claims description 3
- 229920006395 saturated elastomer Polymers 0.000 claims description 3
- HKZLPVFGJNLROG-UHFFFAOYSA-M silver monochloride Chemical compound [Cl-].[Ag+] HKZLPVFGJNLROG-UHFFFAOYSA-M 0.000 claims description 3
- 238000001228 spectrum Methods 0.000 claims description 3
- GZCWLCBFPRFLKL-UHFFFAOYSA-N 1-prop-2-ynoxypropan-2-ol Chemical compound CC(O)COCC#C GZCWLCBFPRFLKL-UHFFFAOYSA-N 0.000 claims description 2
- 102000013563 Acid Phosphatase Human genes 0.000 claims description 2
- 108010051457 Acid Phosphatase Proteins 0.000 claims description 2
- 102000002260 Alkaline Phosphatase Human genes 0.000 claims description 2
- 108020004774 Alkaline Phosphatase Proteins 0.000 claims description 2
- 102400000739 Corticotropin Human genes 0.000 claims description 2
- 101800000414 Corticotropin Proteins 0.000 claims description 2
- 108010051696 Growth Hormone Proteins 0.000 claims description 2
- MHAJPDPJQMAIIY-UHFFFAOYSA-N Hydrogen peroxide Chemical compound OO MHAJPDPJQMAIIY-UHFFFAOYSA-N 0.000 claims description 2
- 102000011782 Keratins Human genes 0.000 claims description 2
- 108010076876 Keratins Proteins 0.000 claims description 2
- 102100036893 Parathyroid hormone Human genes 0.000 claims description 2
- 108090000445 Parathyroid hormone Proteins 0.000 claims description 2
- 102100024819 Prolactin Human genes 0.000 claims description 2
- 108010057464 Prolactin Proteins 0.000 claims description 2
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 claims description 2
- 102100038803 Somatotropin Human genes 0.000 claims description 2
- 229910000147 aluminium phosphate Inorganic materials 0.000 claims description 2
- 229960004407 chorionic gonadotrophin Drugs 0.000 claims description 2
- 239000000122 growth hormone Substances 0.000 claims description 2
- VNWKTOKETHGBQD-UHFFFAOYSA-N methane Chemical compound C VNWKTOKETHGBQD-UHFFFAOYSA-N 0.000 claims description 2
- 239000004570 mortar (masonry) Substances 0.000 claims description 2
- 230000003169 placental effect Effects 0.000 claims description 2
- 239000012286 potassium permanganate Substances 0.000 claims description 2
- 229940097325 prolactin Drugs 0.000 claims description 2
- 239000013049 sediment Substances 0.000 claims description 2
- 239000007787 solid Substances 0.000 claims description 2
- 239000006228 supernatant Substances 0.000 claims description 2
- 239000003643 water by type Substances 0.000 claims description 2
- 239000000463 material Substances 0.000 abstract description 9
- CETPSERCERDGAM-UHFFFAOYSA-N ceric oxide Chemical compound O=[Ce]=O CETPSERCERDGAM-UHFFFAOYSA-N 0.000 abstract 1
- 238000013399 early diagnosis Methods 0.000 abstract 1
- 238000003018 immunoassay Methods 0.000 abstract 1
- 239000008204 material by function Substances 0.000 abstract 1
- 239000000758 substrate Substances 0.000 abstract 1
- 239000000126 substance Substances 0.000 description 7
- 150000001875 compounds Chemical class 0.000 description 6
- 238000000034 method Methods 0.000 description 5
- 238000004020 luminiscence type Methods 0.000 description 4
- 238000011896 sensitive detection Methods 0.000 description 3
- 102000004190 Enzymes Human genes 0.000 description 2
- 108090000790 Enzymes Proteins 0.000 description 2
- 201000011510 cancer Diseases 0.000 description 2
- 238000003745 diagnosis Methods 0.000 description 2
- 239000000439 tumor marker Substances 0.000 description 2
- 238000002965 ELISA Methods 0.000 description 1
- 238000003556 assay Methods 0.000 description 1
- 230000005540 biological transmission Effects 0.000 description 1
- 238000001311 chemical methods and process Methods 0.000 description 1
- 230000007812 deficiency Effects 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 229940088597 hormone Drugs 0.000 description 1
- 239000005556 hormone Substances 0.000 description 1
- 230000001900 immune effect Effects 0.000 description 1
- 230000028993 immune response Effects 0.000 description 1
- 231100000053 low toxicity Toxicity 0.000 description 1
- 230000002503 metabolic effect Effects 0.000 description 1
- 239000002245 particle Substances 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 230000004044 response Effects 0.000 description 1
- 230000035945 sensitivity Effects 0.000 description 1
- 238000011895 specific detection Methods 0.000 description 1
- 238000005728 strengthening Methods 0.000 description 1
- 238000012956 testing procedure Methods 0.000 description 1
- 238000012546 transfer Methods 0.000 description 1
- 210000004881 tumor cell Anatomy 0.000 description 1
Landscapes
- Investigating Or Analysing Materials By The Use Of Chemical Reactions (AREA)
Abstract
The invention belongs to the technical field of nanometer functional materials, immunoassay and biosensing and provides a preparation method and application of an electrochemiluminescence immunosensor constructed on the basis of GO/MWCNTs-COOH/Au @ CeO2. According to the electrochemiluminescence immunosensor, GO/MWCNTs-COOH/Au is adopted as the substrate material, and cerium dioxide is adopted as the luminescent material; the electrochemiluminescence immunosensor has important practicality in early diagnosis of tumors and clinical application.
Description
Technical field
The present invention relates to a kind of based on GO/MWCNTs-COOH/AuCeO
2the preparation method of the electrochemiluminescence immunosensor built and application.Specifically adopt GO/MWCNTs-COOH/Au as base material and CeO
2as luminescent material, prepare a kind of unmarked type electrochemiluminescence immunosensor detecting tumor markers, belong to new function material and bio-sensing detection technique field.
Background technology
Cancer is the general designation of a large class malignant tumour.The incidence of disease of tumour is high, and growth and transfer velocity soon, have great harm to the health of the mankind.Tumor marker be tumour cell produce and release with the metabolic product of the forms such as antigen, enzyme, hormone, and can be used for diagnosis and the auxiliary diagnosis of some tumour clinical.Therefore, in clinical research, develop the extensive concern that a kind of quick, easy, sensitive detection tumor markers method causes people.
The clinical testing procedure of current existing tumor markers is a lot, as radiommunoassay, enzyme-linked immuno assay, chemiluminescence immune assay, and Timed resolved fluoroimmunoassay etc.Immunosensor is a kind of biology sensor combined with analytical chemistry method by immunological method, is combined by the characteristic between antigen with antibody, makes it have the advantages such as high sensitivity, high selectivity, analysis be quick and easy and simple to handle.
Electrochemiluminescence immunosensor have highly sensitive, selectivity good, easy and simple to handle, be easy to miniaturization, can the advantage such as continuous, rapid automatized detection analysis, therefore the present invention has prepared a kind of based on GO/MWCNTs-COOH/AuCeO
2electrochemiluminescence immunosensor, achieves the detection to tumor markers.
Graphene oxide and carboxylic carbon nano-tube have the advantages such as specific surface area is large, electric conductivity is excellent, carboxylic carbon nano-tube better water-soluble, and it can improve the stability of this electrochemiluminescence immunosensor.Au NPs can improve the electric conductivity of compound substance, thus improves electron transport ability.Therefore the graphene oxide modified by golden nanometer particle of the present invention and carboxylic carbon nano-tube are incorporated in the preparation of immunosensor, utilize its specific surface area large, the features such as electron transmission ability is excellent, thus play the effect strengthening electrochemical luminescence signals.CeO
2there is low toxicity, good biocompatibility, the features such as chemical stability is good.Therefore, the present invention utilizes the good biocompatibility of Au atom, achieves antibody fixing at electrode surface; By compound substance GO/MWCNTs-COOH/AuCeO
2introduce in the preparation of sensor, construct a kind of overdelicate unmarked type electrochemiluminescence immunosensor.CeO
2apply in immunosensor as electrochemiluminescence material first time.Compound substance GO/MWCNTs-COOH/AuCeO
2good electrochemical luminescence signals can be produced in testing process, significantly reduce the detection limit of sensor, can be used for the analysis of Diagnostic Value of Several Serum Tumor Markers.The method has that cost is low, highly sensitive, specificity is good, detect the advantages such as quick, and preparation process is comparatively simple, effectively overcomes the deficiency of current tumor-marker object detecting method.
Summary of the invention
(1) based on compound substance GO/MWCNTs-COOH/AuCeO
2, construct a kind of without enzyme, quick and sensitive unmarked type electrochemiluminescence immunosensor.
(2) this unmarked type electrochemiluminescence immunosensor is applied to the detection of Diagnostic Value of Several Serum Tumor Markers.
technical scheme of the present invention is as follows:
1. a kind of based on GO/MWCNTs-COOH/AuCeO
2the preparation method of the electrochemiluminescence immunosensor built
(1) by diameter be the glass-carbon electrode Al of 4 mm
2o
3burnishing powder is polished, and ultrapure water cleans up;
(2) GO/MWCNTs-COOH/AuCeO of 6 μ L, 2.0 ~ 14.0 mg/mL is got
2chitosan solution is added drop-wise to electrode surface, dries under room temperature;
(3) the tumor markers antibody of 6 μ L, 6 ~ 12 μ g/mL is dripped, ultrapure water electrode surface, dry in 4 DEG C of refrigerators;
(4) drip 3 μ L, massfraction be the BSA solution of 1 ~ 2% in order to enclosed-electrode nonspecific activity site on the surface, ultrapure water electrode surface, dries in 4 DEG C of refrigerators;
(5) the tumor markers antigenic solution of a series of variable concentrations of 6 μ L, 0.05 ~ 100 ng/mL is dripped, dry in 4 DEG C of refrigerators, obtained a kind of based on GO/MWCNTs-COOH/AuCeO
2the electrochemiluminescence immunosensor built.
2. the preparation of graphene oxide
By 36 mL, the sulfuric acid of 98% and 4 mL, the phosphoric acid of 85% adds in the there-necked flask of 500 mL, even by magnetic stirrer, adds 300 mg dags, stirs; Slowly add the potassium permanganate powder of 1.8 g; Under 50 C oil baths, magnetic agitation 12 h, is cooled to room temperature; Under agitation, pour in 40 mL ice; Add 300 μ L, massfraction is the superoxol of 30%, and Keep agitation 1 h; With rotating speed 9000 r/min, centrifuging 10 min, removes supernatant; With the hydrochloric acid solution washing precipitation twice of 0.2 mol/L, absolute ethanol washing precipitation twice, finally uses absolute ether washing precipitation one time; Vacuum drying 12 h under 35 C; Because the volatility of ether is comparatively strong, easily splash, to be placed in large centrifuge tube when vacuum drying, wraps by filter paper packet, with antispray; Finally obtain yellow solid powder, ground with agate mortar, weigh, obtained graphene oxide.
3. GO/MWCNTs-COOH/AuCeO
2preparation
100 mg graphene oxide powder are dissolved in 100 mL ultrapure waters, ultrasonic 2 h; Add 100 mg carboxylic carbon nano-tubes, ultrasonic 1 h; Add the Ce (NO of 2.17 g
3)
36H
2o, Keep agitation 30 min; Add the NaOH of 10 mg, make pH value of solution be 10, under continuous stirring state, add NH
3h
2o, makes pH value of solution be 11; Add 5mL, massfraction is the HAuCl of 1%
44H
2o solution, gained suspending liquid is heated to 80 ° of C and maintains 6 h, centrifugal treating, and the sediment ultrapure water obtained and washes of absolute alcohol are washed 3 times, and product is dry 12 h in 40 ° of C vacuum drying chambers, in 300 C tubular furnace, calcine 1 h.
4.the detection method of Diagnostic Value of Several Serum Tumor Markers
(1) use electrochemical workstation with three-electrode system, saturated Ag/AgCl electrode is contrast electrode, platinum electrode is auxiliary electrode, and prepared sensor is working electrode, includes 0.1 mol/L KCl and 20 ~ 140 mmol/L K at the pH 7.5 ~ 8.5 of 10 mL, 50 mmol/L
2s
2o
8pBS buffer solution in test;
(2) with MPI-F Flow Injection Analysis/Chemiluminescence instrument, analysis thing is detected, end liquid be 10 mL, pH 8.0 containing 0.1 mol/L KCl and 120 mmol/L K
2s
2o
8pBS buffer solution in scan, scanning voltage scope is-2 ~ 0 V, and photomultiplier high pressure is 750 V, and sweep speed is 0.1 V s
-1;
(3) after peak value tends towards stability, record electrochemiluminescence spectrum.
5.above-mentioned tumor markers is selected from one of following: CEA, AFP, parathyroid hormone PTH squamous cell related antigen SCCA, people's nuclear matrix protein NMP-22, prostate specific antigen PSA, human chorionic gonadotrophin HCG, acid phosphatase ACP, human placental prolactin HPL, oophoroma CA125, breast cancer susceptibility gene CAl5-3, glycoprotein antigen CA50, CA19-9, CA549, CA72-4, cytokeratin, phosphatization Proteins p53, alkaline phosphatase, NSE NSE, corticotropin ACTH, growth hormone GH.
useful achievement of the present invention
(1) GO/MWCNTs-COOH has larger surface area can increase the charge capacity of Au NPs on its surface, and then increase the charge capacity of antibody, in addition, its electronic conduction ability is excellent, be incorporated in electrochemiluminescence immunosensor, make sensor achieve Sensitive Detection to tumor markers.
(2) CeO
2one step reverts on base material GO/MWCNTs-COOH, compound substance GO/MWCNTs-COOH/AuCeO
2prepare comparatively simple.CeO
2have strong and stable electrochemical luminescence signals, in immunosensor, first time is applied as electrochemiluminescence material.
(3) use of MWCNTs-COOH, had both utilized the electronic conduction ability that it is good compared with Large ratio surface sum, can increase the water-soluble of base material again, made it have more stable electrochemical luminescence signals, thus improved the stability of sensor preferably.
(4), in the middle of preparation shitosan being applied to electrochemical immunosensor, utilize the filming performance of its excellence, prevent compound substance GO/MWCNTs-COOH/AuCeO
2come off from electrode surface, utilize shitosan to have good biocompatibility and hypotoxicity simultaneously, be conducive to antibody fixing at electrode surface.
(5) the present invention utilizes the immune response of antigen, antibody, improves the specificity of detection method.
(6) the electrochemiluminescence immunosensor prepared of the present invention is for the detection of Diagnostic Value of Several Serum Tumor Markers, response time is short, and detectability is low, and the range of linearity is wide, simple, quick, highly sensitive and specific detection can be realized, 0.02 ng/mL can be reached to common tumor markers detectability.
Embodiment
embodiment 1a kind of based on GO/MWCNTs-COOH/AuCeO
2the preparation of electrochemiluminescence immunosensor
(1) by diameter be the glass-carbon electrode Al of 4 mm
2o
3burnishing powder is polished, and ultrapure water cleans up;
(2) GO/MWCNTs-COOH/AuCeO of 6 μ L, 2.0 mg/mL is got
2chitosan solution is added drop-wise to electrode surface, dries under room temperature;
(3) the tumor markers antibody of 6 μ L, 6 μ g/mL is dripped, ultrapure water electrode surface, dry in 4 DEG C of refrigerators;
(4) drip 3 μ L, BSA solution that massfraction is 1 % in order to enclosed-electrode nonspecific activity site on the surface, ultrapure water electrode surface, dries in 4 DEG C of refrigerators;
(5) the tumor markers antigenic solution of a series of variable concentrations of 6 μ L, 0.05 ~ 100 ng/mL is dripped, dry in 4 DEG C of refrigerators, obtained a kind of based on GO/MWCNTs-COOH/AuCeO
2the electrochemiluminescence immunosensor built.
embodiment 2a kind of based on GO/MWCNTs-COOH/AuCeO
2the preparation of electrochemiluminescence immunosensor
(1) by diameter be the glass-carbon electrode Al of 4 mm
2o
3burnishing powder is polished, and ultrapure water cleans up;
(2) GO/MWCNTs-COOH/AuCeO of 6 μ L, 8.0 mg/mL is got
2chitosan solution is added drop-wise to electrode surface, dries under room temperature;
(3) the tumor markers antibody of 6 μ L, 10 μ g/mL is dripped, ultrapure water electrode surface, dry in 4 DEG C of refrigerators;
(4) drip 3 μ L, massfraction be the BSA solution of 1.5% in order to enclosed-electrode nonspecific activity site on the surface, ultrapure water electrode surface, dries in 4 DEG C of refrigerators;
(5) the tumor markers antigenic solution of a series of variable concentrations of 6 μ L, 0.05 ~ 100 ng/mL is dripped, dry in 4 DEG C of refrigerators, obtained a kind of based on GO/MWCNTs-COOH/AuCeO
2the electrochemiluminescence immunosensor built.
embodiment 3a kind of based on GO/MWCNTs-COOH/AuCeO
2the preparation of electrochemiluminescence immunosensor
(1) by diameter be the glass-carbon electrode Al of 4 mm
2o
3burnishing powder is polished, and ultrapure water cleans up;
(2) GO/MWCNTs-COOH/AuCeO of 6 μ L, 14.0 mg/mL is got
2chitosan solution is added drop-wise to electrode surface, dries under room temperature;
(3) the tumor markers antibody of 6 μ L, 12 μ g/mL is dripped, ultrapure water electrode surface, dry in 4 DEG C of refrigerators;
(4) drip 3 μ L, massfraction be the BSA solution of 2% in order to enclosed-electrode nonspecific activity site on the surface, ultrapure water electrode surface, dries in 4 DEG C of refrigerators;
(5) the tumor markers antigenic solution of a series of variable concentrations of 6 μ L, 0.05 ~ 100 ng/mL is dripped, dry in 4 DEG C of refrigerators, obtained a kind of based on GO/MWCNTs-COOH/AuCeO
2the electrochemiluminescence immunosensor built.
embodiment 4electrochemiluminescence immunosensor is used for the detection of CEA
(1) use electrochemical workstation with three-electrode system, saturated Ag/AgCl electrode is contrast electrode, and platinum electrode is auxiliary electrode, and prepared sensor is working electrode, at 10 mL, pH 7.5 ~ 8.5 containing 0.1 mol/L KCl and 20 ~ 140 mmol/L K
2s
2o
8pBS buffer solution in test;
(2) with MPI-F Flow Injection Analysis/Chemiluminescence instrument, analysis thing is detected, end liquid be 10 mL, pH 8.0 containing 0.1 mol/L KCl and 120 mmol/L K
2s
2o
8pBS buffer solution in scan, scanning voltage scope is-2 ~ 0 V, and photomultiplier high pressure is 750 V, and sweep speed is 0.1 V s
-1;
(3) after peak value tends towards stability, record electrochemiluminescence spectrum.
(4) according to the linear relationship between gained electrochemiluminescence intensity and CEA concentration, drawing curve, recording the range of linearity is 0.05 ~ 100 ng/mL, detects and is limited to 0.02 ng/mL.
embodiment 5electrochemiluminescence immunosensor is used for the detection of AFP
Operate according to embodiment 4, according to the linear relationship between gained electrochemiluminescence intensity and AFP concentration, drawing curve, recording the range of linearity is 0.04 ~ 110 ng/mL, detects and is limited to 0.02 ng/mL.
embodiment 6electrochemiluminescence immunosensor is used for the detection of oophoroma CA125
Operate according to embodiment 4, according to the linear relationship between gained electrochemiluminescence intensity and oophoroma CA125 concentration, drawing curve, recording the range of linearity is 0.04 ~ 100 ng/mL, detects and is limited to 0.02 ng/mL.
embodiment 7electrochemiluminescence immunosensor is used for the detection of squamous cell related antigen SCCA
Operate according to embodiment 4, according to the linear relationship between gained electrochemiluminescence intensity and squamous cell related antigen SCCA concentration, drawing curve, recording the range of linearity is 0.06 ~ 100 ng/mL, detects and is limited to 0.03 ng/mL.
embodiment 8electrochemiluminescence immunosensor is used for the detection of prostate specific antigen PSA
Operate according to embodiment 4, according to the linear relationship between gained electrochemiluminescence intensity and prostate specific antigen PSA concentration, drawing curve, recording the range of linearity is 0.05 ~ 110 ng/mL, detects and is limited to 0.02 ng/mL.
embodiment 9electrochemiluminescence immunosensor is used for the detection of people's nuclear matrix protein NMP-22
Operate according to embodiment 4, according to the linear relationship between gained electrochemiluminescence intensity and people's nuclear matrix protein NMP-22 concentration, drawing curve, recording the range of linearity is 0.04 ~ 120 ng/mL, detects and is limited to 0.02 ng/mL.
embodiment 10electrochemiluminescence immunosensor is used for the detection of breast cancer susceptibility gene CAl5-3
Operate according to embodiment 4, according to the linear relationship between gained electrochemiluminescence intensity and breast cancer susceptibility gene CAl5-3 concentration, drawing curve, recording the range of linearity is 0.03 ~ 100 ng/mL, detects and is limited to 0.01 ng/mL.
embodiment 11electrochemiluminescence immunosensor is used for the detection of glycoprotein antigen CA50
Operate according to embodiment 4, according to the linear relationship between gained electrochemiluminescence intensity and glycoprotein antigen CA50 concentration, drawing curve, recording the range of linearity is 0.05 ~ 100 ng/mL, detects and is limited to 0.02 ng/mL.
Claims (5)
1. one kind based on GO/MWCNTs-COOH/AuCeO
2the preparation method of the electrochemiluminescence immunosensor built, is characterized in that, comprise following step:
(1) by diameter be the glass-carbon electrode Al of 4 mm
2o
3burnishing powder is polished, and ultrapure water cleans up;
(2) GO/MWCNTs-COOH/AuCeO of 6 μ L, 2.0 ~ 14.0 mg/mL is got
2chitosan solution is added drop-wise to electrode surface, dries under room temperature;
(3) the tumor markers antibody of 6 μ L, 6 ~ 12 μ g/mL is dripped, ultrapure water electrode surface, dry in 4 DEG C of refrigerators;
(4) drip 3 μ L, massfraction be the BSA solution of 1 ~ 2% in order to enclosed-electrode nonspecific activity site on the surface, ultrapure water electrode surface, dries in 4 DEG C of refrigerators;
(5) the tumor markers antigenic solution of a series of variable concentrations of 6 μ L, 0.05 ~ 100 ng/mL is dripped, dry in 4 DEG C of refrigerators, obtained a kind of based on GO/MWCNTs-COOH/AuCeO
2the electrochemiluminescence immunosensor built.
2. as claimed in claim 1 a kind of based on GO/MWCNTs-COOH/AuCeO
2the preparation method of the electrochemiluminescence immunosensor built, the preparation of described graphene oxide, is characterized in that, step is as follows:
By 36 mL, the sulfuric acid of 98% and 4 mL, the phosphoric acid of 85% adds in the there-necked flask of 500 mL, even by magnetic stirrer, adds 300 mg dags, stirs; Slowly add the potassium permanganate powder of 1.8 g; Under 50 C oil baths, magnetic agitation 12 h, is cooled to room temperature; Under agitation, pour in 40 mL ice; Add 300 μ L, massfraction is the superoxol of 30%, and Keep agitation 1 h; With rotating speed 9000 r/min, centrifuging 10 min, removes supernatant; With the hydrochloric acid solution washing precipitation twice of 0.2 mol/L, absolute ethanol washing precipitation twice, finally uses absolute ether washing precipitation one time; Vacuum drying 12 h under 35 C; Because the volatility of ether is comparatively strong, easily splash, to be placed in large centrifuge tube when vacuum drying, wraps by filter paper packet, with antispray; Finally obtain yellow solid powder, ground with agate mortar, weigh, obtained graphene oxide.
3. as claimed in claim 1 a kind of based on GO/MWCNTs-COOH/AuCeO
2the preparation method of the electrochemiluminescence immunosensor built, is characterized in that, described GO/MWCNTs-COOH/AuCeO
2preparation, step is as follows:
100 mg graphene oxide powder are dissolved in 100 mL ultrapure waters, ultrasonic 2 h; Add 100 mg carboxylic carbon nano-tubes, ultrasonic 1 h; Add the Ce (NO of 2.17 g
3)
36H
2o, Keep agitation 30 min; Add the NaOH of 10 mg, make pH value of solution be 10, under continuous stirring state, add NH
3h
2o, makes pH value of solution be 11; Add 5mL, massfraction is the HAuCl of 1%
44H
2o solution, gained suspending liquid is heated to 80 ° of C and maintains 6 h, centrifugal treating, and the sediment ultrapure water obtained and washes of absolute alcohol are washed 3 times, and product is dry 12 h in 40 ° of C vacuum drying chambers, in 300 C tubular furnace, calcine 1 h.
4. the sensor prepared of preparation method as claimed in claim 1 is for the detection method of Diagnostic Value of Several Serum Tumor Markers, and it is characterized in that, step is as follows:
(1) use electrochemical workstation with three-electrode system, saturated Ag/AgCl electrode is contrast electrode, platinum electrode is auxiliary electrode, and prepared sensor is working electrode, includes 0.1 mol/L KCl and 20 ~ 140 mmol/L K at the pH 7.5 ~ 8.5 of 10 mL, 50 mmol/L
2s
2o
8pBS buffer solution in test;
(2) with MPI-F Flow Injection Analysis/Chemiluminescence instrument, analysis thing is detected, end liquid be 10 mL, pH 8.0 containing 0.1 mol/L KCl and 120 mmol/L K
2s
2o
8pBS buffer solution in scan, scanning voltage scope is-2 ~ 0 V, and photomultiplier high pressure is 750 V, and sweep speed is 0.1 V s
-1;
(3) after peak value tends towards stability, record electrochemiluminescence spectrum.
5. one according to claim 1 is based on GO/MWCNTs-COOH/AuCeO
2the electrochemiluminescence immunosensor built, described tumor markers is selected from one of following: CEA, AFP, parathyroid hormone PTH squamous cell related antigen SCCA, people's nuclear matrix protein NMP-22, prostate specific antigen PSA, human chorionic gonadotrophin HCG, acid phosphatase ACP, human placental prolactin HPL, oophoroma CA125, breast cancer susceptibility gene CAl5-3, glycoprotein antigen CA50, CA19-9, CA549, CA72-4, cytokeratin, phosphatization Proteins p53, alkaline phosphatase, NSE NSE, corticotropin ACTH, growth hormone GH.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201510262821.7A CN104880456B (en) | 2015-05-22 | 2015-05-22 | A kind of based on GO/MWCNTs-COOH/Au@CeO2the preparation method and application of the electrochemiluminescence immunosensor built |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201510262821.7A CN104880456B (en) | 2015-05-22 | 2015-05-22 | A kind of based on GO/MWCNTs-COOH/Au@CeO2the preparation method and application of the electrochemiluminescence immunosensor built |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN104880456A true CN104880456A (en) | 2015-09-02 |
| CN104880456B CN104880456B (en) | 2016-10-12 |
Family
ID=53948031
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201510262821.7A Expired - Fee Related CN104880456B (en) | 2015-05-22 | 2015-05-22 | A kind of based on GO/MWCNTs-COOH/Au@CeO2the preparation method and application of the electrochemiluminescence immunosensor built |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN104880456B (en) |
Cited By (11)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105758920A (en) * | 2015-11-05 | 2016-07-13 | 佳木斯大学 | Preparation method of electrochemical micro-detection system based on disposable pipette tip and lead core electrode array |
| CN105842460A (en) * | 2016-03-25 | 2016-08-10 | 济南大学 | Preparation method of electro-chemiluminescence immunosensor based on silver-hybridized bismuth sulfide |
| CN105911128A (en) * | 2016-04-13 | 2016-08-31 | 中国石油大学(华东) | Enzyme-free glucose electrochemical sensor and application thereof |
| CN106290521A (en) * | 2016-09-30 | 2017-01-04 | 重庆医科大学 | A kind of electrochemical sensor preparation method for ADRB1 1165G ＞ C genetic polymorphism detection |
| CN106442994A (en) * | 2016-09-14 | 2017-02-22 | 山东理工大学 | Preparation method and application of electrochemical immunosensor based on Ag@Au nanocomposite |
| CN106802314A (en) * | 2017-03-20 | 2017-06-06 | 山东理工大学 | A kind of preparation method and application of the immunosensor of the Graphene based on load TaC and gold cladding decahedron Nano silver grain |
| CN108927116A (en) * | 2018-07-20 | 2018-12-04 | 大连医科大学 | A kind of preparation method for the stannic oxide/graphene nano material that phenyl boric acid functionalization is chitosan-modified |
| CN108982481A (en) * | 2018-07-31 | 2018-12-11 | 济南大学 | A kind of preparation method and application of the electrochemical luminescence sensor based on the titanium doped bromine oxygen bismuth microballoon of golden functionalization |
| CN109342527A (en) * | 2018-11-26 | 2019-02-15 | 阜阳师范学院 | It is a kind of for measuring the preparation method of phenyl ethylamine electrode |
| CN110455786A (en) * | 2019-08-26 | 2019-11-15 | 济南大学 | One kind being based on CeO2@SnS2Promote the preparation method of luminol Electrochemiluminescsensor sensor |
| CN110702754A (en) * | 2019-11-08 | 2020-01-17 | 广西师范大学 | Novel method for measuring human chorionic gonadotropin |
Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101995402B (en) * | 2010-10-15 | 2012-08-29 | 济南大学 | Preparation and application of electrochemiluminescence sensor for detecting trace pesticide residue |
| CN103018231A (en) * | 2012-12-27 | 2013-04-03 | 济南大学 | Preparation method and application of composite nano material paper chip electrochemical luminescence immunosensor |
| CN103913496A (en) * | 2014-02-28 | 2014-07-09 | 汕头大学 | Preparation method and application of modified glassy carbon electrode |
| CN102520036B (en) * | 2011-12-30 | 2014-08-13 | 汕头大学 | Method for fixing terpyridyl ruthenium on surface of electrochemical electrode |
| CN104007153A (en) * | 2014-06-09 | 2014-08-27 | 上海师范大学 | Electrochemical-luminescence sensor for quickly detecting diphenhydramine hydrochloride by high sensitivity and preparation method thereof |
| US20150093840A1 (en) * | 2013-09-27 | 2015-04-02 | National Tsing Hua University | Enzyme-free colorimetric immunoassay |
-
2015
- 2015-05-22 CN CN201510262821.7A patent/CN104880456B/en not_active Expired - Fee Related
Patent Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101995402B (en) * | 2010-10-15 | 2012-08-29 | 济南大学 | Preparation and application of electrochemiluminescence sensor for detecting trace pesticide residue |
| CN102520036B (en) * | 2011-12-30 | 2014-08-13 | 汕头大学 | Method for fixing terpyridyl ruthenium on surface of electrochemical electrode |
| CN103018231A (en) * | 2012-12-27 | 2013-04-03 | 济南大学 | Preparation method and application of composite nano material paper chip electrochemical luminescence immunosensor |
| US20150093840A1 (en) * | 2013-09-27 | 2015-04-02 | National Tsing Hua University | Enzyme-free colorimetric immunoassay |
| CN103913496A (en) * | 2014-02-28 | 2014-07-09 | 汕头大学 | Preparation method and application of modified glassy carbon electrode |
| CN104007153A (en) * | 2014-06-09 | 2014-08-27 | 上海师范大学 | Electrochemical-luminescence sensor for quickly detecting diphenhydramine hydrochloride by high sensitivity and preparation method thereof |
Non-Patent Citations (2)
| Title |
|---|
| R.RAJENDRAN, ET AL.: "Dimensionally integrated nanoarchitectonics for a novel composite from 0D,1D,and 2D nanomaterials: RGO/CNT/CeO2 ternary nanocomposites with electrochemical performance", <J.MATER.CHEM.A>, no. 2, 28 February 2014 (2014-02-28), pages 18480 - 18487 * |
| 李俊华等: "基于氧化石墨烯/碳纳米管复合薄膜修饰电极制备L-色氨酸电化学传感器", <分析化学>, vol. 41, no. 1, 31 January 2013 (2013-01-31), pages 98 - 104 * |
Cited By (18)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105758920B (en) * | 2015-11-05 | 2019-06-14 | 佳木斯大学 | A kind of preparation method of the micro- detection system of electrochemistry based on disposable liquid transfer gun head and lead for retractable pencil electrod-array |
| CN105758920A (en) * | 2015-11-05 | 2016-07-13 | 佳木斯大学 | Preparation method of electrochemical micro-detection system based on disposable pipette tip and lead core electrode array |
| CN105842460B (en) * | 2016-03-25 | 2017-10-17 | 济南大学 | A kind of preparation method of the electrochemiluminescimmunosensor immunosensor based on silver-colored hydridization bismuth sulfide |
| CN105842460A (en) * | 2016-03-25 | 2016-08-10 | 济南大学 | Preparation method of electro-chemiluminescence immunosensor based on silver-hybridized bismuth sulfide |
| CN105911128B (en) * | 2016-04-13 | 2018-11-09 | 中国石油大学(华东) | A kind of enzyme-free glucose electrochemical sensor and its application |
| CN105911128A (en) * | 2016-04-13 | 2016-08-31 | 中国石油大学(华东) | Enzyme-free glucose electrochemical sensor and application thereof |
| CN106442994A (en) * | 2016-09-14 | 2017-02-22 | 山东理工大学 | Preparation method and application of electrochemical immunosensor based on Ag@Au nanocomposite |
| CN106442994B (en) * | 2016-09-14 | 2018-01-23 | 山东理工大学 | A kind of preparation method and application of the electrochemical immunosensor based on Ag@Au nano composite materials |
| CN106290521B (en) * | 2016-09-30 | 2020-02-07 | 重庆医科大学 | Preparation method of electrochemical sensor for ADRB1-1165G & gtC gene polymorphism detection |
| CN106290521A (en) * | 2016-09-30 | 2017-01-04 | 重庆医科大学 | A kind of electrochemical sensor preparation method for ADRB1 1165G ＞ C genetic polymorphism detection |
| CN106802314A (en) * | 2017-03-20 | 2017-06-06 | 山东理工大学 | A kind of preparation method and application of the immunosensor of the Graphene based on load TaC and gold cladding decahedron Nano silver grain |
| CN108927116A (en) * | 2018-07-20 | 2018-12-04 | 大连医科大学 | A kind of preparation method for the stannic oxide/graphene nano material that phenyl boric acid functionalization is chitosan-modified |
| CN108982481A (en) * | 2018-07-31 | 2018-12-11 | 济南大学 | A kind of preparation method and application of the electrochemical luminescence sensor based on the titanium doped bromine oxygen bismuth microballoon of golden functionalization |
| CN109342527A (en) * | 2018-11-26 | 2019-02-15 | 阜阳师范学院 | It is a kind of for measuring the preparation method of phenyl ethylamine electrode |
| CN109342527B (en) * | 2018-11-26 | 2020-09-01 | 阜阳师范学院 | Preparation method of electrode for measuring phenylethylamine |
| CN110455786A (en) * | 2019-08-26 | 2019-11-15 | 济南大学 | One kind being based on CeO2@SnS2Promote the preparation method of luminol Electrochemiluminescsensor sensor |
| CN110455786B (en) * | 2019-08-26 | 2021-08-20 | 济南大学 | Based on CeO2@SnS2Preparation method of luminol-promoted electrochemiluminescence sensor |
| CN110702754A (en) * | 2019-11-08 | 2020-01-17 | 广西师范大学 | Novel method for measuring human chorionic gonadotropin |
Also Published As
| Publication number | Publication date |
|---|---|
| CN104880456B (en) | 2016-10-12 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN104880456B (en) | A kind of based on GO/MWCNTs-COOH/Au@CeO2the preparation method and application of the electrochemiluminescence immunosensor built | |
| CN103116023B (en) | ECL (electrochemiluminescence) immunosensor for detecting tumor markers and preparation method and applications thereof | |
| CN108896638B (en) | Preparation method and application of immunosensor based on titanium dioxide doped graphene loaded sea cucumber-like gold-palladium core-shell nanoparticles | |
| CN102980925B (en) | Sandwich type electrochemical immunosensor, preparation method and application thereof | |
| CN104459132B (en) | A kind of is preparation method and the application of the cancer of pancreas immunosensor of label based on golden electro-deposition and Au@Ag/CuO-GS | |
| CN104391117B (en) | Preparation method and application of gastric cancer antigen electrogenerated chemiluminescence sensor based on PP<y>-NH2GO-Ag2Se@CdSe | |
| CN104122309B (en) | A kind of preparation of cyclodextrin-Cu@Ag electrochemical immunosensor | |
| CN105319254B (en) | Preparation and application of electrochemical immunosensor based on Pt/PdCu-three-dimensional graphene markers | |
| CN105301241B (en) | Preparing method and application of sandwich type electrochemical luminescence immunosensor for detecting tumor marker | |
| CN104391113B (en) | A kind of preparation method of the biology sensor based on golden copper nano-wire-three mercaptopropyltriethoxysilane functionalized graphene structure and application | |
| CN107328930B (en) | A kind of preparation and application based on dual signal response ratio type screen printing electrode immunosensor | |
| CN108226252B (en) | Preparation method and application of current type immunosensor for detecting breast cancer | |
| CN104655616A (en) | Preparation method and application of electrochemiluminescence aptamer sensor for detecting tumor marker MUC1 | |
| CN104391123B (en) | A kind of preparation method of the biology sensor built based on flower-like nanometer ZnO microsphere and golden palladium nano flower composite material and application | |
| CN105954339A (en) | Preparation method and application of sandwich type immunosensor based on CeO2@Cu2O/Au@Pt | |
| CN105241939A (en) | Preparation method for immunosensor based on gold/silver core-shell magnetic graphene adsorption cadmium ion and application | |
| CN104297478B (en) | A kind of preparation method of the immunosensor based on acid site compound and application | |
| CN105842460A (en) | Preparation method of electro-chemiluminescence immunosensor based on silver-hybridized bismuth sulfide | |
| CN109613244A (en) | A kind of preparation method and application of the immunosensor of Ag@Pt-CuS label | |
| CN105115961A (en) | Method for preparing electrochemical luminescence sensor made of nano-composites | |
| CN104569427A (en) | Preparation method and application of immunosensor established based on manganese dioxide loaded silver nanoparticle multiwalled carbon nanotube | |
| CN110058020A (en) | A kind of preparation method and application of the electrochemical immunosensor of PdCu nano wire functionalization porous graphene | |
| CN105606681B (en) | A kind of preparation method and application of the biosensor based on golden copper-multi-walled carbon nanotube-manganese dioxide structure | |
| CN104931698A (en) | Preparation method and application of NP-NiGd@Au-based gastric cancer marker gold nano-cluster electrogenerated chemiluminescence sensor | |
| CN104849458B (en) | A kind of based on KNbO3-Au NPs@Bi2s3the preparation method and application of the electrochemical luminescence immunosensor built |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| EXSB | Decision made by sipo to initiate substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| CB03 | Change of inventor or designer information |
Inventor after: Wei Qin Inventor after: Wu Dan Inventor after: Yan Tao Inventor after: Li Yueyun Inventor after: Li Jianxiu Inventor after: Ma Hongmin Inventor after: Pang Xuehui Inventor after: Wang Xiaodong Inventor after: Du Bin Inventor after: Fan Dawei Inventor after: Cao Wei Inventor after: Hu Lihua Inventor before: Li Jianxiu Inventor before: Wu Dan Inventor before: Yan Tao Inventor before: Li Yueyun Inventor before: Wei Qin Inventor before: Ma Hongmin Inventor before: Pang Xuehui Inventor before: Wang Xiaodong Inventor before: Du Bin Inventor before: Fan Dawei Inventor before: Cao Wei Inventor before: Hu Lihua |
|
| COR | Change of bibliographic data | ||
| CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20161012 |
|
| CF01 | Termination of patent right due to non-payment of annual fee |