CN104990913A - Gold nanoparticle growth based method for detection of glucose - Google Patents
Gold nanoparticle growth based method for detection of glucose Download PDFInfo
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- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 title claims abstract description 77
- 239000008103 glucose Substances 0.000 title claims abstract description 77
- 238000000034 method Methods 0.000 title claims abstract description 30
- PCHJSUWPFVWCPO-UHFFFAOYSA-N gold Chemical compound [Au] PCHJSUWPFVWCPO-UHFFFAOYSA-N 0.000 title claims abstract description 23
- 239000010931 gold Substances 0.000 title claims abstract description 23
- 229910052737 gold Inorganic materials 0.000 title claims abstract description 23
- 238000001514 detection method Methods 0.000 title claims abstract description 14
- 239000002105 nanoparticle Substances 0.000 title claims abstract description 7
- 108010015776 Glucose oxidase Proteins 0.000 claims abstract description 38
- 239000004366 Glucose oxidase Substances 0.000 claims abstract description 38
- 229940116332 glucose oxidase Drugs 0.000 claims abstract description 38
- 235000019420 glucose oxidase Nutrition 0.000 claims abstract description 38
- MHAJPDPJQMAIIY-UHFFFAOYSA-N Hydrogen peroxide Chemical compound OO MHAJPDPJQMAIIY-UHFFFAOYSA-N 0.000 claims abstract description 30
- 238000002835 absorbance Methods 0.000 claims abstract description 12
- 238000011534 incubation Methods 0.000 claims abstract description 10
- 239000002245 particle Substances 0.000 claims description 30
- FDWREHZXQUYJFJ-UHFFFAOYSA-M gold monochloride Chemical compound [Cl-].[Au+] FDWREHZXQUYJFJ-UHFFFAOYSA-M 0.000 claims description 23
- 241001247197 Cephalocarida Species 0.000 claims description 17
- 230000012447 hatching Effects 0.000 claims description 17
- 229960002163 hydrogen peroxide Drugs 0.000 claims description 14
- 239000008363 phosphate buffer Substances 0.000 claims description 9
- 230000002255 enzymatic effect Effects 0.000 claims description 7
- 230000009467 reduction Effects 0.000 claims description 5
- SJUCACGNNJFHLB-UHFFFAOYSA-N O=C1N[ClH](=O)NC2=C1NC(=O)N2 Chemical compound O=C1N[ClH](=O)NC2=C1NC(=O)N2 SJUCACGNNJFHLB-UHFFFAOYSA-N 0.000 claims description 3
- 238000002474 experimental method Methods 0.000 claims description 3
- 239000000243 solution Substances 0.000 abstract description 39
- 230000008859 change Effects 0.000 abstract description 5
- 102000004190 Enzymes Human genes 0.000 abstract description 4
- 108090000790 Enzymes Proteins 0.000 abstract description 4
- 229940088598 enzyme Drugs 0.000 abstract description 4
- 230000008569 process Effects 0.000 abstract description 4
- RGHNJXZEOKUKBD-UHFFFAOYSA-N D-gluconic acid Natural products OCC(O)C(O)C(O)C(O)C(O)=O RGHNJXZEOKUKBD-UHFFFAOYSA-N 0.000 abstract description 3
- RGHNJXZEOKUKBD-SQOUGZDYSA-N Gluconic acid Natural products OC[C@@H](O)[C@@H](O)[C@H](O)[C@@H](O)C(O)=O RGHNJXZEOKUKBD-SQOUGZDYSA-N 0.000 abstract description 3
- 235000012208 gluconic acid Nutrition 0.000 abstract description 3
- 239000000174 gluconic acid Substances 0.000 abstract description 3
- 239000002253 acid Substances 0.000 abstract 2
- 238000004519 manufacturing process Methods 0.000 abstract 1
- 239000008055 phosphate buffer solution Substances 0.000 abstract 1
- 238000012360 testing method Methods 0.000 description 9
- 239000008280 blood Substances 0.000 description 8
- 210000004369 blood Anatomy 0.000 description 8
- 206010012601 diabetes mellitus Diseases 0.000 description 8
- 230000008033 biological extinction Effects 0.000 description 3
- 201000010099 disease Diseases 0.000 description 3
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 3
- 102000003992 Peroxidases Human genes 0.000 description 2
- 238000006243 chemical reaction Methods 0.000 description 2
- 238000010586 diagram Methods 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 239000011259 mixed solution Substances 0.000 description 2
- 238000012216 screening Methods 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- 230000000007 visual effect Effects 0.000 description 2
- 206010008190 Cerebrovascular accident Diseases 0.000 description 1
- 206010020772 Hypertension Diseases 0.000 description 1
- 208000001647 Renal Insufficiency Diseases 0.000 description 1
- 208000006011 Stroke Diseases 0.000 description 1
- 238000013459 approach Methods 0.000 description 1
- 238000003556 assay Methods 0.000 description 1
- 230000008901 benefit Effects 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 238000004737 colorimetric analysis Methods 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 230000009849 deactivation Effects 0.000 description 1
- 230000007812 deficiency Effects 0.000 description 1
- 238000013461 design Methods 0.000 description 1
- 238000003745 diagnosis Methods 0.000 description 1
- 235000005911 diet Nutrition 0.000 description 1
- 230000000378 dietary effect Effects 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 238000013399 early diagnosis Methods 0.000 description 1
- 230000003203 everyday effect Effects 0.000 description 1
- 230000004907 flux Effects 0.000 description 1
- 108010046301 glucose peroxidase Proteins 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 208000019622 heart disease Diseases 0.000 description 1
- 150000002500 ions Chemical class 0.000 description 1
- 201000006370 kidney failure Diseases 0.000 description 1
- 239000011159 matrix material Substances 0.000 description 1
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- 230000007246 mechanism Effects 0.000 description 1
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- 108040007629 peroxidase activity proteins Proteins 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 230000035945 sensitivity Effects 0.000 description 1
- 239000000758 substrate Substances 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
Abstract
The invention relates to a gold nanoparticle growth based method for detection of glucose. The steps include: (1) adding glucose oxidase and a glucose solution with a concentration of 0-1.0mM into a phosphate buffer solution respectively, conducting incubation at room temperature for 10min to obtain an incubation solution; and 2) adding 5nm gold nanoparticles and chloroauric acid into the incubation solution, carrying out room temperature incubation for 30min, taking photos, and using a Synergy 2 micropore plate reader to read the absorbance value at the place of 530nm correspondingly. The method employs glucose oxidase to catalyze production of gluconic acid from glucose, generates hydrogen peroxide and reduces chloroauric acid into elemental gold, promotes the growth of 5nm gold nanoparticles, also causes color change so as to conduct glucose content determination. The whole detection process only needs one enzyme, i.e. glucose oxidase, has no need of expensive instrument, not only reduces the cost, but also is fast, convenient and is easy to implement.
Description
Technical field
The present invention relates to the method quantitatively detecting glucose content, particularly a kind of method detecting glucose based on gold nano growth.
Background technology
Along with the raising of people's living standard, the change of dietary structure and blue-collar minimizing, the incidence of disease of diabetes grows with each passing day.At present, about there is diabetic more than 200,000,000 in the whole world, become one of major disease threatening human health, in addition, diabetes also can cause a series of complication, as hypertension, kidney failure, blind, apoplexy, heart disease etc., therefore, the Diagnosis and Treat of diabetes always is the key subjects that world medicine faces.Blood sugar concentration is the important clinical indices of of diabetes, and the early diagnosis of diabetes can realize by detecting blood sugar, and in order to control this disease better, diabetic needs every day and measures its blood-sugar content.Therefore, develop simple, the cheap and effective method for monitor patients blood glucose levels is all very important in family expenses detection and Clinical screening.
In recent years, for improving the sensitivity of glucose sensor, specificity and reliability, scientists has paid a lot of effort.In these sensing technologies, most is representational is blood glucose meter, and it plays very important effect in monitoring diabetes with providing in clinical information.But blood glucose meter relies in galvanochemistry, electrode is very expensive and easily by matrix deactivation in testing process, cause measurement result inaccurate.In addition, blood glucose meter is mainly family expenses detection design, and is not suitable for the high flux examination that clinical laboratory carries out great amount of samples.
At present, the method that the Clinical screening of blood sugar is the most frequently used is the assay method based on colorimetric.These methods are all that the hydrogen peroxide produced by detecting glucose oxidase enzymatic glucose detects blood sugar.The detection of hydrogen peroxide then needs its chromogen substrate of Catalyzed Synthesis By Peroxidase to generate colored solutions in the presence of hydrogen peroxide.A drawback of this method is compared with glucose oxidase, and peroxidase is very expensive, and its activity is affected by environment comparatively large, uses inconvenience and cost is higher.
Summary of the invention
The present invention seeks to the deficiency overcoming prior art existence, there is provided a kind of quick, convenient, cheap in gold nano growth glucose sensing approach, the method generates gluconic acid by glucose oxidase enzymatic glucose, produce hydrogen peroxide simultaneously, hydrogen-peroxide reduction gold chloride generates golden simple substance, impel the growth of golden nanometer particle, produce color change simultaneously, carry out the mensuration of glucose content with this.Only need in whole testing process to use a kind of enzyme of glucose oxidase, testing result is visual, does not need, with expensive instrument, to reduce testing cost.
Technical scheme of the present invention:
Detect a method for glucose based on gold nano growth, comprise the steps:
1) be 0.01M, pH in concentration be mixed with glucose oxidase (GOx) respectively by the glucose solution of one group of certain concentration in the phosphate buffer of 7.2-7.4, incubated at room temperature 2-10min, obtains one group of artemia hatching solution a;
2) 5nm golden nanometer particle and gold chloride are joined in above-mentioned each artemia hatching solution respectively, incubation at room temperature 5-60min, obtain one group of artemia hatching solution b, take artemia hatching solution b photo respectively and read them at the corresponding absorbance in 530nm place with Synergy 2 microplate reader, according to concentration of glucose and corresponding absorbance fit standard curve thereof;
3) be 0.01M, pH in concentration be in the phosphate buffer of 7.2-7.4, the glucose solution sample of concentration to be measured is mixed with glucose oxidase (GOx), incubated at room temperature 2-10min, add golden nanometer particle and the chlorauric acid solution of 5nm, incubation at room temperature 5-60min, it is read at the corresponding absorbance in 530nm place with Synergy 2 microplate reader, utilize above-mentioned typical curve, calculate the concentration of glucose solution sample to be measured.
Described step 1) concentration of glucose oxidase is 1.0-3.0mg/mL in artemia hatching solution; The concentration of the glucose solution of certain concentration is respectively 0,0.1,0.3,0.5,0.7,0.9,1.0mM.
Described step 2) concentration of 5nm golden nanometer particle is 4-10nM in artemia hatching solution b, the concentration of gold chloride is 0.05-0.4mM.
Described step 3) in the concentration of golden nanometer particle and the concentration of gold chloride and step 2) in concentration identical.
The described method detecting glucose based on gold nano growth, for heterogeneous experiment, step is as follows:
1) with card punch, filter paper is broken into the filter paper that diameter is 1cm;
2) be 0.01M, pH in concentration be in the phosphate buffer of 7.3, the solution of gold nanoparticles of glucose oxidase (GOx), gold chloride and 5nm is mixed, obtain mixed liquor, mixed liquor is dripped on above-mentioned filter paper, incubated at room 2-10min;
3) on the above-mentioned scraps of paper, drip the glucose solution of one group of certain concentration, incubated at room 5-60min, glucose oxidase enzymatic glucose produces hydrogen peroxide, hydrogen-peroxide reduction gold chloride, golden nanometer particle is impelled to grow, filter paper reddens gradually, and the red weight of the scraps of paper becomes positive correlation with glucose concentration.
4) by step 2) in mixed liquor drip on filter paper, incubated at room 2-10min, drips the glucose solution of concentration to be measured, incubated at room 5-60min, with step 3) in gained filter paper carry out color contrast, the concentration of qualitative detection glucose solution.
Described step 2) concentration of glucose oxidase is the final concentration of 1.0-3.0mg/mL, 5nm golden nanometer particle in mixed liquor is 4-10nM, the final concentration of gold chloride is 0.05-0.4mM.
Described step 3) in the concentration of glucose solution of certain concentration be respectively 0,1,10,20,50,100mM.
Working mechanism of the present invention:
Golden nanometer particle has very high extinction coefficient because of it, is often used to build colorimetric methods to detect the multiple analytes such as enzyme, DNA, organic molecule, ion.And the extinction coefficient of golden nanometer particle and its size are closely related, size is larger, and extinction coefficient is larger, and therefore, at low concentrations, small size collaurum is colourless, and the large scale collaurum under same concentrations is then red.The hydrogen-peroxide reduction gold chloride utilizing glucose oxidase enzymatic glucose to produce in the present invention, impels the growth of gold nano grain, thus produces the mensuration that glucose content is carried out in color change.
Advantage of the present invention is:
This detection method generates gluconic acid by glucose oxidase enzymatic glucose, produce hydrogen peroxide, hydrogen peroxide can reduce gold chloride and generate golden simple substance, impels the golden nanometer particle of 5nm to grow simultaneously, produce color change simultaneously, carry out the mensuration of glucose content with this.Only need to use a kind of enzyme of glucose oxidase in whole testing process, testing result is visual, does not need, with expensive instrument, to not only reduce testing cost, and fast, convenient, easy to implement.
Accompanying drawing explanation
Fig. 1 is the method schematic diagram of the growth detection glucose based on golden nanometer particle.
To be concentration be Fig. 20,0.1,0.3,0.5,0.7,0.9, the testing result colour developing figure of the glucose solution of 1.0mM.
Fig. 3 is that the glucose solution of variable concentrations is at A
530nmthe fitting a straight line figure at place, wherein A
530nmread absorbance at 530nm place add gold chloride and 5nm golden nanometer particle room temperature reaction 30min in glucose oxidase and glucose mixed solution after with microplate reader.
Fig. 4 is the result colour developing figure of method on filter paper of the growth detection glucose based on golden nanometer particle.
Embodiment
Below in conjunction with embodiment, the present invention is further described.Unless separately there are other to illustrate in the application's context, technical term used and the conventional sense all had known to those skilled in the art of abridging in the application; Except as otherwise noted, in following embodiment, raw materials used compound is commercially available.
Embodiment:
Detect a method for glucose based on gold nano growth, comprise the steps:
1) be 0.01M, pH in concentration be in the phosphate buffer of 7.4, the glucose solution of one group of certain concentration is mixed with glucose oxidase (GOx) respectively, incubated at room temperature 10min, obtain one group of artemia hatching solution a, the concentration of the glucose solution of certain concentration be respectively 0,0.1,0.3,0.5,0.7,0.9,1.0mM;
2) 5nm golden nanometer particle and gold chloride are joined in above-mentioned each artemia hatching solution respectively, in artemia hatching solution b, the concentration of 5nm golden nanometer particle is 8.3nM, the concentration of gold chloride is 0.16mM, incubation at room temperature 30min, obtain one group of artemia hatching solution b, take artemia hatching solution b photo respectively and read them at the corresponding absorbance in 530nm place with Synergy 2 microplate reader, according to concentration of glucose and corresponding absorbance fit standard curve thereof;
3) be 0.01M, pH in concentration be in the phosphate buffer of 7.2-7.4, the glucose solution sample of concentration to be measured is mixed with glucose oxidase (GOx), incubated at room temperature 10min, add golden nanometer particle and the chlorauric acid solution of 5nm, the concentration of golden nanometer particle and the concentration of gold chloride and step 2) in concentration identical, incubation at room temperature 30min, it is read at the corresponding absorbance in 530nm place with Synergy 2 microplate reader, utilize above-mentioned typical curve, calculate the concentration of glucose solution sample to be measured.
Fig. 1 is the method schematic diagram of the growth detection glucose based on golden nanometer particle.
To be concentration be Fig. 20,0.1,0.3,0.5,0.7,0.9, the testing result colour developing figure of the glucose solution of 1.0mM, show in figure: along with the increase of concentration of glucose, golden nanometer particle is grown up gradually, and solution colour reddens gradually.
Fig. 3 is that the glucose solution of variable concentrations is at A
530nmthe fitting a straight line figure at place, wherein A
530nmread absorbance at 530nm place add gold chloride and 5nm golden nanometer particle room temperature reaction 30min in glucose oxidase and glucose mixed solution after with microplate reader.Show in figure: after growth 30min, A
530nmlinear with the concentration of glucose.
The described method detecting glucose based on gold nano growth, for heterogeneous experiment, step is as follows:
1) with card punch, filter paper is broken into the filter paper that diameter is 1cm;
2) be 0.01M, pH in concentration be in the phosphate buffer of 7.2-7.4, the solution of gold nanoparticles of glucose oxidase (GOx), gold chloride and 5nm is mixed, obtain mixed liquor, in mixed liquor, the concentration of glucose oxidase is 1.0mg/mL, the final concentration of 5nm golden nanometer particle is 8.3nM, the final concentration of gold chloride is 0.16mM, mixed liquor is dripped on above-mentioned filter paper, incubated at room 10min;
3) on the above-mentioned scraps of paper, drip the glucose solution of one group of certain concentration, incubated at room 5-60min, glucose oxidase enzymatic glucose produces hydrogen peroxide, hydrogen-peroxide reduction gold chloride, golden nanometer particle is impelled to grow, filter paper reddens gradually, and the red weight of the scraps of paper becomes positive correlation with glucose concentration.
4) by step 2) in mixed liquor drip on filter paper, incubated at room 2-10min, drips the glucose solution of concentration to be measured, incubated at room 5-60min, with step 3) in gained filter paper carry out color contrast, the concentration of qualitative detection glucose solution.
Fig. 4 is the result colour developing figure of method on filter paper of the growth detection glucose based on golden nanometer particle, shows: along with the increase of concentration of glucose, golden nanometer particle is grown up gradually, and the scraps of paper redden gradually in figure.
Claims (7)
1. detect a method for glucose based on gold nano growth, it is characterized in that comprising the steps:
1) be 0.01M, pH in concentration be mixed with glucose oxidase (GOx) respectively by the glucose solution of one group of certain concentration in the phosphate buffer of 7.2-7.4, incubated at room temperature 2-10min, obtains one group of artemia hatching solution a;
2) 5nm golden nanometer particle and gold chloride are joined in above-mentioned each artemia hatching solution respectively, incubation at room temperature 5-60min, obtain one group of artemia hatching solution b, take artemia hatching solution b photo respectively and read them at the corresponding absorbance in 530nm place with Synergy 2 microplate reader, according to concentration of glucose and corresponding absorbance fit standard curve thereof;
3) be 0.01M, pH in concentration be in the phosphate buffer of 7.2-7.4, the glucose solution sample of concentration to be measured is mixed with glucose oxidase (GOx), incubated at room temperature 2-10min, add golden nanometer particle and the chlorauric acid solution of 5nm, incubation at room temperature 5-60min, it is read at the corresponding absorbance in 530nm place with Synergy 2 microplate reader, utilize above-mentioned typical curve, calculate the concentration of glucose solution sample to be measured.
2. detecting the method for glucose based on gold nano growth according to claim 1, it is characterized in that: described step 1) concentration of glucose oxidase is 1.0-3.0mg/mL in artemia hatching solution; The concentration of the glucose solution of certain concentration is respectively 0,0.1,0.3,0.5,0.7,0.9,1.0mM.
3. detecting the method for glucose based on gold nano growth according to claim 1, it is characterized in that: described step 2) concentration of 5nm golden nanometer particle is 4-10nM in artemia hatching solution b, the concentration of gold chloride is 0.05-0.4mM.
4. detect the method for glucose based on gold nano growth according to claim 1, it is characterized in that: described step 3) in the concentration of golden nanometer particle and the concentration of gold chloride and step 2) in concentration identical.
5. detect a method for glucose as claimed in claim 1 based on gold nano growth, it is characterized in that step is as follows for heterogeneous experiment:
1) with card punch, filter paper is broken into the filter paper that diameter is 1cm;
2) be 0.01M, pH in concentration be in the phosphate buffer of 7.3, the solution of gold nanoparticles of glucose oxidase (GOx), gold chloride and 5nm is mixed, obtain mixed liquor, mixed liquor is dripped on above-mentioned filter paper, incubated at room 2-10min;
3) on the above-mentioned scraps of paper, drip the glucose solution of one group of certain concentration, incubated at room 5-60min, glucose oxidase enzymatic glucose produces hydrogen peroxide, hydrogen-peroxide reduction gold chloride, golden nanometer particle is impelled to grow, filter paper reddens gradually, and the red weight of the scraps of paper becomes positive correlation with glucose concentration.
4) by step 2) in mixed liquor drip on filter paper, incubated at room 2-10min, drips the glucose solution of concentration to be measured, incubated at room 5-60min, with step 3) in gained filter paper carry out color contrast, the concentration of qualitative detection glucose solution.
6. the method for glucose is detected according to claim 5 based on gold nano growth, it is characterized in that: described step 2) concentration of glucose oxidase is 1.0-3.0mg/mL in mixed liquor, the final concentration of 5nm golden nanometer particle is 4-10nM, and the final concentration of gold chloride is 0.05-0.4mM.
7. detect the method for glucose based on gold nano growth according to claim 5, it is characterized in that: described step 3) in the concentration of glucose solution of certain concentration be respectively 0,1,10,20,50,100mM.
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN105296595A (en) * | 2015-10-29 | 2016-02-03 | 北京物资学院 | Bio-enzyme activity detection method based on nanogold growth |
| CN107561074A (en) * | 2017-09-27 | 2018-01-09 | 安徽师范大学 | A kind of biological colorimetric sensor, preparation method and its application for detecting glucose |
| CN110672544A (en) * | 2019-10-12 | 2020-01-10 | 天津科技大学 | Glucose visualization sensor based on cyclic peptide recognition element and preparation method and application thereof |
| WO2020109986A1 (en) * | 2018-11-27 | 2020-06-04 | Module Innovations Private Limited | Method for testing of antibiotic susceptibility in microorganisms |
| CN114295819A (en) * | 2021-12-30 | 2022-04-08 | 南京邮电大学 | Label-free high-flux quantitative detection method for glucose |
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| CN1912622A (en) * | 2006-08-10 | 2007-02-14 | 福建省洪诚生物药业有限公司 | Chemiluminescence investigating method of glucose in body fluid |
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| CN1912622A (en) * | 2006-08-10 | 2007-02-14 | 福建省洪诚生物药业有限公司 | Chemiluminescence investigating method of glucose in body fluid |
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Cited By (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105296595A (en) * | 2015-10-29 | 2016-02-03 | 北京物资学院 | Bio-enzyme activity detection method based on nanogold growth |
| CN105296595B (en) * | 2015-10-29 | 2019-04-09 | 北京物资学院 | A kind of bioenzyme activity detection method based on nanogold growth |
| CN107561074A (en) * | 2017-09-27 | 2018-01-09 | 安徽师范大学 | A kind of biological colorimetric sensor, preparation method and its application for detecting glucose |
| WO2020109986A1 (en) * | 2018-11-27 | 2020-06-04 | Module Innovations Private Limited | Method for testing of antibiotic susceptibility in microorganisms |
| GB2594619A (en) * | 2018-11-27 | 2021-11-03 | Module Innovations Private Ltd | Method for testing of antibiotic susceptibility in microorganisms |
| GB2594619B (en) * | 2018-11-27 | 2022-12-07 | Module Innovations Private Ltd | Method for testing of antibiotic susceptibility in microorganisms |
| CN110672544A (en) * | 2019-10-12 | 2020-01-10 | 天津科技大学 | Glucose visualization sensor based on cyclic peptide recognition element and preparation method and application thereof |
| CN114295819A (en) * | 2021-12-30 | 2022-04-08 | 南京邮电大学 | Label-free high-flux quantitative detection method for glucose |
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