CN114570936B - Preparation method of glutathione-S-transferase-gold-platinum nanocluster and application of glutathione-S-transferase-gold-platinum nanocluster in aureomycin detection - Google Patents
Preparation method of glutathione-S-transferase-gold-platinum nanocluster and application of glutathione-S-transferase-gold-platinum nanocluster in aureomycin detection Download PDFInfo
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Abstract
The invention relates to the technical field of gold-platinum nanocluster synthesis, in particular to a preparation method of glutathione-S-transferase-gold-platinum nanocluster and application thereof in aureomycin detection, which comprises the following specific steps: and (3) taking glutathione-S-transferase as a template, preparing glutathione-S-transferase-gold-platinum nanocluster stock solution under a specific pH condition, and further performing dialysis and purification. Under 375nm excitation wavelength, gold-platinum nanoclusters exhibit red fluorescence, and have two emission peaks at 470nm and 656nm, respectively; gold-platinum nanoclusters as an effective, environmentally friendly ratio fluorescent probe can be used to detect aureomycin (CTC) in solution. The gold-platinum nanocluster provided by the invention has unique photophysical characteristics, almost no toxicity and excellent biocompatibility, is rapid and simple in detection of aureomycin, has good detection sensitivity, and is an ideal fluorescent nanomaterial applied to the fields of biology and medicine.
Description
Technical Field
The invention relates to the technical field of gold-platinum nanocluster synthesis, in particular to a preparation method of glutathione-S-transferase-gold-platinum nanocluster and application thereof in aureomycin detection.
Background
Glutathione s transferase is a multifunctional enzyme that is widely distributed throughout the body and is capable of binding to the cytoplasm or membrane, playing an important role in the organism. glutathione-S-transferase can catalyze a large amount of chemical reactions such as nucleophilic aromatic substitution, michael addition and the like, can generate various electrophilic substances by catalyzing and reducing endogenous glutathione and exogenous compounds in organisms, and protects cells from chemical substances such as medicines, endogenous peroxide free radicals, toxic metabolites and the like. glutathione-S-transferase exhibits high expression in certain cancer cells compared to normal tissues, playing an important role in the development of drug resistance to chemotherapeutic drugs. Thus, glutathione s transferase is an important biological enzyme that defends against the reducing and toxic electrophiles produced by normal metabolic processes.
The wavelength of the metal nanocluster (the diameter is less than 2 nm) is equivalent to the Fermi wavelength of electrons, and the metal nanocluster has the properties of molecular-like substances, can perform photoinduced fluorescence, and has the advantages of good stability, mild synthesis conditions, low toxicity, high biocompatibility and the like. The bimetallic nanoclusters are more beneficial to improving fluorescence intensity due to the addition of the second metal element. Because the protein has reducing capability and rich binding sites and groups, the protein usually acts on a metal center in the form of a stabilizer and a reducing agent, the protein-protected metal nanocluster shows stronger fluorescence under the action of a proper ligand, the secondary structure and biological function of the protein are not obviously changed, and the structural stability of the protein can ensure that the fluorescence performance of the protein is not quenched due to aggregation.
Aureomycin (CTC) is a broad-spectrum tetracycline antibiotic, which is widely used in animal husbandry because of its ability to effectively inhibit infections with gram-positive and negative bacteria and its low cost, and also can be used as a growth promoter in animal feed; however, due to improper dosage, many side effects are generated, such as water and soil pollution, bacterial drug resistance and the like. The traditional detection method for aureomycin has the limitations of high cost, low sensitivity, poor selectivity, long time consumption and the like, so that development of a rapid, efficient and low-cost detection method is urgently needed. Therefore, the development of new technologies such as fluorescence sensing for detecting CTCs is of great importance.
Disclosure of Invention
The invention aims to solve the defects in the prior art, and provides a preparation method of glutathione-S-transferase-gold-platinum nanoclusters and application thereof in aureomycin detection.
In order to achieve the above purpose, the present invention adopts the following technical scheme:
a preparation method of glutathione-S-transferase-gold platinum nanocluster comprises the following specific steps:
s1, adding chloroauric acid solution into glutathione-S-transferase solution, uniformly mixing, adding chloroplatinic acid solution, and uniformly mixing to obtain solution A;
s2, adding sodium hydroxide solution into the solution A obtained in the step S1, adjusting the pH of the solution to 11-13, and heating the solution under the metal bath condition for 1-9 hours to obtain solution B;
s3, dialyzing the solution B obtained in the step S2 in a phosphate buffer solution for 24 hours by using a dialysis bag with a molecular retention amount of 12-14kDa to remove redundant reactants, recovering the protein to a natural state as much as possible, and storing the product at the temperature of 4 ℃ to obtain the glutathione-S-transferase-gold platinum nanocluster.
Preferably, in the S1, the concentration ratio of chloroauric acid to chloroplatinic acid is 8:1, and the concentration of glutathione-S-transferase is 20mg ml -1 。
Preferably, in S2, sodium hydroxide adjusts the pH of the solution to 11, and the reaction temperature is 80 ℃ for 5 hours.
The invention also provides application of the glutathione-S-transferase-gold-platinum nanocluster obtained by the preparation method of the glutathione-S-transferase-gold-platinum nanocluster in aureomycin detection, wherein the glutathione-S-transferase-gold-platinum nanocluster is diluted by a phosphate buffer solution, aureomycin with different concentrations is added and mixed uniformly, and incubated at room temperature, and under the excitation wavelength condition, the fluorescence intensity of the glutathione-S-transferase-gold-platinum nanocluster is gradually enhanced along with the gradual increase of the aureomycin concentration, so that the ratio type detection is realized.
Preferably, the detection is achieved at 375nm excitation wavelength by incubation for 5 minutes at room temperature.
Compared with the prior art, the invention has the following beneficial effects:
1. the invention takes glutathione-S-transferase as a template, adopts a one-step synthesis method to prepare the glutathione-S-transferase-gold-platinum nanocluster, and has the advantages of unique photophysical characteristics, almost no toxicity, excellent biocompatibility and low cost.
2. The glutathione-S-transferase-gold-platinum nanocluster prepared by the invention has the advantages of rapid and simple method for detecting the aureomycin and good detection sensitivity, and is an ideal fluorescent nanomaterial applied to the fields of biology and medicine.
Drawings
FIG. 1 is a graph showing excitation spectrum and emission spectrum of glutathione-S-transferase-gold-platinum nanoclusters prepared by the present invention;
FIG. 2 is a fluorescence spectrum chart of glutathione-S-transferase-gold-platinum nanoclusters prepared under different pH conditions according to the present invention;
FIG. 3 is a graph of fluorescence spectra of glutathione-S-transferase-gold-platinum nanoclusters prepared from chloroauric acid and chloroplatinic acid in different proportions according to the invention;
FIG. 4 is a fluorescence spectrum chart of glutathione-S-transferase-gold-platinum nanoclusters prepared at different temperatures according to the present invention;
FIG. 5 is a fluorescence spectrum chart of glutathione-S-transferase-gold-platinum nanoclusters prepared at different times according to the present invention;
FIG. 6 is a graph showing fluorescence spectra of glutathione-S-transferase-gold-platinum nanoclusters prepared by glutathione-S-transferase with different concentrations according to the present invention;
FIG. 7 is a transmission electron microscope image and a size distribution diagram of glutathione-S-transferase-gold-platinum nanoclusters after condition optimization according to the present invention;
FIG. 8 is a graph showing fluorescence emission spectra of glutathione-S-transferase-gold-platinum nanocluster solutions after the addition of different concentrations of aureomycin in the present invention.
Detailed Description
The following technical solutions in the embodiments of the present invention will be clearly and completely described with reference to the accompanying drawings, so that those skilled in the art can better understand the advantages and features of the present invention, and thus the protection scope of the present invention is more clearly defined. The described embodiments of the present invention are intended to be only a few, but not all embodiments of the present invention, and all other embodiments that may be made by one of ordinary skill in the art without inventive faculty are intended to be within the scope of the present invention.
In order to improve the luminous performance of the product, the method comprises the following steps:
adding chloroauric acid solution into glutathione-S-transferase solution, mixing uniformly, adding chloroplatinic acid into the solution, and mixing uniformly; and (3) regulating the pH value of the mixed solution to 11, and heating the mixed solution in a metal bath to obtain the product. The excitation spectrum and the emission spectrum of the product were detected by means of a fluorescence spectrometer. The pH of the mixed solution was changed, and a fluorescence spectrum was observed, and as the pH was lowered, the fluorescence intensity was gradually decreased, and when the pH of the solution was 11, the fluorescence intensity was optimal. Thus, a pH of 11 was chosen as the optimal pH condition for the preparation of glutathione S transferase-gold platinum nanoclusters.
Adding 50 mu L of chloroauric acid solution with the concentration of 10mM into 125 mu L of glutathione-S-transferase solution, uniformly mixing, adding 16 mu L of chloroplatinic acid with the concentration of 10mM into the solution, and uniformly mixing; and (3) regulating the pH value of the mixed solution to 11, and heating the mixed solution in a metal bath to obtain the product. The excitation spectrum and the emission spectrum of the product were detected with a fluorescence spectrometer. Fluorescence spectra were observed by changing the chloroplatinic acid volume to change the concentration ratio of chloroauric acid to chloroplatinic acid, and the fluorescence intensity was optimal when the concentration ratio of chloroauric acid to chloroplatinic acid was 8:1. Thus, a concentration ratio of chloroauric acid to chloroplatinic acid of 8:1 was chosen as the optimal ratio for the preparation of glutathione s transferase-gold platinum nanoclusters.
Adding 50 mu L of chloroauric acid solution with the concentration of 10mM into 125 mu L of glutathione-S-transferase solution, uniformly mixing, adding 6.25 mu L of chloroplatinic acid with the concentration of 10mM into the solution, and uniformly mixing; the pH of the mixed solution was adjusted to 11, and the mixed solution was heated in a metal bath for 1 hour to obtain the product. The excitation spectrum and the emission spectrum of the product were detected by means of a fluorescence spectrometer. The reaction time is increased, the fluorescence spectrum is observed, the fluorescence intensity is gradually enhanced along with the increase of the reaction time, and the fluorescence intensity is optimal when the reaction time is 5 hours. Thus 5 hours was chosen as the optimal time for the preparation of glutathione s-transferase-gold platinum nanoclusters.
Adding 50 mu L of chloroauric acid solution with the concentration of 10mM into 125 mu L of glutathione-S-transferase solution, uniformly mixing, adding 6.25 mu L of chloroplatinic acid with the concentration of 10mM into the solution, and uniformly mixing; the pH of the mixed solution was adjusted to 11, and the mixed solution was heated in a metal bath at 37℃for 5 hours to obtain the product. The excitation spectrum and the emission spectrum of the product were detected by means of a fluorescence spectrometer. The reaction temperature is increased, the fluorescence spectrum is observed, the fluorescence intensity is gradually enhanced along with the increase of the reaction temperature, and the fluorescence intensity is optimal when the reaction temperature is 80 ℃. Thus 80 ℃ was chosen as the optimal temperature for the preparation of glutathione s-transferase-gold-platinum nanoclusters.
mu.L of chloroauric acid solution having a concentration of 10mM was added to 125. Mu.L of 5mg ml -1 Adding 6.25 mu L of chloroplatinic acid with the concentration of 10mM into the solution, and uniformly mixing; the pH of the mixed solution was adjusted to 11, and the mixed solution was heated in a metal bath at 80℃for 5 hours to obtain the product. The excitation spectrum and the emission spectrum of the product were detected by means of a fluorescence spectrometer. Increasing the concentration of glutathione-S-transferase, observing fluorescence spectrum, and increasing the fluorescence intensity with increasing concentration of glutathione-S-transferase, when 20mg ml was used -1 The fluorescence intensity is optimal when glutathione s transferase; thus 20mg ml was chosen -1 glutathione-S-transferase was used as the optimal concentration for the preparation of glutathione-S-transferase-gold-platinum nanoclusters. The glutathione S-transferase-gold platinum nanocluster prepared after the condition optimization has uniform size distribution, and the average particle diameter is about 1.7nm.
Detecting aureomycin by taking glutathione-S-transferase-gold platinum nanoclusters as fluorescent probes:
diluting the glutathione-S-transferase-gold-platinum nanocluster by using a phosphate buffer solution, adding aureomycin with the concentration range of 5-70 mu M, incubating the mixed solution for 5 minutes at room temperature, and gradually enhancing the fluorescence emission of the glutathione-S-transferase-gold-platinum nanocluster at 475nm along with the gradual increase of the aureomycin concentration when the excitation wavelength is 375 nm.
Preparation and optimization of glutathione-S-transferase-gold-platinum nanoclusters
Embodiment one: mu.L of chloroauric acid solution having a concentration of 10mM was added to 125. Mu.L of solution having a concentration of 20mg ml -1 Is dissolved in glutathione S-transferaseMixing the above materials in the solution; then adding 6.25 mu L of chloroplatinic acid with the concentration of 10mM into the solution, and uniformly mixing; regulating the pH value of the mixed solution to 3, heating the mixed solution in a metal bath at 80 ℃ for 5 hours, and dialyzing in a phosphate buffer solution for 24 hours by using a dialysis bag with the molecular retention of 12-14kDa to obtain the glutathione-S-transferase-gold-platinum nanocluster.
Embodiment two: mu.L of chloroauric acid solution having a concentration of 10mM was added to 125. Mu.L of solution having a concentration of 20mg ml -1 Is evenly mixed in the glutathione-S-transferase solution; then adding 6.25 mu L of chloroplatinic acid with the concentration of 10mM into the solution, and uniformly mixing; and (3) regulating the pH value of the mixed solution to 7, heating the mixed solution in a metal bath at 80 ℃ for 5 hours, and dialyzing in a phosphate buffer solution for 24 hours by using a dialysis bag with the molecular retention of 12-14kDa to obtain the glutathione-S-transferase-gold platinum nanocluster.
Embodiment III: mu.L of chloroauric acid solution having a concentration of 10mM was added to 125. Mu.L of solution having a concentration of 20mg ml -1 Is evenly mixed in the glutathione-S-transferase solution; then adding 6.25 mu L of chloroplatinic acid with the concentration of 10mM into the solution, and uniformly mixing; regulating the pH value of the mixed solution to 11, heating the mixed solution in a metal bath at 80 ℃ for 5 hours, and dialyzing in a phosphate buffer solution for 24 hours by using a dialysis bag with the molecular retention of 12-14kDa to obtain the glutathione-S-transferase-gold platinum nanocluster.
Embodiment four: the glutathione-S-transferase-gold-platinum nanoclusters obtained in example one, example two and example three were determined to have the strongest fluorescence emission intensity at pH 11 by comparing their fluorescence intensities at 375nm excitation wavelength as shown in FIG. 2.
Fifth embodiment: mu.L of chloroauric acid solution having a concentration of 10mM was added to 125. Mu.L of solution having a concentration of 20mg ml -1 Is evenly mixed in the glutathione-S-transferase solution; then adding 16 mu L of chloroplatinic acid with the concentration of 10mM into the solution, and uniformly mixing; regulating pH of the mixed solution to 11, heating the mixed solution in metal bath at 80deg.C for 5 hr, dialyzing with dialysis bag with molecular retention of 12-14kDa in phosphate buffer solution for 24 hr to obtain glutathione S-transfer productEnzyme-gold platinum nanoclusters.
Example six: mu.L of chloroauric acid solution having a concentration of 10mM was added to 125. Mu.L of solution having a concentration of 20mg ml -1 Is evenly mixed in the glutathione-S-transferase solution; adding 10 mu L of chloroplatinic acid with the concentration of 10mM into the solution, and uniformly mixing; regulating the pH value of the mixed solution to 11, heating the mixed solution in a metal bath at 80 ℃ for 5 hours, and dialyzing in a phosphate buffer solution for 24 hours by using a dialysis bag with the molecular retention of 12-14kDa to obtain the glutathione-S-transferase-gold platinum nanocluster.
Embodiment seven: the glutathione-S-transferase-gold-platinum nanoclusters obtained in the third, fifth and sixth embodiments are shown in fig. 3, and when the fluorescence intensity of the glutathione-S-transferase-gold-platinum nanoclusters is determined to be 8:1 compared with that of chloroplatinic acid by comparing the fluorescence intensity of the glutathione-S-transferase-gold-platinum nanoclusters at 375nm excitation wavelength.
Example eight: mu.L of chloroauric acid solution having a concentration of 10mM was added to 125. Mu.L of solution having a concentration of 20mg ml -1 Is evenly mixed in the glutathione-S-transferase solution; then adding 6.25 mu L of chloroplatinic acid with the concentration of 10mM into the solution, and uniformly mixing; and (3) regulating the pH value of the mixed solution to 11, heating the mixed solution in a metal bath at 37 ℃ for 5 hours, and dialyzing the mixed solution in a phosphate buffer solution for 24 hours by using a dialysis bag with the molecular retention of 12-14kDa to obtain the glutathione-S-transferase-gold platinum nanocluster.
Example nine: mu.L of chloroauric acid solution having a concentration of 10mM was added to 125. Mu.L of solution having a concentration of 20mg ml -1 Is evenly mixed in the glutathione-S-transferase solution; then adding 6.25 mu L of chloroplatinic acid with the concentration of 10mM into the solution, and uniformly mixing; regulating the pH value of the mixed solution to 11, heating the mixed solution in a metal bath at 50 ℃ for 5 hours, and dialyzing in a phosphate buffer solution for 24 hours by using a dialysis bag with the molecular retention of 12-14kDa to obtain the glutathione-S-transferase-gold platinum nanocluster.
Example ten: the glutathione-S-transferase-gold-platinum nanoclusters obtained in the third, eighth and ninth embodiments are shown in FIG. 4, and the fluorescence emission intensity of the glutathione-S-transferase-gold-platinum nanoclusters is the strongest when the reaction temperature is determined to be 80℃by comparing the fluorescence intensities thereof at 375nm excitation wavelength.
Example eleven: mu.L of chloroauric acid solution having a concentration of 10mM was added to 125. Mu.L of solution having a concentration of 20mg ml -1 Is evenly mixed in the glutathione-S-transferase solution; then adding 6.25 mu L of chloroplatinic acid with the concentration of 10mM into the solution, and uniformly mixing; regulating the pH value of the mixed solution to 11, heating the mixed solution in a metal bath at 80 ℃ for 3 hours, and dialyzing in a phosphate buffer solution for 24 hours by using a dialysis bag with the molecular retention of 12-14kDa to obtain the glutathione-S-transferase-gold platinum nanocluster.
Embodiment twelve: mu.L of chloroauric acid solution having a concentration of 10mM was added to 125. Mu.L of solution having a concentration of 20mg ml -1 Is evenly mixed in the glutathione-S-transferase solution; then adding 6.25 mu L of chloroplatinic acid with the concentration of 10mM into the solution, and uniformly mixing; regulating the pH value of the mixed solution to 11, heating the mixed solution in a metal bath at 80 ℃ for 7 hours, and dialyzing in a phosphate buffer solution for 24 hours by using a dialysis bag with the molecular retention of 12-14kDa to obtain the glutathione-S-transferase-gold platinum nanocluster.
Embodiment thirteen: the glutathione s transferase-gold platinum nanoclusters obtained in the third, eleventh and twelfth examples are shown in fig. 5, and the fluorescence emission intensity is strongest when the reaction time is 5 hours by comparing the respective fluorescence emission intensities at 375nm excitation wavelength.
Fourteen examples: mu.L of chloroauric acid solution having a concentration of 10mM was added to 125. Mu.L of 5mg ml -1 Is evenly mixed in the glutathione-S-transferase solution; then adding 6.25 mu L of chloroplatinic acid with the concentration of 10mM into the solution, and uniformly mixing; regulating the pH value of the mixed solution to 11, heating the mixed solution in a metal bath at 70 ℃ for 5 hours, and dialyzing in a phosphate buffer solution for 24 hours by using a dialysis bag with the molecular retention of 12-14kDa to obtain the glutathione-S-transferase-gold platinum nanocluster.
Example fifteen: mu.L of chloroauric acid solution having a concentration of 10mM was added to 125. Mu.L of solution having a concentration of 10mg ml -1 Is evenly mixed in the glutathione-S-transferase solution; 6.25. Mu.L of chloroplatinic acid at a concentration of 10mM was then introducedAdding acid into the solution, and uniformly mixing; regulating the pH value of the mixed solution to 11, heating the mixed solution in a metal bath at 70 ℃ for 5 hours, and dialyzing in a phosphate buffer solution for 24 hours by using a dialysis bag with the molecular retention of 12-14kDa to obtain the glutathione-S-transferase-gold platinum nanocluster.
Example sixteen: mu.L of chloroauric acid solution having a concentration of 10mM was added to 125. Mu.L of solution having a concentration of 15mg ml -1 Is evenly mixed in the glutathione-S-transferase solution; then adding 6.25 mu L of chloroplatinic acid with the concentration of 10mM into the solution, and uniformly mixing; regulating the pH value of the mixed solution to 11, heating the mixed solution in a metal bath at 70 ℃ for 5 hours, and dialyzing in a phosphate buffer solution for 24 hours by using a dialysis bag with the molecular retention of 12-14kDa to obtain the glutathione-S-transferase-gold platinum nanocluster.
Example seventeenth: glutathione-S-transferase-gold-platinum nanoclusters obtained in example three, fourteen, fifteen and sixteen were compared with each other at 375nm excitation wavelength to determine the concentration of glutathione-S-transferase at 20mg ml by comparing the respective fluorescence emission intensities as shown in FIG. 6 -1 The fluorescence emission intensity is strongest.
(II) glutathione-S-transferase-gold platinum nanocluster as fluorescent probe for detecting aureomycin
Example eighteenth: diluting glutathione-S-transferase-gold-platinum nanocluster stock solution with phosphate buffer solution, then adding aureomycin solution with final concentration of 0-70 mu M, incubating for 10 minutes at room temperature, and detecting fluorescence spectrum under 375nm excitation wavelength with a fluorescence spectrometer.
The description and practice of the invention disclosed herein will be readily apparent to those skilled in the art, and may be modified and adapted in several ways without departing from the principles of the invention. Accordingly, modifications or improvements may be made without departing from the spirit of the invention and are also to be considered within the scope of the invention.
Claims (2)
1. The preparation method of the glutathione-S-transferase-gold-platinum nanocluster is characterized by comprising the following specific steps:
s1, adding chloroauric acid solution into glutathione-S-transferase solution, uniformly mixing, adding chloroplatinic acid solution, and uniformly mixing to obtain solution A;
s2, adding a sodium hydroxide solution into the solution A obtained in the step S1, adjusting the pH of the solution to 11, and heating the solution for 5 hours under the metal bath condition of 80 ℃ to obtain a solution B;
s3, dialyzing the solution B obtained in the step S2 in a phosphate buffer solution for 24 hours by using a dialysis bag with a molecular retention amount of 12-14kDa to remove redundant reactants, recovering the protein to a natural state as much as possible, and storing the product at the temperature of 4 ℃ to obtain glutathione-S-gold-platinum nanoclusters;
in the S1, the concentration ratio of chloroauric acid to chloroplatinic acid is 8:1, and the concentration of glutathione-S-transferase is 20mg ml -1 。
2. The application of the glutathione-S-transferase-gold-platinum nanocluster obtained by the preparation method of the glutathione-S-transferase-gold-platinum nanocluster according to claim 1 in aureomycin detection is characterized in that the glutathione-S-transferase-gold-platinum nanocluster is diluted by a phosphate buffer solution, aureomycin with different concentrations is added and mixed uniformly, and incubated for 5 minutes at room temperature, and under the excitation wavelength condition of 375 and nm, the fluorescence intensity of the glutathione-S-transferase-gold-platinum nanocluster is gradually enhanced along with the gradual increase of the aureomycin concentration, so that the detection is realized.
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| DE102010033924A1 (en) * | 2010-08-03 | 2012-02-09 | Fraunhofer-Gesellschaft zur Förderung der angewandten Forschung e.V. | Process for the preparation of nanoparticles from a noble metal and the use of the nanoparticles thus produced |
| TWI530674B (en) * | 2014-11-06 | 2016-04-21 | 財團法人工業技術研究院 | Gold nanocluster composition and method for preparing the same and method for detecting thiol-containing compounds |
| CN107118758B (en) * | 2017-05-03 | 2019-03-26 | 吉林大学 | A kind of gold/platinum bimetal nano cluster fluorescence probe based on polyethyleneimine protection and its application in detection aureomycin |
| WO2020147753A1 (en) * | 2019-01-15 | 2020-07-23 | 南通纺织丝绸产业技术研究院 | Preparation of metal nanocluster wrapped with sericin protein and fluorescence probe |
| CN112175606B (en) * | 2020-10-08 | 2021-08-27 | 南通大学 | Preparation method of gold-silver nanocluster protected by glutathione-S-transferase and application of gold-silver nanocluster in oxytetracycline detection |
| CN113390843A (en) * | 2021-06-16 | 2021-09-14 | 南通大学 | Preparation method of casein-gold nanocluster and application of casein-gold nanocluster in aureomycin detection |
| CN113695585B (en) * | 2021-08-23 | 2023-07-28 | 南通大学 | Preparation method of casein-protected gold and silver nanoclusters and application of casein-protected gold and silver nanoclusters in aureomycin detection |
-
2022
- 2022-03-02 CN CN202210196368.4A patent/CN114570936B/en active Active
Non-Patent Citations (1)
| Title |
|---|
| Yancai Gao等.Glutathione protected bimetallic gold-platinum nanoclusters with near-infrared emission for ratiometric determination of silver ions.《Microchimica Acta》.2021,第1-8页. * |
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