CN114702950A - Preparation method of fluorescent gold nanocluster material, product and application thereof - Google Patents

Preparation method of fluorescent gold nanocluster material, product and application thereof Download PDF

Info

Publication number
CN114702950A
CN114702950A CN202210423826.3A CN202210423826A CN114702950A CN 114702950 A CN114702950 A CN 114702950A CN 202210423826 A CN202210423826 A CN 202210423826A CN 114702950 A CN114702950 A CN 114702950A
Authority
CN
China
Prior art keywords
acp
solution
gold nanocluster
bsa
auncs
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.)
Pending
Application number
CN202210423826.3A
Other languages
Chinese (zh)
Inventor
祁文静
吴狄
杜呈培
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Chongqing Normal University
Original Assignee
Chongqing Normal University
Priority date (The priority date 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 date listed.)
Filing date
Publication date
Application filed by Chongqing Normal University filed Critical Chongqing Normal University
Priority to CN202210423826.3A priority Critical patent/CN114702950A/en
Publication of CN114702950A publication Critical patent/CN114702950A/en
Pending legal-status Critical Current

Links

Images

Classifications

    • C—CHEMISTRY; METALLURGY
    • C09—DYES; PAINTS; POLISHES; NATURAL RESINS; ADHESIVES; COMPOSITIONS NOT OTHERWISE PROVIDED FOR; APPLICATIONS OF MATERIALS NOT OTHERWISE PROVIDED FOR
    • C09K—MATERIALS FOR MISCELLANEOUS APPLICATIONS, NOT PROVIDED FOR ELSEWHERE
    • C09K11/00—Luminescent, e.g. electroluminescent, chemiluminescent materials
    • C09K11/02—Use of particular materials as binders, particle coatings or suspension media therefor
    • C09K11/025—Use of particular materials as binders, particle coatings or suspension media therefor non-luminescent particle coatings or suspension media
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B22—CASTING; POWDER METALLURGY
    • B22F—WORKING METALLIC POWDER; MANUFACTURE OF ARTICLES FROM METALLIC POWDER; MAKING METALLIC POWDER; APPARATUS OR DEVICES SPECIALLY ADAPTED FOR METALLIC POWDER
    • B22F9/00—Making metallic powder or suspensions thereof
    • B22F9/16—Making metallic powder or suspensions thereof using chemical processes
    • B22F9/18—Making metallic powder or suspensions thereof using chemical processes with reduction of metal compounds
    • B22F9/24—Making metallic powder or suspensions thereof using chemical processes with reduction of metal compounds starting from liquid metal compounds, e.g. solutions
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B82—NANOTECHNOLOGY
    • B82Y—SPECIFIC USES OR APPLICATIONS OF NANOSTRUCTURES; MEASUREMENT OR ANALYSIS OF NANOSTRUCTURES; MANUFACTURE OR TREATMENT OF NANOSTRUCTURES
    • B82Y20/00—Nanooptics, e.g. quantum optics or photonic crystals
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B82—NANOTECHNOLOGY
    • B82Y—SPECIFIC USES OR APPLICATIONS OF NANOSTRUCTURES; MEASUREMENT OR ANALYSIS OF NANOSTRUCTURES; MANUFACTURE OR TREATMENT OF NANOSTRUCTURES
    • B82Y30/00—Nanotechnology for materials or surface science, e.g. nanocomposites
    • C—CHEMISTRY; METALLURGY
    • C09—DYES; PAINTS; POLISHES; NATURAL RESINS; ADHESIVES; COMPOSITIONS NOT OTHERWISE PROVIDED FOR; APPLICATIONS OF MATERIALS NOT OTHERWISE PROVIDED FOR
    • C09K—MATERIALS FOR MISCELLANEOUS APPLICATIONS, NOT PROVIDED FOR ELSEWHERE
    • C09K11/00—Luminescent, e.g. electroluminescent, chemiluminescent materials
    • C09K11/08—Luminescent, e.g. electroluminescent, chemiluminescent materials containing inorganic luminescent materials
    • C09K11/58—Luminescent, e.g. electroluminescent, chemiluminescent materials containing inorganic luminescent materials containing copper, silver or gold
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/64—Fluorescence; Phosphorescence
    • G01N21/6428—Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes"
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/64—Fluorescence; Phosphorescence
    • G01N21/6428—Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes"
    • G01N2021/6432—Quenching
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/64—Fluorescence; Phosphorescence
    • G01N21/6428—Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes"
    • G01N2021/6439—Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes" with indicators, stains, dyes, tags, labels, marks

Landscapes

  • Chemical & Material Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Nanotechnology (AREA)
  • Physics & Mathematics (AREA)
  • Health & Medical Sciences (AREA)
  • Materials Engineering (AREA)
  • Organic Chemistry (AREA)
  • Optics & Photonics (AREA)
  • General Physics & Mathematics (AREA)
  • Crystallography & Structural Chemistry (AREA)
  • Immunology (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • General Chemical & Material Sciences (AREA)
  • Biophysics (AREA)
  • Condensed Matter Physics & Semiconductors (AREA)
  • Composite Materials (AREA)
  • Inorganic Chemistry (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Analytical Chemistry (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • Pathology (AREA)
  • Investigating Or Analysing Materials By The Use Of Chemical Reactions (AREA)

Abstract

The invention relates to a preparation method of a fluorescent gold nanocluster material, and a product and application thereof, and belongs to the technical field of acid phosphatase (ACP) detection. The invention discloses a preparation method of a fluorescent gold nanocluster material (BSA-AuNCs) with bovine serum albumin as a reducing agent and a protective agent, wherein the prepared fluorescent gold nanocluster material has peroxidase simulation activity and fluorescence characteristics and can be used for detecting the content of acid phosphatase (ACP), and a sensitive double-quenching double-enhancement ACP detection method is established by using the gold nanocluster material (BSA-AuNCs) for the first time, is high in ACP detection sensitivity, has a linear range of 0.01-2U/L and a detection limit of 0.003U/L, and has higher selectivity on the acid phosphatase (ACP) compared with other enzymes (such as horseradish peroxidase, lysozyme, glucose oxidase and tyrosinase) or metal ions.

Description

Preparation method of fluorescent gold nanocluster material, product and application thereof
Technical Field
The invention belongs to the technical field of acid phosphatase (ACP) detection, and relates to a preparation method of a fluorescent gold nanocluster material, and a product and application thereof.
Background
Acid phosphatase (ACP) is a common enzyme found in mammals and plants that hydrolyzes the phosphoester bond to dissociate the phosphate group from the substrate. The content of acid phosphatase (ACP) in serum is considered to be a major factor in relevant pathological diagnosis, and abnormal expression of ACP can cause some common diseases, such as prostate cancer, liver cirrhosis, hemolytic anemia, chronic nephritis, and the like. Since acid phosphatase (ACP) has important clinical significance, there is a need to establish a simple and sensitive method for detecting ACP. The detection methods of acid phosphatase (ACP) reported so far include electrochemical methods, fluorescence methods, ratiometric fluorescence methods, colorimetric methods, chromatography methods, and the like. Fluorescence detection methods have been widely used for the detection of molecules in vivo due to their advantages of high sensitivity, high selectivity, and simplicity of use.
Peroxidase is an oxidoreductase, which is distributed in body fluids or cells and catalyzes H2O2Directly oxidizing phenolic or amine compounds. Compared with the defect that the activity of natural enzyme is easy to change under the influence of pH and temperature, the mimic enzyme has the advantages of simple structure, stable chemical property, high efficiency, low cost, easy obtainment and the like. With the development of nanotechnology and biotechnology, various nanomaterials have been proved to have activities simulating peroxidase, and can be used as nanoenzymes, such as mesoporous nickel oxide nanoflowers (NiONFs), C-dots/Fe3O4、FeS2Nanoparticles, graphene quantum dot-supported CuO nanoneedles (GQDs-CuO), and the like have been demonstrated to have peroxide-like activity. However, most of these materials require complicated synthetic methods and strict experimental conditions. In contrast, gold nanoclusters having peroxidase-like properties and fluorescence characteristics have been widely used for detection of chemical substances due to their simple and mild synthesis manner. Cerium (Ce)3+) Is one of important Rare Earth Elements (REEs), which is easily ionized from 4f and releases electrons, and has unique catalysis, magnetism and propertyAn electronic characteristic. Deng et al convert Ce3+Adding into gold nanoparticles with peroxidase activity to enhance the catalytic activity of the gold nanoparticles. And Ce4+Is generally used for synthesizing nano enzyme with strong oxidizing property. LucaArtigia et al reported TiO2@ CeOx can catalyze H2O2Adding Ce3+Oxidation to Ce4+、
Figure BDA0003607644550000011
Redox mechanism of (2) and different proportions of Ce3+And Ce4+To TiO 22The catalytic activity of @ CeOx is affected to a different extent, even better than that of the natural peroxidase.
Fluorescence Resonance Energy Transfer (FRET) is widely used as a sensitive analysis method, and provides more possibilities for the development of the analysis method. Shi et al synthesized Fe/Eu-MOF materials with simulated peroxidase activity and fluorescence properties, capable of catalyzing H2O2Oxidizing TMB to generate oxTMB, wherein the oxTMB has stronger ultraviolet absorption at 652nm, so that Fe/Eu-MOF is used as a donor, and the oxTMB is used as an acceptor to construct a FRET mechanism. Ni and his colleagues developed a colorimetric fluorescence two-channel method for the detection of alkaline phosphatase (ALP) based on the FRET mechanism using gold nanoclusters (BSA-AuNCs) with mimicking peroxidase activity and fluorescence. Double quenching is also a good method to reduce background signal, increase sensitivity and selectivity. Zhang and his colleagues developed a fluorescent double quenching method for the selective detection of intracellular SO2To overcome the interference of long term exposure to cysteine.
Therefore, the search for efficient dual "enhanced" detection methods combined with Fluorescence Resonance Energy Transfer (FRET) and dual quenching responses has great potential in improving sensitivity and selectivity.
Disclosure of Invention
In view of the above, an object of the present invention is to provide a method for preparing a fluorescent gold nanocluster material; the second purpose of the invention is to provide a fluorescent gold nanocluster material; the invention also aims to provide an application of the fluorescent gold nanocluster material in acid phosphatase (ACP).
In order to achieve the purpose, the invention provides the following technical scheme:
1. a preparation method of a fluorescent gold nanocluster material comprises the following steps:
chloroauric acid trihydrate (HAuCl)4-3H2O) adding the aqueous solution into a Bovine Serum Albumin (BSA) aqueous solution, mixing, stirring in a water bath at 20-50 ℃, adding an alkali solution to adjust the pH of the solution to 10-11, continuously stirring for 10-18h in a dark place, and dialyzing by using a cutoff membrane and ultrapure water to remove unreacted raw materials to obtain the fluorescent gold nanocluster material ((BSA-AuNCs)).
Preferably, the chloroauric acid trihydrate (HAuCl)4-3H2O) and Bovine Serum Albumin (BSA) in a molar mass ratio of 1:3-8, mol: g.
Preferably, the chloroauric acid trihydrate (HAuCl)4-3H2O) by containing chloroauric acid trihydrate (HAuCl) at a concentration of 8-12mM4-3H2O) is added in the form of an aqueous solution;
the mass concentration of the Bovine Serum Albumin (BSA) in the Bovine Serum Albumin (BSA) aqueous solution is 40-80 mg/mL.
Further preferably, the alkali solution is a sodium hydroxide solution.
Preferably, the molecular weight of the cut-off membrane is 3 KDa.
2. The fluorescent gold nanocluster material prepared according to the preparation method.
3. The fluorescent gold nanocluster material is applied to preparation of an acid phosphatase (ACP) detection probe.
The invention has the beneficial effects that: the invention discloses a preparation method of a fluorescent gold nanocluster material (BSA-AuNCs) by taking bovine serum albumin as a reducing agent and a protective agent, wherein the prepared product has peroxidase simulation activity and fluorescence characteristics and can be used for detecting the content of acid phosphatase (ACP), and a sensitive double-quenching double-enhanced phosphatase (ACP) detection method is established by using the fluorescent gold nanocluster material (BSA-AuNCs) for the first time, and is high in ACP detection sensitivity, the linear range is 0.01-2U/L, the detection limit is 0.003U/L, and the method is relative to other enzymes(such as horseradish peroxidase, lysozyme, glucose oxidase, tyrosinase) or metal ions, and has high selectivity to acid phosphatase (ACP); more importantly, the following advantages are also provided: first, oxTMB is used as a quenching platform, the absorption spectrum of the oxTMB is overlapped with the fluorescence spectrum of BSA-AuNCs, and the BSA-AuNCs realize Fluorescence Resonance Energy Transfer (FRET) from the BSA-AuNCs to the oxTMB and carry out fluorescence quenching firstly. Since the acid phosphatase (ACP) catalyzes the hydrolysis of ascorbic acid 2-phosphate (AA2P) to PO4 3-And Ascorbic Acid (AA), which can reduce oxTMB to TMB and thus prevent FRET from occurring in the presence of ACP, again "enhancing" fluorescence; secondly, Ce is added3+TMB is also oxidized to oxTMB, further quenching the fluorescence; in addition, since PO4 3-And Ce3+The redox reaction is hindered by the presence of ACP and AA 2P. Thus, fluorescence is turned on again in the presence of ACP, and the high quenching and effective enhancement of the initial fluorescent signal enables highly sensitive and selective detection of ACP. Therefore, the fluorescent gold nanocluster material (BSA-AuNCs) prepared by the invention expands the application prospect of a sensitive fluorescence method in biological analysis.
Additional advantages, objects, and features of the invention will be set forth in part in the description which follows and in part will become apparent to those having ordinary skill in the art upon examination of the following or may be learned from practice of the invention. The objectives and other advantages of the invention may be realized and attained by the means of the instrumentalities and combinations particularly pointed out hereinafter.
Drawings
For the purposes of promoting a better understanding of the objects, aspects and advantages of the invention, reference will now be made to the following detailed description taken in conjunction with the accompanying drawings in which:
FIG. 1 is a Transmission Electron Microscope (TEM) image (A) and an ultraviolet absorption spectrum (B) of the fluorescent gold nanocluster material (BSA-AuNCs) prepared in example 1;
FIG. 2 is a graph of a 5mM fluorescent gold nanocluster material (BSA-AuNCs) solution under visible light (a) and ultraviolet radiation (b) at 365 nm;
FIG. 3 is a graph showing an excitation spectrum, an emission spectrum and an oxTMB absorption spectrum of a 5mM gold nanocluster material (BSA-AuNCs) solution;
FIG. 4 shows the fluorescent gold nanocluster material (BSA-AuNCs) prepared in example 1 versus H2O2Effect of oxidation reaction of 3,3',5,5' -Tetramethylbenzidine (TMB), wherein a is color change under visible light condition and B is change of absorption intensity, wherein a is addition of 0.2 mM 3,3',5,5' -Tetramethylbenzidine (TMB) to gold nanocluster material (BSA-AuNCs) solution with concentration of 5mM, and B is addition of 0.4mM hydrogen peroxide (H) to a2O2) C TMB at 0.2mM and H at 0.4mM2O2And d is the addition of 0.2mM TMB and 0.4mM H to c2O2；
FIG. 5 shows the color change (A) and fluorescence intensity change (B) of different solutions under visible light, wherein a is a fluorescent gold nanocluster material (BSA-AuNCs) solution, B is a 0.1mM ascorbic acid 2-phosphoric acid (AA2P) solution added to a, c is 1.4U/L acid phosphatase (ACP) added to a, and d is 0.4mM hydrogen peroxide (H)2O2) 0.25mM of a mixed solution of 3,3',5,5' -Tetramethylbenzidine (TMB) and 0.1mM of ascorbic acid 2-phosphoric acid (AA2P), e is the addition of 50M Ce to d3+F is acid phosphatase (ACP) added to e at a rate of 1.4U/L;
FIG. 6 shows different Ce3+Different solutions (BSA-AuNCs) were mixed with 5mM of a fluorescent gold nanocluster material (BSA-AuNCs) in a HAc-NaAc buffer solution having a pH of 4, and 0.25mM of ascorbic acid 2-phosphate (AA2P) and 0.4mM of hydrogen peroxide (H) were mixed in a HAc-NaAc buffer solution having a pH of 42O2) To a solution of 0.25mM of 3,3',5,5' -Tetramethylbenzidine (TMB) (ACP-free control solution) and a HAc-NaAc buffer solution having a pH of 4, 1.4U/L of acid phosphatase (ACP), 0.25mM of ascorbic acid 2-phosphate (AA2P), and 0.4mM of hydrogen peroxide (H)2O2) 0.25mM of 3,3',5,5' -Tetramethylbenzidine (TMB) (ACP-containing test group solution));
FIG. 7 shows different pH values of the buffer solution versus different solutions (0.25 mM ascorbic acid 2-phosphate (AA2P) and 0.4mM per phosphate in HAc-NaAc buffer solutionHydrogen oxide (H)2O2) 0.25mM of 3,3',5,5' -Tetramethylbenzidine (TMB), 50M of Ce3+The solution formed (control solution without ACP) and HAc-NaAc buffer solution were added with 1.4U/L acid phosphatase (ACP), 0.25mM ascorbic acid 2-phosphate (AA2P), 0.4mM hydrogen peroxide (H)2O2) 0.25mM of 3,3',5,5' -Tetramethylbenzidine (TMB), 50M of Ce3+Solution formed solution (ACP containing experimental group solution));
FIG. 8 shows the addition of 0.25mM ascorbic acid 2-phosphate (AA2P) and 0.4mM hydrogen peroxide (H2) to different solutions (HAc-NaAc buffer solution at pH 4) at different temperatures2O2) 0.25mM of 3,3',5,5' -Tetramethylbenzidine (TMB), 50M of Ce3+To the solution (ACP-free control solution) and the HAc-NaAc buffer solution at pH 4, 1.4U/L of acid phosphatase (ACP), 0.25mM of ascorbic acid 2-phosphate (AA2P), and 0.4mM of hydrogen peroxide (H2) were added2O2) 0.25mM of 3,3',5,5' -Tetramethylbenzidine (TMB), 50M of Ce3+Solution formed solution (ACP containing experimental group solution));
FIG. 9 shows the reaction time for different solutions (HAc-NaAc buffer at pH 4 supplemented with 0.25mM ascorbic acid 2-phosphate (AA2P), 0.4mM hydrogen peroxide (H2)2O2) 0.25mM of 3,3',5,5' -Tetramethylbenzidine (TMB), 50M of Ce3+To the solution (ACP-free control solution) and the HAc-NaAc buffer solution at pH 4, 1.4U/L of acid phosphatase (ACP), 0.25mM of ascorbic acid 2-phosphate (AA2P), and 0.4mM of hydrogen peroxide (H2) were added2O2) 0.25mM of 3,3',5,5' -Tetramethylbenzidine (TMB), 50M of Ce3+Solution formed solution (ACP containing experimental group solution));
in fig. 10, a is an influence of acid phosphatase (ACP) solutions of different concentrations on a fluorescence spectrum of a HAc-NaAc buffer solution (pH 4) of a fluorescent gold nanocluster material (BSA-AuNCs) of which the concentration is 5mM, B is a graph of a relationship between a concentration of an added acid phosphatase (ACP) solution and a fluorescence intensity of the solution, and C is a linear fitting result of a concentration of the added acid phosphatase (ACP) solution and a fluorescence intensity of the solution;
FIG. 11 shows the results of selectivity tests of fluorescent gold nanocluster material (BSA-AuNCs) for ACP. A Blank solution (Blank), alkaline phosphatase (ALP), horseradish peroxidase (HRP), lysozyme (Lys), tyrosine (Try), Glucose Oxidase (GOX), tyrosinase (T yr), cysteine (Cys), glycine (Gly), alanine (Ala), and K were added to a HAc-NaAc buffer solution (pH 4) of a fluorescent gold nanocluster material (BSA-AuNCs) at a concentration of 5mM, respectively+、Na+、Ca2+、Mg2+、Fe2+And the result of fluorescence intensity change after ACP;
FIG. 12 is a diagram of the mechanism of action of the gold nanocluster material (BSA-AuNCs) prepared by the present invention for generating peroxidase mimic activity and fluorescence characteristics.
Detailed Description
The embodiments of the present invention are described below with reference to specific embodiments, and other advantages and effects of the present invention will be easily understood by those skilled in the art from the disclosure of the present specification. The invention is capable of other and different embodiments and of being practiced or of being carried out in various ways, and its several details are capable of modification in various respects, all without departing from the spirit and scope of the present invention. It should be noted that the drawings provided in the following embodiments are only for illustrating the basic idea of the present invention in a schematic way, and the features in the following embodiments and examples may be combined with each other without conflict.
Example 1
A fluorescent gold nanocluster material (BSA-AuNCs) is prepared by the following specific preparation method:
(1) 15mL of chloroauric acid trihydrate (HAuCl)4-3H2Adding O) solution (wherein the concentration of chloroauric acid trihydrate is 10mM) into 15mL of Bovine Serum Albumin (BSA) aqueous solution (wherein the concentration of bovine serum albumin is 50mg/mL), immediately stirring vigorously in a water bath at 37 ℃ for 2min, adding sodium hydroxide (NaOH) solution (wherein the concentration of sodium hydroxide is 1M) to adjust the pH value of the reaction system to 10, stirring continuously in the dark for 12h to obtain gold nanocluster material (BSA-AuNCs) solution with the concentration of 5mM, and storing in 4Keeping in a refrigerator at the temperature of DEG C for later use;
(2) and (2) dialyzing the solution synthesized in the step (1) for 24 hours by using a cut-off membrane with the molecular weight of 3KDa and ultrapure water to obtain the fluorescent gold nanocluster material (BSA-AuNCs).
Example 2
A fluorescent gold nanocluster material (BSA-AuNCs) is prepared by the following specific preparation method:
(1) 15mL of chloroauric acid trihydrate (HAuCl)4-3H2O) solution (wherein the concentration of chloroauric acid trihydrate is 8mM) is added into 9mL of Bovine Serum Albumin (BSA) aqueous solution (wherein the concentration of bovine serum albumin is 40mg/mL), sodium hydroxide (NaOH) solution (wherein the concentration of sodium hydroxide is 1M) is added to adjust the pH value of the reaction system to 10 after the bovine serum albumin aqueous solution is stirred vigorously in a water bath at 37 ℃ for 2min, and the mixture is stirred continuously in a dark place for 12h to obtain gold nanocluster material (BSA-AuNCs) solution with the concentration of 5mM, and the gold nanocluster material solution is stored in a refrigerator at 4 ℃ for later use;
(2) and (3) dialyzing the solution synthesized in the step (1) for 24h by using a 3KDa molecular weight cut-off membrane and ultrapure water to obtain the fluorescent gold nanocluster material (BSA-AuNCs).
Example 3
A fluorescent gold nanocluster material (BSA-AuNCs) is prepared by the following specific preparation method:
(1) 15mL of chloroauric acid trihydrate (HAuCl)4-3H2O) solution (wherein the concentration of chloroauric acid trihydrate is 12mM) is added into 18mL of Bovine Serum Albumin (BSA) aqueous solution (wherein the concentration of bovine serum albumin is 80mg/mL), sodium hydroxide (NaOH) solution (wherein the concentration of sodium hydroxide is 1M) is added to adjust the pH value of the reaction system to be 11 immediately after vigorous stirring in a water bath at 37 ℃ for 2min, and stirring is continuously carried out in a dark place for 12h, so that gold nanocluster material (BSA-AuNCs) solution with the concentration of 5mM is obtained and is stored in a refrigerator at 4 ℃ for later use;
(2) and (3) dialyzing the solution synthesized in the step (1) for 24h by using a 3KDa molecular weight cut-off membrane and ultrapure water to obtain the fluorescent gold nanocluster material (BSA-AuNCs).
Performance testing
The fluorescent gold nanocluster material (BSA-AuNCs) solution used in the following performance test procedure was a fluorescent gold nanocluster material (BSA-AuNCs) solution having a concentration of 5mM, which was prepared by dissolving the fluorescent gold nanocluster material (BSA-AuNCs) solution prepared in example 1 in a HAc-NaAc buffer solution having a pH of 4, and the fluorescent gold nanocluster material (BSA-AuNCs) was the product obtained in step (2) in example 1.
The performance characterization of the fluorescent gold nanocluster material (BSA-AuNCs) prepared in example 1 was performed: fig. 1 a is a diagram of a Transmission Electron Microscope (TEM) showing a morphology characterization of the gold nanocluster material (BSA-AuNCs) prepared in example 1, and it can be seen that the gold nanocluster material (BSA-AuNCs) is successfully synthesized by the preparation method in example 1, and the particle size of the gold nanocluster material is in a range of 3.0-4.5 nm. In addition, when the fluorescent gold nanocluster material (BSA-AuNCs) prepared in example 1 is subjected to ultraviolet absorption detection, as can be seen from an ultraviolet absorption spectrogram (B in FIG. 1), the prepared fluorescent gold nanocluster material (BSA-AuNCs) has no obvious plasmon resonance absorption peak, and only a shoulder peak appears at 275 nm.
Fig. 2 is a graph of a fluorescent gold nanocluster material (BSA-AuNCs) solution with a concentration of 5mM under irradiation of visible light (a) and ultraviolet light (b) at a wavelength of 365nm, from which it can be seen that the fluorescent gold nanocluster material (BSA-AuNCs) solution is yellow under visible light and red fluorescent under irradiation of ultraviolet light at a wavelength of 365nm, indicating that the fluorescent gold nanocluster material (BSA-AuNCs) having a fluorescent characteristic is indeed successfully synthesized in example 1. Fig. 3 shows an excitation spectrum and an emission spectrum of a fluorescent gold nanocluster material (BSA-AuNCs) solution with a concentration of 5mM and an absorption spectrum of oxTMB formed after TMB is oxidized, from which it can be seen that the emission peak wavelength of the gold nanocluster material (BSA-AuNCs) prepared in example 1 is 635nm and the excitation peak wavelength is 370nm, and the emission spectrum of the fluorescent gold nanocluster material and the absorption spectrum of oxTMB are greatly overlapped, which conforms to the principle of Fluorescence Resonance Energy Transfer (FRET), further illustrating that the method in example 1 can indeed prepare the fluorescent gold nanocluster material (BSA-AuNCs) and can also be used for constructing a FRET experimental system together with oxTMB.
FIG. 4 shows the fluorescent gold nanocluster material (BSA-AuNCs) prepared in example 1 versus H2O2Influence of reaction for oxidizing 3,3',5,5' -Tetramethylbenzidine (TMB), whichWhere A is the color change under visible light conditions and B is the change in absorption intensity. As can be seen from a in fig. 4, in the HAc-NaAc buffer solution having a pH of 4 under visible light conditions, the mixed solution (a) of 3,3',5,5' -Tetramethylbenzidine (TMB) having a concentration of 0.2mM and the gold nanocluster material (BSA-AuNCs) having a concentration of 5mM was a colorless transparent solution, and hydrogen peroxide (H) having a concentration of 0.4mM was added2O2) The mixed solution (b) was a colorless transparent solution, and 3,3',5,5' -Tetramethylbenzidine (TMB) was added at a concentration of 0.2mM and hydrogen peroxide (H) was added at a concentration of 0.4mM2O2) The mixed solution (c) formed can be H-substituted with 3,3',5,5' -Tetramethylbenzidine (TMB)2O2OxTMB oxidized to blue appeared pale blue but its blue color was not apparent, but to 3,3',5,5' -Tetramethylbenzidine (TMB) at a concentration of 0.2mM and hydrogen peroxide (H) at a concentration of 0.4mM2O2) The blue color of the mixed solution (d) formed after the fluorescent gold nanocluster material (BSA-AuNCs) with the concentration of 5mM is added into the formed mixed solution (c) is deepened to be dark blue, which shows that the concentration of the blue oxTMB formed in the mixed solution is increased; similarly, as can be seen from B in fig. 4, the two mixed solutions a and B have no distinct absorption peak, and the mixed solution c has a distinct oxTMB absorption peak at 652nm, and the intensity of the oxTMB absorption peak is also significantly increased with the addition of the fluorescent gold nanocluster material (BSA-AuNCs) to the mixed solution d. It can be seen that the fluorescent gold nanocluster material (BSA-AuNCs) prepared by the invention can indeed enhance hydrogen peroxide (H)2O2) The oxidation of 3,3',5,5' -Tetramethylbenzidine (TMB) shows that the fluorescent gold nanocluster material (BSA-AuNCs) prepared by the invention has the effect of enhancing the oxidation of hydrogen peroxide similar to peroxidase.
FIG. 5 shows the color change (A) and fluorescence intensity change (B) of different solutions under visible light, wherein a is a fluorescent gold nanocluster material (BSA-AuNCs) solution, B is a 0.1mM ascorbic acid 2-phosphoric acid (AA2P) solution added to a, c is 1.4U/L acid phosphatase (ACP) added to a, and d is 0.4mM hydrogen peroxide (H)2O2) 0.25mM of a mixed solution of 3,3',5,5' -Tetramethylbenzidine (TMB) and 0.1mM of ascorbic acid 2-phosphoric acid (AA2P), e is the addition of 50M Ce to d3+And f is the addition of 1.4U/L of acid phosphatase (ACP) to e. Since, the acid phosphatase (ACP) can decompose the substrate ascorbic acid 2-phosphate (AA2P) into Ascorbic Acid (AA) and Phosphate (PO)4 3-)，Ce3+With phosphate radical (PO)4 3-) Has high coordination. Therefore, in A of FIG. 5, the mixed solutions d and e have changed the color of the system due to oxidation, while in f, phosphate ions can react with Ce due to generation of phosphate ions3+The coordination effect is generated, and the oxidation of TMB is prevented from changing into blue, so that the blue color is changed into light; in contrast, in B of fig. 5, in the HAc-NaAc buffer solution having a pH of 4, the fluorescence intensity of the solution (a) of the fluorescent gold nanocluster material (BSA-AuNCs) and the solutions formed after ascorbic acid 2-phosphate (AA2P) (B) and acid phosphatase (ACP) (c) were added thereto, respectively, did not change relatively greatly under excitation at 370nm, but hydrogen peroxide (H) was added2O2) The fluorescence intensity of the solution (d) formed by 3,3',5,5' -Tetramethylbenzidine (TMB) and ascorbic acid 2-phosphate (AA2P) was greatly reduced, and 50M Ce was continuously added3+The fluorescence intensity of the solution (e) formed continued to decrease, and then the fluorescence intensity of the solution (f) formed by adding 1.4U/L of acid phosphatase (ACP) was significantly increased. Thus, the influence of oxTMB and other substances on the fluorescence intensity of the fluorescent gold nanocluster material (BSA-AuNCs) at 652nm is shown, and only PO is found through research4 3-When the BSA-AuNCs exist, the fluorescence of the BSA-AuNCs does not change, and meanwhile, because AA is a reducing agent, the blue oxTMB is reduced into colorless TMB solution by the AA, so that the absorption peak of the solution at 652nm disappears, and the fluorescence of the fluorescent gold nanocluster material (BSA-AuNCs) at 652nm is enhanced again. More importantly, PO4 3-These two fluorescence enhancing effects with AA are present simultaneously in the fluorescence "on-on" detection of ACP.
FIG. 6 shows different Ce3+Different solutions (BSA-AuNCs) were mixed with 5mM of a fluorescent gold nanocluster material (BSA-AuNCs) in a HAc-NaAc buffer solution having a pH of 4, and 0.25mM of ascorbic acid 2-phosphate (AA2P) and 0.4mM of hydrogen peroxide (H) were mixed in a HAc-NaAc buffer solution having a pH of 42O2) 0.25mM of 3,3',5,5' -Tetramethylbenzidine (TMB)(control solution without ACP) and HAc-NaAc buffer solution at pH 4 were added with 1.4U/L acid phosphatase (ACP), 0.25mM ascorbic acid 2-phosphate (AA2P), and 0.4mM hydrogen peroxide (H2)2O2) 0.25mM of 3,3',5,5' -Tetramethylbenzidine (TMB) (ACP-containing test group solution)). As can be seen from FIG. 6, the phosphate ester bond of ascorbic acid 2-phosphate (AA2P) was not decomposed in the absence of acid phosphatase (ACP), and Ascorbic Acid (AA) and Phosphate (PO) were not present4 3-) Formation of oxTMB cannot be reduced, and Ce is present3+Is not capable of mixing with phosphate radical (PO)4 3-) Coordination and combination are carried out, the amount of oxTMB is large, energy transfer can occur, and system fluorescence is quenched; when ACP is present, Ascorbic Acid (AA) and Phosphate (PO)4 3-) In addition, the oxTMB is reduced by Ascorbic Acid (AA), energy transfer is inhibited, and Ce3+Can also be mixed with phosphate radical (PO)4 3-) The amount of oxTMB is reduced by coordination bonding, and energy transfer is prevented, thereby also leading to the enhancement of system fluorescence. Ce at a concentration of 20-60M3+Has the effects of fluorescence quenching and quenching in a fluorescent gold nanocluster material (BSA-AuNCs) solution.
FIG. 7 shows different pH values of the buffer solutions (0.25 mM ascorbic acid 2-phosphate (AA2P) and 0.4mM hydrogen peroxide (H) were added to HAc-NaAc buffer solution)2O2) 0.25mM of 3,3',5,5' -Tetramethylbenzidine (TMB), 50M of Ce3+The solution formed (control solution without ACP) and HAc-NaAc buffer solution were added with 1.4U/L acid phosphatase (ACP), 0.25mM ascorbic acid 2-phosphate (AA2P), 0.4mM hydrogen peroxide (H)2O2) 0.25mM of 3,3',5,5' -Tetramethylbenzidine (TMB), 50M of Ce3+Solution formed solution (ACP containing experimental group solution)) fluorescence intensity. The experimental exploration and a large number of literature reports show that the activation of acid phosphatase (ACP) cannot be exerted under neutral or alkaline conditions, and the enzyme activity can be exerted only under acidic conditions. As can be seen from FIG. 7, the addition of acid phosphatase (ACP) can decompose the phosphate ester bond of ascorbic acid 2-phosphate (AA2P) to produce Ascorbic Acid (AA) and Phosphate (PO)4 3-) Since the system fluorescence can be enhanced under acidic conditions of pH 3.6 to 5.6, it is suggested that the acid phosphatase (ACP) can function under pH 3.6 to 5.6, and thus ACP can be detected under pH 3.6 to 5.6.
FIG. 8 shows the addition of 0.25mM ascorbic acid 2-phosphate (AA2P) and 0.4mM hydrogen peroxide (H2) to different solutions (HAc-NaAc buffer solution at pH 4) at different temperatures2O2) 0.25mM of 3,3',5,5' -Tetramethylbenzidine (TMB), 50M of Ce3+To the solution (ACP-free control solution) and HAc-NaAc buffer solution at pH 4 were added 1.4U/L acid phosphatase (ACP), 0.25mM ascorbic acid 2-phosphate (AA2P), and 0.4mM hydrogen peroxide (H)2O2) 0.25mM of 3,3',5,5' -Tetramethylbenzidine (TMB), 50M of Ce3+Solution formed solution (ACP containing experimental group solution)) fluorescence intensity. As can be seen from FIG. 8, when the detection temperature is between 15 ℃ and 50 ℃, the added acid phosphatase (ACP) can decompose the phosphate ester bond of ascorbic acid 2-phosphate (AA2P) to generate Ascorbic Acid (AA) and Phosphate (PO)4 3-) The enhancement of the fluorescence signal is realized, which indicates that the acid phosphatase (ACP) can play a role under the condition of 15-50 ℃, so the detection of the ACP can be carried out under the condition of 15-50 ℃.
FIG. 9 shows the reaction time for different solutions (HAc-NaAc buffer at pH 4 supplemented with 0.25mM ascorbic acid 2-phosphate (AA2P), 0.4mM hydrogen peroxide (H2)2O2) 0.25mM of 3,3',5,5' -Tetramethylbenzidine (TMB), 50M of Ce3+To the solution (ACP-free control solution) and HAc-NaAc buffer solution at pH 4 were added 1.4U/L acid phosphatase (ACP), 0.25mM ascorbic acid 2-phosphate (AA2P), and 0.4mM hydrogen peroxide (H)2O2) 0.25mM of 3,3',5,5' -Tetramethylbenzidine (TMB), 50M of Ce3+Solution formed solution (ACP containing experimental group solution)) fluorescence intensity. As can be seen from FIG. 8, when the reaction time is between 1-60min, the addition of acid phosphatase (ACP) can decompose the phosphate ester bond of ascorbic acid 2-phosphate (AA2P) to generate Ascorbic Acid (AA) and Phosphate (PO)4 3-) Is aA rapid process, with a fluorescence enhanced response at substantially 1 min. The enhancement effect is better along with the prolonging of the time, and a stable fluorescence enhancement effect is basically achieved after 30min, which indicates that the acid phosphatase (ACP) has completely decomposed the phosphate ester bond of ascorbic acid 2-phosphate (AA2P), so that the detection of the ACP is a quick response process, and the detection of the ACP can be realized after 1min of action.
In fig. 10, a is an influence of acid phosphatase (ACP) solutions of different concentrations on a fluorescence spectrum of a HAc-NaAc buffer solution (pH 4) of a fluorescent gold nanocluster material (BSA-AuNCs) of 5mM concentration, B is a graph of a relationship between a concentration of an added acid phosphatase (ACP) solution and a fluorescence intensity of the solution, and C is a linear fitting result of a concentration of the added acid phosphatase (ACP) solution and a fluorescence intensity of the solution. To the HAc-NaAc buffer solution (pH 4) of the gold nanocluster material (BSA-AuNCs) having a concentration of 5mM, acid phosphatase (ACP) solutions having concentrations of 0U/L, 0.01U/L, 0.03U/L, 0.05U/L, 0.08U/L, 0.1U/L, 0.25U/L, 0.4U/L, 0.55U/L, 0.7U/L, 1U/L, 1.2U/L, 1.4U/L, 1.6U/L, 1.8U/L, 2U/L, 3U/L, 5U/L, 7U/L, and 10U/L were added at different concentrations, and the change in fluorescence spectrum was as shown in a in fig. 10, with the increase in the concentration of the added ACP, the intensity of the fluorescence spectrum also gradually increased, and a graph showing the relationship between the concentration of the added ACP solution and the fluorescence intensity of the solution is shown as B in fig. 10. The concentration of the added ACP solution is from 0U/L to 2.0U/L, the fluorescence intensity of the fluorescent gold nano-cluster material (BSA-AuNCs) solution is gradually increased, then the plateau phase is reached, the concentration of the added ACP solution is from 2.0U/L to 10.0U/L, and the fluorescence intensity of the fluorescent gold nano-cluster material (BSA-AuNCs) solution is not changed any more. The concentration of the ACP solution was linearly fitted to the fluorescence intensity of the corresponding fluorescent gold nanocluster material (BSA-AuNCs) solution at a concentration of 2.0U/L to 10.0U/L, and the result is shown in fig. 10C, where the linear equation obtained is I ═ 21.18CACP+17.33c (U/L), correlation coefficient (r) 0.9930, and limit of detection (LOD) 0.003U/L (3, n ═ 11). LOD is calculated from the Standard Deviation (SD) of the response and the slope of the calibration curve. Compared with other ACP detection methods, the double-enhanced fluorescence combined double-quenching strategy has high sensitivity and wide linear range (shown in Table 1).
TABLE 1 detection of ACP Linear Range and detection Limit by different methods
Figure BDA0003607644550000101
FIG. 11 shows the results of a selectivity test of fluorescent gold nanocluster material (BSA-AuNCs) for ACP. A Blank solution (Blank), alkaline phosphatase (ALP), horseradish peroxidase (HRP), lysozyme (Lys), tyrosine (Try), Glucose Oxidase (GOX), tyrosinase (T yr), cysteine (Cys), glycine (Gly), alanine (Ala), and K were added to a HAc-NaAc buffer solution (pH 4) of a fluorescent gold nanocluster material (BSA-AuNCs) at a concentration of 5mM, respectively+、Na+、Ca2+、Mg2+、Fe2+And the result of the change of the fluorescence intensity after the ACP, it can be seen that the fluorescent gold nanocluster material (BSA-AuNCs) prepared in example 1 of the present invention has good fluorescence selectivity to ACP.
Similarly, the fluorescent gold nanocluster materials (BSA-AuNCs) prepared in example 2 and example 3 were used for the above performance test, and the obtained results were similar to those of the fluorescent gold nanocluster materials (BSA-AuNCs) prepared in example 1.
In conclusion, the fluorescent gold nanocluster material (BSA-AuNCs) prepared by the invention has peroxidase mimic activity and fluorescence characteristics. The mechanism of action is shown in fig. 12. The fluorescent gold nanocluster material (BSA-AuNCs) can catalyze colorless TMB to be oxidized into blue oxTMB, and can also be used as a donor to establish FRET with the blue oxTMB so as to cause fluorescence to be quenched, which is the first key factor of fluorescence quenching; when adding Ce3+Then, the fluorescent gold nanocluster material (BSA-AuNCs) catalyzes H2O2Ce is mixed3+Oxidation to Ce4+And Ce4+The material has stronger oxidizability, can further oxidize TMB into oxTMB, and enhances FRET established between gold nanocluster materials (BSA-AuNCs) and blue oxTMB, which is the second key factor of fluorescence quenching. Since ACP hydrolyzes ascorbic acid 2-phosphoric acid (AA2P) into phosphate ion (PO)4 3-) And Ascorbic Acid (AA), first, PO4 3-And Ce3+The high coordination between the two can block the oxidation-reduction reaction; the second reduction of blue oxTMB by AA resulted in its disappearance of absorbance at 652nm, which prevented FRET from the gold nanocluster material (BSA-AuNCs) to oxTMB. Based on this, a method for improving detection of ACP using double quenching and FRET mechanisms can be developed.
The gold nanocluster material (BSA-AuNCs) prepared by the method is used for ACP detection in an actual serum sample, and the specific steps are as follows:
(1) at room temperature (25 ℃), 20L of ascorbic acid 2-phosphate (AA2P) solution with a concentration of 5mM was first reacted with different concentrations of acidic phosphate (ACP) in 200L of HAc-NaAc buffer with a concentration of 0.2M, pH ═ 4 for 30 min;
(2) then 25L of 2mM Ce was added to the sample3+Solution, 20L of 20mM H2O2Solutions and 40L of BSA-AuNCs solution;
(3) finally, 50. mu.L of TMB solution with a concentration of 5mM and an appropriate serum solution diluted 50 times are added continuously to make the final volume of the whole solution 1mL, and the reaction is carried out for 30 min;
(4) the final volume of the whole solution was kept at 1mL by adding HAc-NaAc buffer at a concentration of 0.2M, pH ═ 4, and after thorough vortex mixing, fluorescence spectra were recorded (slit widths for excitation and emission were 10nm and 10nm, respectively, and photomultiplier tube voltage (PMT) was kept at 800V).
In order to verify the accuracy of the fluorescent gold nanocluster material (BSA-AuNCs) prepared by the method in the aspect of actually detecting ACP, a commercially available goat serum sample is diluted and the method for detecting ACP is verified. In other steps, the same as the ACP detection method, ACP of 0.5U/L, ACP of 1.0U/L and ACP of 1.5U/L were added to the goat serum sample, and the content of the ACP and the sample recovery rate obtained by detection are shown in Table 2, and the Relative Standard Deviation (RSD) is less than 5%. The result shows that the method for detecting the ACP by adopting the fluorescent gold nanocluster material (BSA-AuNCs) has higher accuracy and has wide application prospect in the field of actual ACP analysis.
Table 2 test results for ACP detection using fluorescent gold nanocluster material (BSA-AuNCs)
Figure BDA0003607644550000111
In conclusion, the invention discloses a fluorescent gold nanocluster material (BSA-AuNCs) taking bovine serum albumin as a reducing agent and a protective agent, which has peroxidase simulation activity and fluorescence characteristic and can be used for detecting the content of acid phosphatase (ACP), and the fluorescent gold nanocluster material (BSA-AuNCs) is used for establishing a sensitive double-quenching double-enhanced ACP detection method for detecting ACP for the first time, wherein the method has high sensitivity for detecting ACP, the linear range is 0.01-2U/L, the detection limit is 0.003U/L, and the method has higher selectivity for acid phosphatase (ACP) compared with other enzymes (such as horseradish peroxidase, lysozyme, glucose oxidase and tyrosinase) or metal ions; more importantly, the following advantages are also provided: firstly, the oxTMB is used as a quenching platform, the absorption spectrum of the oxTMB is overlapped with the fluorescence spectrum of BSA-AuNCs, and the BSA-AuNCs realizes Fluorescence Resonance Energy Transfer (FRET) from the BSA-AuNCs to the oxTMB, thereby realizing the quenching of fluorescence. Since the acid phosphatase (ACP) catalyzes the hydrolysis of ascorbic acid 2-phosphate (AA2P) to PO4 3-And Ascorbic Acid (AA), which can reduce oxTMB to TMB and thus prevent FRET from occurring in the presence of ACP, again "enhancing" fluorescence; secondly, Ce is added3+TMB is also oxidized to oxTMB, further quenching the fluorescence; in addition, since PO4 3-And Ce3+The redox reaction is hindered by the presence of ACP and AA 2P. Thus, fluorescence is again "enhanced" in the presence of ACP, and the high quenching and efficient enhancement of the initial fluorescent signal enables highly sensitive and selective detection of ACP. Therefore, the fluorescent gold nanocluster material (BSA-AuNCs) provided by the invention expands the application prospect of a sensitive fluorescence method in biological analysis.
Finally, the above embodiments are only intended to illustrate the technical solutions of the present invention and not to limit the present invention, and although the present invention has been described in detail with reference to the preferred embodiments, it will be understood by those skilled in the art that modifications or equivalent substitutions may be made on the technical solutions of the present invention without departing from the spirit and scope of the technical solutions, and all of them should be covered by the claims of the present invention.

Claims (7)

1. A preparation method of a fluorescent gold nanocluster material is characterized by comprising the following steps:
adding chloroauric acid trihydrate aqueous solution into bovine serum albumin aqueous solution, mixing, stirring in a water bath at 20-50 ℃, adding alkali solution to adjust the pH of the solution to 10-11, continuously stirring in a dark place for 10-18h, and removing unreacted raw materials by using a cutoff membrane and ultrapure water dialysis to obtain the fluorescent gold nanocluster material.
2. The method according to claim 1, wherein the molar mass ratio of chloroauric acid trihydrate to bovine serum albumin is 1:3-8, mol: g.
3. The method according to claim 1, wherein the chloroauric acid trihydrate is added in the form of an aqueous solution containing chloroauric acid trihydrate at a concentration of 8 to 12 mM;
the mass concentration of the bovine serum albumin in the bovine serum albumin water solution is 40-80 mg/mL.
4. The production method according to claim 3, wherein the alkali solution is a sodium hydroxide solution.
5. The method according to claim 1, wherein the molecular weight of the cut-off film is 3 kDa.
6. The fluorescent gold nanocluster material prepared by the preparation method of any one of claims 1 to 6.
7. The use of the fluorescent gold nanocluster material of claim 6 in the preparation of an acid phosphatase detection probe.
CN202210423826.3A 2022-04-21 2022-04-21 Preparation method of fluorescent gold nanocluster material, product and application thereof Pending CN114702950A (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
CN202210423826.3A CN114702950A (en) 2022-04-21 2022-04-21 Preparation method of fluorescent gold nanocluster material, product and application thereof

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
CN202210423826.3A CN114702950A (en) 2022-04-21 2022-04-21 Preparation method of fluorescent gold nanocluster material, product and application thereof

Publications (1)

Publication Number Publication Date
CN114702950A true CN114702950A (en) 2022-07-05

Family

ID=82174373

Family Applications (1)

Application Number Title Priority Date Filing Date
CN202210423826.3A Pending CN114702950A (en) 2022-04-21 2022-04-21 Preparation method of fluorescent gold nanocluster material, product and application thereof

Country Status (1)

Country Link
CN (1) CN114702950A (en)

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN116120921A (en) * 2023-01-06 2023-05-16 南方科技大学 Fluorescent-colorimetric bimodal chloride ion probe based on luminescent gold nanoclusters and preparation method and application thereof
CN116660231A (en) * 2023-06-05 2023-08-29 四川大学 Ce-based material 4+ Visualization method for detecting phosphate by using fluorescence-colorimetric dual-mode self-verification as center

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN107389635A (en) * 2017-07-07 2017-11-24 西安科技大学 The synthetic method of functional gold nanoparticles cluster based on bovine serum albumin(BSA) and application
WO2021223317A1 (en) * 2020-05-07 2021-11-11 青岛大学 Preparation method for ratio fluorescent lactic acid probe based on novel nanocomposite system and enzyme catalytic reaction

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN107389635A (en) * 2017-07-07 2017-11-24 西安科技大学 The synthetic method of functional gold nanoparticles cluster based on bovine serum albumin(BSA) and application
WO2021223317A1 (en) * 2020-05-07 2021-11-11 青岛大学 Preparation method for ratio fluorescent lactic acid probe based on novel nanocomposite system and enzyme catalytic reaction

Non-Patent Citations (4)

* Cited by examiner, † Cited by third party
Title
CHENGPEI DU: "Fluorescence dual "turn-on" detection of acid phosphatase via fluorescence resonance energy transfer and dual quenching strategy to improve sensitivity", 《DYES AND PIGMENTS》, pages 1 - 7 *
FANPENG RAN等: "A fluorescent and colorimetric dual-channel sensor based on acid phosphatase-triggered blocking of internal filtration effect", 《MICROCHIMICA ACTA》, vol. 188, no. 8, pages 2, XP037543229, DOI: 10.1007/s00604-021-04951-6 *
JIANPING XIE等: "Protein-Directed Synthesis of Highly Fluorescent Gold Nanoclusters", 《J.AM.CHEM.SOC》, no. 131, pages 3 - 4 *
丛剑涵等: "牛血清蛋白纳米金团簇基于荧光共振能量转移法灵敏检测尿酸", 《光谱学与光谱分析》, vol. 42, no. 2, pages 484 *

Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN116120921A (en) * 2023-01-06 2023-05-16 南方科技大学 Fluorescent-colorimetric bimodal chloride ion probe based on luminescent gold nanoclusters and preparation method and application thereof
CN116120921B (en) * 2023-01-06 2024-01-30 南方科技大学 Fluorescent-colorimetric bimodal chloride ion probe based on luminescent gold nanoclusters and preparation method and application thereof
CN116660231A (en) * 2023-06-05 2023-08-29 四川大学 Ce-based material 4+ Visualization method for detecting phosphate by using fluorescence-colorimetric dual-mode self-verification as center

Similar Documents

Publication Publication Date Title
Chen et al. Logically regulating peroxidase-like activity of gold nanoclusters for sensing phosphate-containing metabolites and alkaline phosphatase activity
CN114702950A (en) Preparation method of fluorescent gold nanocluster material, product and application thereof
Wang et al. Intrinsic enzyme mimicking activity of gold nanoclusters upon visible light triggering and its application for colorimetric trypsin detection
Meng et al. Protein‐protected metal nanoclusters: An emerging ultra‐small nanozyme
Qi et al. PA-Tb-Cu MOF as luminescent nanoenzyme for catalytic assay of hydrogen peroxide
Song et al. Enzyme-triggered in situ formation of Ag nanoparticles with oxidase-mimicking activity for amplified detection of alkaline phosphatase activity
Yi et al. MIL-53 (Fe) MOF-mediated catalytic chemiluminescence for sensitive detection of glucose
Wei et al. Fluorometric determination of pesticides and organophosphates using nanoceria as a phosphatase mimic and an inner filter effect on carbon nanodots
Song et al. A fluorescence turn-on H 2 O 2 probe exhibits lysosome-localized fluorescence signals
Amjadi et al. An enzyme-free fluorescent probe based on carbon dots–MnO2 nanosheets for determination of uric acid
Ma et al. Copper (II) ions enhance the peroxidase-like activity and stability of keratin-capped gold nanoclusters for the colorimetric detection of glucose
Xue et al. Analyte-triggered citrate-stabilized Au nanoparticle aggregation with accelerated peroxidase-mimicking activity for catalysis-based colorimetric sensing of arsenite
CN109266332A (en) A kind of preparation method for the Ratiometric fluorescent probe of AChE and BChE in quantitative detection blood
Yang et al. A promising method for diabetes early diagnosis via sensitive detection of urine glucose by FePd/rGO
Deng et al. A novel strategy for colorimetric detection of hydroxyl radicals based on a modified Griess test
Ahmed et al. Strong nanozymatic activity of thiocyanate capped gold nanoparticles: an enzyme–nanozyme cascade reaction based dual mode ethanol detection in saliva
Yang et al. Continuous singlet oxygen generation for persistent chemiluminescence in Cu-MOFs-based catalytic system
Kim et al. Poly-γ-glutamic Acid/Chitosan hydrogel nanoparticles entrapping glucose oxidase and magnetic nanoparticles for glucose biosensing
Ali et al. One pot fabrication of fluorescein functionalized manganese dioxide for fluorescence “Turn OFF–ON” sensing of hydrogen peroxide in water and cosmetic samples
Chen et al. Rapid and highly sensitive colorimetric biosensor for the detection of glucose and hydrogen peroxide based on nanoporphyrin combined with bromine as a peroxidase-like catalyst
Deng et al. A mitochondrion-targeted dual-site fluorescent probe for the discriminative detection of SO 3 2− and HSO 3− in living HepG-2 cells
Chen et al. Visual and dual-fluorescence homogeneous sensor for the detection of pyrophosphatase in clinical hyperthyroidism samples based on selective recognition of CdTe QDs and coordination polymerization of Ce 3+
Zhang et al. Luminescent carbon dots with excellent peroxidase mimicking property for fluorometric and colorimetric detection of glucose
Lai et al. Enzyme-controllable just-in-time production system of copper hexacyanoferrate nanoparticles with oxidase-mimicking activity for highly sensitive colorimetric immunoassay
Ning et al. Defect-Rich CoFe-Layered Double Hydroxides as Superior Peroxidase-like Nanozymes for the Detection of Ascorbic Acid

Legal Events

Date Code Title Description
PB01 Publication
PB01 Publication
SE01 Entry into force of request for substantive examination
SE01 Entry into force of request for substantive examination