CN114751400A - Nitrogen and zinc co-doped graphene quantum dot, ratio type immunosensor, and preparation method and application thereof - Google Patents
Nitrogen and zinc co-doped graphene quantum dot, ratio type immunosensor, and preparation method and application thereof Download PDFInfo
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- CN114751400A CN114751400A CN202210561606.7A CN202210561606A CN114751400A CN 114751400 A CN114751400 A CN 114751400A CN 202210561606 A CN202210561606 A CN 202210561606A CN 114751400 A CN114751400 A CN 114751400A
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Abstract
The invention discloses a nitrogen and zinc co-doped graphene quantum dot, a ratio type immunosensor, and a preparation method and application thereof, wherein the method comprises the following steps: ultrasonically and uniformly mixing GO powder and L-cysteine in water, adjusting the pH value to 8-9, then heating at high temperature for reaction to obtain supernatant, removing water, and freeze-drying to obtain N-doped graphene quantum dot powder; heating the N-doped graphene quantum dot powder, dropwise adding a solution containing zinc ions under stirring for light-shielding reaction, centrifuging to obtain a precipitate, washing, freeze-drying, and preparing a solution with deionized water to obtain the nitrogen-zinc co-doped graphene quantum dot, so that the fluorescence intensity of the quantum dot is improved, and the quantum yield of the quantum dot is also improved. Based on graphene oxide and nitrogen-zinc co-doped graphene quantum dot composite materials, a photoelectric double-response ratio type immunosensor is constructed, and the photoelectric double-response ratio type immunosensor is used for detecting disease markers, obtains good results, and is low in detection limit.
Description
Technical Field
The invention relates to the technical field of novel nano composite materials and biosensing detection, in particular to a nitrogen and zinc co-doped graphene quantum dot, a ratio type immunosensor, and a preparation method and application thereof.
Background
In recent years, the prevalence rate of chronic diseases of the middle-aged and the elderly continues to rise, which seriously affects the health and the quality of life of the elderly, and increases the economic burden of society. Chronic diseases mainly include the following diseases: hypertension, diabetes, cardiovascular and cerebrovascular diseases, etc. Therefore, early detection and diagnosis of the above chronic diseases is one of the current research hotspots.
Graphene Oxide (GO) is an ideal material of an electrochemical biosensor, and the biosensor prepared from the GO has good application prospect in detecting protein, glucose and the like in medicine; GO also has excellent light transmittance, photon transmittance is high at 97.4%, more importantly, Graphene Oxide (GO) has higher specific surface area, and the two-dimensional structure of the graphene oxide allows other substances to exist on the surface of the graphene oxide. Due to these unique attributes, graphene oxide has become a good carrier in biological applications. The graphene quantum dots prepared by taking GO as a raw material have excellent physicochemical characteristics, so that the graphene quantum dots have great application potential in the aspect of biological detection. The fluorescence intensity and the quantum yield of the existing graphene quantum dots are not high;
therefore, it is necessary to develop a graphene quantum dot having fluorescence intensity and quantum yield.
Disclosure of Invention
The invention aims to provide a nitrogen and zinc co-doped graphene quantum dot, a ratio type immunosensor, a preparation method and an application thereof. And a photoelectric double-response ratio type immunosensor is constructed on the basis of the graphene oxide and nitrogen and zinc co-doped graphene quantum dot composite material, and the photoelectric double-response ratio type immunosensor can obtain a good result when being used for detecting disease markers, and the detection limit is low. Compared with an electrochemical sensor and an photochemical sensor, the sensor has higher selectivity and strong stability.
In order to achieve the purpose, the invention adopts the following technical scheme:
in a first aspect of the present invention, a preparation method of a nitrogen and zinc co-doped graphene quantum dot is provided, and the method includes:
ultrasonically mixing GO powder and L-cysteine in water uniformly, adjusting the pH value to 8-9, then carrying out high-temperature heating reaction to obtain supernatant, removing water to obtain light yellow solid, and carrying out freeze drying to obtain N-doped graphene quantum dot powder (N-GQDs);
heating the N-doped graphene quantum dot powder, dropwise adding a solution containing zinc ions under stirring for light-shielding reaction, centrifuging to obtain a precipitate, washing, freeze-drying, and preparing a solution with deionized water to obtain the nitrogen-zinc co-doped graphene quantum dot, namely the N, Zn-GQDs nano composite material.
Further, the mass ratio of the GO powder to the L-cysteine is 1: (1-6).
Further, the temperature of the high-temperature heating reaction is 190-210 ℃, and the time of the high-temperature heating reaction is 3-5 hours.
Further, heating the N-doped graphene quantum dot powder to 35-45 ℃; the temperature of the light-resistant reaction is 25-80 ℃, and the time of the light-resistant reaction is 1-5 h.
Further, the solution containing zinc ions comprises one of zinc acetate, zinc sulfate, zinc chloride, zinc nitrate, zinc fluosilicate, zinc fluoroborate and zinc gluconate; the volume ratio of the N-GQDs to the solution containing zinc ions is 1: (1-10).
In a second aspect of the invention, the nitrogen and zinc co-doped graphene quantum dot obtained by the method is provided.
In a third aspect of the present invention, there is provided a method of preparing a ratiometric immunosensor, the method comprising:
and dropwise adding a turbid liquid of GO to the activated electrode surface, after film formation, dropwise adding the nitrogen and zinc co-doped graphene quantum dots to the GO surface, and drying and then sequentially combining an antibody, a sealant and an antigen to obtain the ratio type immunosensor.
Further, the GO suspension is prepared by dispersing powder in deionized water; the concentration of GO turbid liquid is 0.1-10 mg/mL, and the concentration of N, Zn-GQDs is 0.1-20 mg/mL; and the concentration ratio of GO turbid liquid to N, Zn-GQDs is 1: (1-10).
Further, the antibody comprises one of cardiac troponin, A beta protein, myoglobin, procalcitonin, albumin, total protein, urine beta 2 microglobulin, clusterin and an antibody corresponding to prostate specific antigen; the blocking agent comprises one of Bovine Serum Albumin (BSA), serum, casein, skimmed milk powder and a protein-free compound; the antigen is one of cardiac troponin, Abeta protein, myoglobin, procalcitonin, albumin, total protein, urine beta 2 microglobulin, clusterin and prostate specific antigen.
In a fourth aspect of the invention, a ratiometric immunosensor prepared by the method is provided.
In a fourth aspect of the invention, there is provided the use of the ratiometric immunosensor in the detection of a biomarker in a human serum sample.
One or more technical solutions in the embodiments of the present invention have at least the following technical effects or advantages:
1. the invention provides a nitrogen and zinc co-doped graphene quantum dot and a preparation method thereof, wherein graphene oxide is used as a raw material, nitrogen-doped graphene quantum dots (N-GQDs) are synthesized through a high-temperature hydrothermal method, and then the N-GQDs are synthesized into the nitrogen and zinc co-doped graphene quantum dots through a reaction with zinc acetate, so that the fluorescence intensity of the quantum dots is improved, and the quantum yield of the quantum dots is improved to 31.5%.
2. The invention provides a ratio-type immunosensor and a preparation method thereof, wherein the ratio-type immunosensor with photoelectric double responses is constructed on the basis of graphene oxide and nitrogen-zinc co-doped graphene quantum dot composite materials, and the ratio-type immunosensor is used for detecting disease markers, obtains good results and is low in detection limit. Compared with an electrochemical sensor and a photochemical sensor, the biosensor has higher selectivity and strong stability, and can be further applied to the detection of various disease biomarkers.
3. The invention provides an application of a ratio-type immunosensor in detection of biomarkers in human serum samples, and provides a ratio-type analysis method adopting photoelectric dual-signal response. The method is used for detecting the content of disease biomarkers by analyzing the ratio of photochemical signals and current signals instead of detecting the content of the disease biomarkers only according to one signal, and is gradually applied to the fields of protein detection, virus detection, detection of other bioactive substances and the like. Compared with other analysis technologies, the analysis method has the advantages of simplicity, rapidness, high sensitivity, strong practicability and the like, and provides a wider application prospect for the development and research of the biosensor.
Drawings
In order to more clearly illustrate the technical solutions in the embodiments of the present invention, the drawings needed to be used in the description of the embodiments are briefly introduced below, and it is obvious that the drawings in the following description are some embodiments of the present invention, and it is obvious for those skilled in the art to obtain other drawings based on the drawings without creative efforts.
FIG. 1 is a scanning electron and transmission electron microscope image of GO; wherein FIG. 1A is a scanning electron microscope, and FIG. 1B is a transmission electron microscope.
FIG. 2 is a scanning electron microscope image and a transmission electron microscope image of N, Zn-GQDs; wherein FIG. 2A is a scanning electron microscope, and FIG. 2B is a transmission electron microscope.
FIG. 3 is an XPS plot of N, Zn-GQDs nanocomposites.
FIG. 4 is a fluorescence spectrum and an ultraviolet spectrum of N-GQDs and N, Zn-GQDs; wherein FIG. 4A is a fluorescence spectrum; FIG. 4B is a UV spectrum.
Fig. 5 is a graph showing results of concentration of a cardiac troponin I standard solution at different concentrations in a ratio-type immunosensor, in which fig. 5A is a graph showing an ECL signal, fig. 5B is a graph showing a DPV signal, and fig. 5C is a graph showing a linear relationship between a ratio of an ECL signal to a DPV signal and a concentration of a cardiac troponin I standard solution.
Fig. 6 is a graph showing the results of the concentration of the standard a β protein solution at different concentrations in the ratiometric immunosensor, in which fig. 6A is a graph of ECL signal, fig. 6B is a graph of DPV signal, and fig. 6C is a graph showing the linear relationship between the ratio of ECL signal to DPV signal and the concentration of the standard a β protein solution.
Detailed Description
The present invention will be specifically explained below in conjunction with specific embodiments and examples, and the advantages and various effects of the present invention will be more clearly presented thereby. It will be understood by those skilled in the art that these specific embodiments and examples are for the purpose of illustrating the invention and are not to be construed as limiting the invention.
Throughout the specification, unless otherwise specifically noted, terms used herein should be understood as having meanings as commonly used in the art. Accordingly, unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. If there is a conflict, the present specification will control.
Unless otherwise specifically stated, various raw materials, reagents, instruments, equipment and the like used in the present invention are commercially available or can be obtained by an existing method. The steps S1, S2, and S3 … … in the present invention do not represent a strict order relationship, and the order may be appropriately adjusted as necessary.
In order to solve the technical problems, the general idea of the embodiment of the application is as follows:
according to an exemplary embodiment of the present invention, there is provided a method for preparing a nitrogen-zinc co-doped graphene quantum dot, the method including:
s101, ultrasonically mixing GO powder and L-cysteine in water, adjusting the pH value to 8-9, performing high-temperature heating reaction to obtain supernatant, removing water to obtain light yellow solid, and performing freeze drying to obtain N-doped graphene quantum dot powder (N-GQDs);
in the above-mentioned step S101, the user can select,
the mass ratio of the GO powder to the L-cysteine is 1: (1-6). If the mass ratio is too small, the reaction may not be complete and the like; if the mass ratio is too large, the stability of the prepared sample is adversely affected;
the GO powder is graphene oxide powder, can be directly purchased and can also be prepared in the following mode: accurately measuring 30mL of concentrated sulfuric acid, adding 1g of graphite and 0.8g of sodium nitrate into 30mL of concentrated sulfuric acid, placing the mixture in an ice bath, continuously stirring for 4h, then slowly adding 4g of potassium permanganate into the mixed solution to prevent rapid oxidation-reduction reaction from causing bumping, then stirring for 1h at room temperature, adding 100mL of deionized water into the solution after the solution becomes green, heating for 1h at 37 ℃, transferring the solution into an oil bath, heating to 95 ℃, refluxing for 2h, observing that the color of the reaction solution becomes yellow, and then stopping further reaction by using 25mL of 30 w/v% hydrogen peroxide. And finally, washing the prepared GO for multiple times by using 5% hydrochloric acid and deionized water in sequence, and drying the washed GO for 24 hours in vacuum.
The temperature of the high-temperature heating reaction is 190-210 ℃, and the time of the high-temperature heating reaction is 3-5 h. If the temperature of the high-temperature heating reaction is too low to completely carry out the reaction, and if the temperature is too high to improve the fluorescence yield of the quantum dots, the temperature is preferably 200 ℃.
And S102, heating the N-doped graphene quantum dot powder, dropwise adding a solution containing zinc ions under stirring for light-shielding reaction, centrifuging to obtain a precipitate, washing, freeze-drying, and preparing a solution by using deionized water to obtain the nitrogen-zinc co-doped graphene quantum dot, namely the N, Zn-GQDs nano composite material.
In the step S102, the first step is performed,
heating the N-doped graphene quantum dot powder to 35-45 ℃; firstly heating to 35-45 ℃, and then dropwise adding a zinc ion-containing solution under strong stirring to facilitate doping of zinc ions; preferably 40 deg.c.
The temperature of the light-resistant reaction is 25-80 ℃, and the time of the light-resistant reaction is 1-5 h. If the temperature of the light-shielding reaction is too low, the formation of a nitrogen-zinc covalent bond is not facilitated, and if the temperature is too high, the fluorescence stability of the quantum dots is not facilitated;
the solution containing zinc ions comprises one of zinc acetate, zinc sulfate, zinc chloride, zinc nitrate, zinc fluosilicate, zinc fluoborate and zinc gluconate; the volume ratio of the N-GQDs to the solution containing zinc ions is 1: (1-10). If the volume ratio is too low, successful doping of zinc ions is not facilitated, and if the volume ratio is too high, fluorescence stability of the quantum dots is not facilitated;
according to another exemplary embodiment of the invention, the nitrogen and zinc co-doped graphene quantum dot obtained by the method is provided. According to the method, graphene oxide is used as a raw material, nitrogen-doped graphene quantum dots (N-GQDs) are synthesized through a high-temperature hydrothermal method, and then the N-GQDs are reacted with zinc acetate to synthesize the nitrogen-zinc co-doped graphene quantum dots, so that the fluorescence intensity of the quantum dots is improved, and the quantum yield of the quantum dots is improved.
According to another exemplary embodiment of the present invention, there is provided a method of preparing a ratio-type immunosensor, the method including:
and dropwise adding a turbid liquid of GO to the activated electrode surface, after film formation, dropwise adding the nitrogen and zinc co-doped graphene quantum dots to the GO surface, and drying and then sequentially combining an antibody, a sealant and an antigen to obtain the ratio type immunosensor.
The GO suspension is prepared by dispersing powder in deionized water; the concentration of GO turbid liquid is 0.1-10 mg/mL, and the concentration range is favorable for loading N, Zn-GQDs.
The concentration of the N, Zn-GQDs is 0.1-20 mg/mL; the concentration range is favorable for capturing protein antibodies by N, Zn-GQDs.
And the concentration ratio of the GO turbid liquid to the N, Zn-GQDs is 1: (1-10). The concentration ratio is favorable for the reaction to be complete; if the concentration ratio is too small, the utilization of GO surface space is not facilitated, and if the concentration ratio is too large, the capture of the protein antibody by Zn-GQDs is not facilitated.
The antibody comprises one of cardiac troponin, Abeta protein, myoglobin, procalcitonin, albumin, total protein, urine beta 2 microglobulin, clusterin and an antibody corresponding to prostate specific antigen; the blocking agent comprises one of Bovine Serum Albumin (BSA), serum, casein, skimmed milk powder and a protein-free compound; the antigen is one of cardiac troponin, Abeta protein, myoglobin, procalcitonin, albumin, total protein, urine beta 2 microglobulin, clusterin and prostate specific antigen.
The photoelectric double-response ratio type immunosensor is constructed on the basis of the graphene oxide and nitrogen-zinc co-doped graphene quantum dot composite material, and is used for detecting disease markers, good results are obtained, and the detection limit is low. Compared with an electrochemical sensor and a photochemical sensor, the biosensor has higher selectivity and strong stability, and can be further applied to the detection of various disease biomarkers.
A ratio-type immunosensor, a method for manufacturing the same, and applications thereof will be described in detail with reference to examples, comparative examples, and experimental data.
Example 1
The embodiment of the invention provides a preparation method of a nitrogen and zinc co-doped graphene quantum dot, which comprises the following steps:
1. preparation of Graphene Oxide (GO): accurately measuring 30mL of concentrated sulfuric acid, adding 1g of graphite and 0.8g of sodium nitrate into 30mL of concentrated sulfuric acid, placing the mixture in an ice bath, continuously stirring for 4h, then slowly adding 4g of potassium permanganate into the mixed solution to prevent rapid oxidation-reduction reaction from causing bumping, then stirring for 1h at room temperature, adding 100mL of deionized water into the solution after the solution becomes green, heating for 1h at 37 ℃, transferring the solution into an oil bath, heating to 95 ℃, refluxing for 2h, observing that the color of the reaction solution becomes yellow, and then stopping further reaction by using 25mL of 30 w/v% hydrogen peroxide. And finally, washing the prepared GO for multiple times by using 5% hydrochloric acid and deionized water in sequence, and drying the washed GO for 24 hours in vacuum. The scanning electron microscope image and the transmission electron microscope image of the obtained GO result are shown in the attached figure 1, and the prepared graphene oxide sheet is ground into powder by a mortar for later use.
2. Accurately weighing 15mg of GO powder and 45mg of L-cysteine, mixing, adding 15mL of deionized water, performing ultrasonic treatment for 30min to uniformly disperse the GO powder, and then adding 100 mu L of ammonia water to adjust the pH value of the mixed solution to 8-9. And then, after carrying out ultrasonic dispersion in an ultrasonic cleaning machine for 10min, transferring the obtained product into an autoclave lined with polytetrafluoroethylene, placing the autoclave in a muffle furnace for heating reaction at 200 ℃ for 4h, cooling the obtained product to room temperature, transferring the obtained product into a glass bottle for carrying out dark ultrasonic dispersion for 30min, so that quantum dots attached between unreacted graphene oxide sheet layers can be dispersed into an aqueous solution, filtering out black precipitates, and collecting supernatant. And transferring the supernatant into a round-bottom flask, removing water at 60 ℃ by using a rotary evaporator to obtain a light yellow solid, and freeze-drying the light yellow solid to obtain light yellow N-doped graphene quantum dot powder (N-GQDs).
3. Accurately weighing 1mg of N-GQDs prepared in the step (1) in a glass bottle, heating to 40 ℃, then dropwise and slowly adding 2mL of 0.1g/mL zinc acetate solution under high-speed magnetic stirring, and reacting for 3 hours in a dark place. After the reaction is finished, the mixed solution is placed in a 4mL centrifuge tube, then the mixed solution is centrifuged in a centrifuge at 10000rpm for 14min, the supernatant is discarded, and the precipitate is washed three to four times by deionized water to obtain N, Zn-GQDs.
4. The scanning electron microscope image and the transmission electron microscope image of the N, Zn-GQDs composite material are shown in the attached figure 2, the sediment is prepared into 1mg/mL solution by deionized water after being frozen and dried, the XPS spectrum of the obtained composite material N, Zn-GQDs is shown in the attached figure 3, the fluorescence spectrum and the ultraviolet spectrum are shown in the attached figure 4, and the obtained aqueous solution is stored in a dark place for later use. Indicating that the composite material of N, Zn-GQDs is successfully prepared.
5. And calculating the quantum yield. The quantum yield of the invention is improved to 31.5 percent through calculation. Is calculated by the formula
Wherein,
for quantum yield, I and ISThe areas of the fluorescence peaks of N, Zn-GQDs and the reference substance solution, A and ASIs N, ZAbsorbance, eta and eta of solutions of n-GQDs and reference substancessRefractive indices of N, Zn-GQDs and reference solution, respectively (aqueous solution: 1.33); in this example, I is 0.6; i isS＝0.3；As＝40302.5；A＝1387206.5；η＝1.33；ηs＝1.33。
Example 2
The embodiment of the invention provides a preparation method and application of a cardiac troponin I ratio type immunosensor, which comprises the following steps:
1. 4.5mg of N, Zn-GQDs prepared in the above example 1 was dispersed in 1mg/mL of EDC in PBS (pH 7.4) under high speed stirring, and stirred for 2h to activate the carboxyl groups on the surface of N, Zn-GQDs.
2. Continuously and continuously polishing the glassy carbon electrode by using 0.3 mu m and 0.05 mu m aluminum oxide powder, sequentially immersing the electrode in water and ethanol, ultrasonically cleaning and drying. The washed electrode is activated in 0.1mol/L PBS solution and 50mmol/L potassium ferricyanide solution by cyclic voltammetry until the cyclic voltammetry current is stable. And cleaning the activated electrode in ultrapure water and ethanol again, and drying for later use.
3. And (3) uniformly dropwise adding 5uL of 1mg/mL GO turbid liquid on the surface of the electrode pretreated in the step (2), drying at normal temperature, uniformly dropwise adding 5 mu L N Zn-GQDs to the surface of GO after the GO forms a film on the surface of the electrode, and drying. After the above materials are completely dried, 5 μ L (100 μ g/mL) of cardiac troponin I antibody (anti-cTnI) is dripped on the surface of the materials, and then the materials are incubated for 3h at 37 ℃ so that the amino group on the anti-cTnI and the carboxyl group on the N, Zn-GQDs can form an amido bond. Next, 5. mu.L of BSA (1%) was incubated at 4 ℃ for 2h to block non-specific binding sites, unbound BSA was washed off, air dried, and 5. mu.L (100. mu.g/mL) of cardiac troponin I (cTnI) solution was added dropwise and incubated at 4 ℃ for 12h to be assayed.
4. Determination of the content of cardiac troponin I (cTnI) by a ratiometric biosensor
Drawing a standard curve: the method comprises the following steps of preparing cTnI antigen concentration, sequentially increasing from top to bottom (0.001, 0.02, 0.03, 0.05, 0.08 and 0.10ng/mL), drawing a standard curve according to the ratio of an ECL light signal to a DPV current signal and the cTnI concentration, and determining the concentration of cTnI according to the standard curve, wherein the specific method comprises the following steps:
a series of cTnI antigen solutions (5 mu L) with different concentrations are dripped on the surface of a glassy carbon electrode modified by an antibody to react for 120min, and a DPV signal and an ECL signal are detected by using differential pulse voltammetry and cyclic voltammetry under optimized experimental conditions. DPV detection conditions comprise a scanning range of-0.6-0.8V; potential increment of 4m V; pulse width 0.1 s; the pulse amplitude was 50m V. ECL detection conditions: the applied voltage is-2-0V (Ag/AgC1), the scanning rate is 100mV/s, the photomultiplier is set to 800V, and the amplification stage number is 3.
The results are shown in FIG. 5, which shows that the prepared rate type immunosensor can realize sensitive detection of cTnI, and the detection limit is as low as 4.59 pg/L.
Example 3
The embodiment of the invention provides a preparation method and application of an Abeta ratio type immunosensor, which are characterized by comprising the following steps:
1. the working electrode is pretreated by the same method as the embodiment 2, 5uL of GO suspension of 1mg/mL is uniformly dripped on the surface of the pretreated electrode, the electrode is dried at normal temperature, after GO forms a film on the surface of the electrode, 5 mu L N, Zn-GQDs are uniformly dripped on the surface of GO, and the GO is dried. After the materials are completely dried, 5 mu L (100 mu g/mL) of Abeta amyloid antibody (anti-Abeta) is dripped on the surface of the materials, and then the materials are incubated for 3h at 25 ℃ so that amino on the anti-Abeta and carboxyl on the N, Zn-GQDs can form amido bonds. Next, 5. mu.L of BSA (1%) was incubated at 4 ℃ for 2h to block non-specific binding sites, unbound BSA was washed off, air dried, and 5. mu.L (100. mu.g/mL) of A.beta.antigen solution was added dropwise and incubated at 37 ℃ for 3h to be assayed.
2. Measurement of amyloid beta content (a β) by a ratiometric biosensor.
Drawing a standard curve: the concentration of the antigen prepared was increased from top to bottom (1, 2, 30, 50, 80, 100ng/mL), a standard curve was plotted, and the concentration of A.beta.was measured from the standard curve.
A series of A beta antigen solutions (5 mu L) with different concentrations are dripped on the surface of an antibody modified electrode to react for 120min, and Differential Pulse Voltammetry (DPV) signals and ECL signal intensity are detected under optimized experimental conditions. DPV detection conditions comprise a scanning range of-0.6-0.8V; potential increment of 4m V; pulse width 0.1 s; the pulse amplitude was 50m V. ECL detection conditions: the applied voltage is-2-0V (Ag/AgC1), the scanning rate is 100mV/s, the photomultiplier is set to 800V, and the amplification stage number is 3.
The results are shown in fig. 6, which shows that the ratiometric immunosensor prepared above can detect a β protein in human serum, with a limit of detection as low as 52 pg/L.
Finally, it should also be noted that the terms "comprises," "comprising," or any other variation thereof, are intended to cover a non-exclusive inclusion, such that a process, method, article, or apparatus that comprises a list of elements does not include only those elements but may include other elements not expressly listed or inherent to such process, method, article, or apparatus.
While preferred embodiments of the present invention have been described, additional variations and modifications in those embodiments may occur to those skilled in the art once they learn of the basic inventive concepts. Therefore, it is intended that the appended claims be interpreted as including the preferred embodiment and all changes and modifications that fall within the scope of the invention.
It will be apparent to those skilled in the art that various changes and modifications may be made in the present invention without departing from the spirit and scope of the invention. Thus, if such modifications and variations of the present invention fall within the scope of the claims of the present invention and their equivalents, the present invention is also intended to include such modifications and variations.
Claims (10)
1. A preparation method of a nitrogen and zinc co-doped graphene quantum dot is characterized by comprising the following steps:
ultrasonically mixing GO powder and L-cysteine in water, adjusting the pH value to 8-9, then carrying out high-temperature heating reaction to obtain supernatant, removing water to obtain light yellow solid, and freeze-drying to obtain N-doped graphene quantum dot powder (N-GQDs);
heating the N-doped graphene quantum dot powder, dropwise adding a solution containing zinc ions under stirring for light-shielding reaction, centrifuging to obtain a precipitate, washing, freeze-drying, and preparing a solution with deionized water to obtain the nitrogen-zinc co-doped graphene quantum dot, namely the N, Zn-GQDs nano composite material.
2. The preparation method of the nitrogen-zinc co-doped graphene quantum dot according to claim 1, wherein the mass ratio of the GO powder to the L-cysteine is 1: (1-6).
3. The preparation method of the nitrogen-zinc co-doped graphene quantum dot according to claim 1, wherein the temperature of the high-temperature heating reaction is 190-210 ℃, and the time of the high-temperature heating reaction is 3-5 h.
4. The preparation method of the nitrogen-zinc co-doped graphene quantum dot according to claim 1, wherein the N-doped graphene quantum dot powder is heated to 35-45 ℃; the temperature of the light-resistant reaction is 25-80 ℃, and the time of the light-resistant reaction is 1-5 h.
5. The method for preparing the nitrogen-zinc co-doped graphene quantum dot according to claim 1, wherein the solution containing the zinc ions comprises one of zinc acetate, zinc sulfate, zinc chloride, zinc nitrate, zinc fluorosilicate, zinc fluoroborate and zinc gluconate; the volume ratio of the N-GQDs to the solution containing zinc ions is 1: (1-10).
6. The nitrogen and zinc co-doped graphene quantum dot prepared by the method of any one of claims 1 to 5.
7. A method of making a ratiometric immunosensor, comprising:
and dropwise adding a GO turbid liquid on the surface of the activated electrode, after film formation, dropwise adding the nitrogen and zinc co-doped graphene quantum dots of claim 6 on the surface of GO, drying, and then sequentially combining an antibody, a sealant and an antigen to obtain the ratio type immunosensor.
8. The method of claim 7, wherein the GO suspension is formed by dispersing a powder in ionized water; the concentration of GO turbid liquid is 0.1-10 mg/mL, and the concentration of N, Zn-GQDs is 0.1-20 mg/mL; and the concentration ratio of the GO turbid liquid to the N, Zn-GQDs is 1: (1-10).
9. A ratiometric immunosensor prepared by the method of any one of claims 7 to 8.
10. Use of a ratiometric immunosensor of claim 9 for the detection of a biomarker in a human serum sample.
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| CN113148988A (en) * | 2021-04-14 | 2021-07-23 | 南昌大学 | Preparation method of nitrogen atom doped graphene quantum dots |
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| CN116023939A (en) * | 2022-12-21 | 2023-04-28 | 宁波慈溪生物医学工程研究所 | Metal doped carbon quantum dot, colorimetric sensing solution, colorimetric array sensor, preparation method of colorimetric array sensor and method for detecting biological mercaptan |
| CN116023939B (en) * | 2022-12-21 | 2023-12-19 | 宁波慈溪生物医学工程研究所 | Metal doped carbon quantum dot, colorimetric sensing solution, colorimetric array sensor, preparation method of colorimetric array sensor and method for detecting biological mercaptan |
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Application publication date: 20220715 Assignee: Wuhan geditai Health Technology Co.,Ltd. Assignor: HUBEI University OF TECHNOLOGY Contract record no.: X2023980047145 Denomination of invention: A nitrogen zinc co doped graphene quantum dot, ratio type immunosensor and its preparation method and application Granted publication date: 20230825 License type: Common License Record date: 20231120 |