CN111410951A - Composite quantum dot encoding microsphere based on natural spiny pollen and preparation method thereof - Google Patents
Composite quantum dot encoding microsphere based on natural spiny pollen and preparation method thereof Download PDFInfo
- Publication number
- CN111410951A CN111410951A CN202010303050.2A CN202010303050A CN111410951A CN 111410951 A CN111410951 A CN 111410951A CN 202010303050 A CN202010303050 A CN 202010303050A CN 111410951 A CN111410951 A CN 111410951A
- Authority
- CN
- China
- Prior art keywords
- pollen
- quantum dot
- layer
- microsphere
- substrate
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
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
-
- 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
-
- 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
- C09K11/56—Luminescent, e.g. electroluminescent, chemiluminescent materials containing inorganic luminescent materials containing sulfur
- C09K11/562—Chalcogenides
- C09K11/565—Chalcogenides with zinc cadmium
-
- 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
- C09K11/88—Luminescent, e.g. electroluminescent, chemiluminescent materials containing inorganic luminescent materials containing selenium, tellurium or unspecified chalcogen elements
- C09K11/881—Chalcogenides
- C09K11/883—Chalcogenides with zinc or cadmium
Landscapes
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Materials Engineering (AREA)
- Organic Chemistry (AREA)
- Inorganic Chemistry (AREA)
- Investigating, Analyzing Materials By Fluorescence Or Luminescence (AREA)
Abstract
The invention discloses a composite quantum dot coding microsphere based on natural spiny pollen and a preparation method thereof, wherein the preparation method takes the natural spiny pollen as a coding substrate, and quantum dots with different characteristic emission peak positions and intensities are modified layer by layer on the surface; after the spontaneous fluorescence of the pollen substrate is eliminated through carbonization treatment, the porous and thorn-shaped surface structure provides high porosity and large specific surface area, so that the adsorption quantity of target molecules can be increased, the fluorescence signal can be obviously enhanced, and the detection sensitivity can be improved; by adopting a polyelectrolyte layer-by-layer deposition technology, the outer surface of the pollen substrate can adsorb quantum dots with different layers, different types or different sizes layer by layer, the method is simple and easy to implement, can uniformly combine multiple types of quantum dots, and has good coding stability; the combination of different types and different layers of quantum dots can realize the coding form of multiple characteristic emission peaks and intensity, expand the coding amount, and simultaneously can accurately control the fluorescent coding information of the coding microspheres, thereby meeting the requirements of multi-index and high-throughput biological detection.
Description
Technical Field
The invention relates to the technical field of biological materials, relates to a quantum dot coding microsphere taking natural spiny pollen as a coding substrate and a preparation method thereof, and particularly relates to a composite quantum dot coding microsphere based on the natural spiny pollen and a preparation method thereof.
Background
In order to study a large number of intermolecular interactions, such as protein-nucleic acid, protein-protein and protein-drug interactions, high-throughput assay platforms have emerged in succession, which require high-throughput molecular vectors, so-called coding vectors, to identify the different molecules and the interactions occurring between the molecules, followed by analysis of the results by reading the corresponding coding information. In a common coding carrier, an array biochip uses different positions, namely coordinates, where molecules are fixed as codes to distinguish different biomolecules, but the coordinates of the array biochip can only be fixed in a two-dimensional space, so that the speed of identification reaction and application thereof are limited. The microsphere as a novel solid phase coding carrier has become a focus of attention in the fields of high-throughput screening, combinatorial chemistry and the like. Compared with other forms of solid phase carriers, the microspheres have the following remarkable advantages: firstly, the microspheres have large specific surface area, so that the surface chemical reaction can be carried out in a smaller volume; secondly, the microspheres used as the carrier can be combined with other auxiliary means such as stirring, liquid scouring and the like to realize a reaction system between solid-liquid reaction and liquid-liquid reaction, so that the reaction speed of the system is accelerated; thirdly, the molecules bound on the surface of the microsphere can be conveniently separated from the solution after the reaction is finished; fourthly, the surface of the microsphere is modified with diversified functional groups, so that the application of the microsphere can be greatly expanded. The advantages enable the coding microspheres to play a role in the fields of anti-counterfeiting, biological analysis, environmental monitoring and the like.
The existing manufacturing method of the coding microspheres mainly comprises chemical synthesis, a template method, photoetching, a microfluidic technology and the like. The microspheres obtained by these methods tend to exhibit a relatively simple spherical shape and a smooth surface structure. The absence of complex surface morphologies greatly reduces the efficiency of chemical reactions or photoelectric interactions that are carried out at active sites that rely on structural derivatization, and also hinders the integration of a variety of physicochemical properties and functions with the encoding vector. In addition, most of these manufacturing methods involve precision processing techniques, are complicated in equipment, time-consuming, labor-consuming, and costly, and thus limit the mass production and widespread use of encoded microspheres.
The nature provides abundant materials for human beings, the materials are endowed with fine and complex microstructures and extraordinary functions after being evolved and transformed for hundreds of millions of years, and the complicated process of manual preparation is omitted by directly utilizing natural materials. Pollen is a germ cell in the stamen of a flowering plant, is powdery in appearance, is micron-sized in particle size, has a highly monodisperse particle size distribution, has a fine multi-thorn surface structure, and has durable firmness even under harsh natural conditions. In addition, the natural thorny-shaped pollen is huge in quantity and extremely easy to obtain, so that the production cost can be remarkably reduced by using the pollen as a base material of the coding carrier.
Compared with the traditional organic fluorescent molecules, the quantum dots have obvious superiority: the excitation spectrum is wide, the emission spectrum is symmetrically distributed and narrow, the color is adjustable, the photochemical stability is high, and photolysis is not easy to occur. The quantum dots have continuous excitation spectrum, and continuous emission spectrum can be obtained by adjusting the size of crystal particles. The emission spectrum of the quantum dots is narrow, and the quantum dots with various sizes can be simultaneously excited by using light with one wavelength, so that the emission light with various colors can be obtained by using the minimum optical band gap. Based on the coding advantages of the quantum dots, the semiconductor nanocrystal luminescent quantum dots with different fluorescent colors and fluorescent intensities are combined with the natural thorny-shaped pollen substrate, so that an ideal coding microcarrier for the multivariate biological detection can be established.
Therefore, in the invention, natural spiny pollen is used as an encoding carrier, quantum dots are modified on the encoding carrier layer by layer, and the composite quantum dot encoding microsphere capable of simultaneously measuring multiple targets is designed and invented and can be used for biological detection and analysis.
Disclosure of Invention
The invention aims to solve the technical problems of the prior art and provides a composite quantum dot coding microsphere based on natural spiny pollen and a preparation method thereof.
In order to achieve the technical purpose, the technical scheme adopted by the invention is as follows: a preparation method of a composite quantum dot coding microsphere based on natural spiny pollen comprises the following steps:
(1) carrying out dispersive screening on the thorny-shaped pollen, and carrying out carbonization treatment on the screened pollen grains to obtain a pollen substrate after autofluorescence is eliminated;
(2) the method comprises the steps of sequentially soaking a pollen substrate in a negative polyelectrolyte solution and a positive polyelectrolyte solution to obtain a microsphere substrate with a positively charged surface, and adsorbing quantum dots with different layers, different types or different sizes layer by layer on the outer surface of the microsphere substrate with the positively charged surface by utilizing a polyelectrolyte layer-by-layer deposition technology, so as to obtain the composite quantum dot coding microsphere taking natural multi-thorn-shaped pollen as the substrate.
Further, in the step (1), the thorny-shaped pollen is soaked in absolute ethyl alcohol and vibrated to obtain a pollen dispersion liquid with fully stripped pollen grains, the pollen dispersion liquid is dried to disperse and screen out the dry pollen grains with clear grains, the dry pollen grains are calcined at the high temperature of 300 ℃ and continuously introduced with nitrogen for 12 hours to completely carbonize the fluorescent organic substances so as to eliminate autofluorescence.
Further, the thorny pollen is one selected from sunflower pollen, ragweed pollen and chrysanthemum pollen.
Further, the positive polyelectrolyte is selected from one of polyacrylamide hydrochloride (PAH) and Polyethyleneimine (PEI), and the negative polyelectrolyte is selected from one of sodium polystyrene sulfonate (PSS) and polyacrylic acid (PAA).
Further, the preparation method of the positive polyelectrolyte solution or the negative polyelectrolyte solution comprises the step of dissolving the positive polyelectrolyte or the negative polyelectrolyte solid in 0.5 mol/L NaCl solution to finally obtain the positive polyelectrolyte solution or the negative polyelectrolyte solution with the concentration of 1mg/m L.
Further, in the step (2), the positively charged microsphere substrate is soaked in the negatively charged quantum dot solution, the quantum dots are adsorbed on the outer surface of the microsphere substrate, and then the quantum dots with different layers, different types or different sizes are adsorbed layer by the electrostatic acting force between the positive-negative-positive polyelectrolyte layer (formed by sequentially adsorbing the positive-negative-positive polyelectrolyte solution, the negative polyelectrolyte solution and the positive polyelectrolyte solution, and the positive-negative-positive polyelectrolyte layer separates the quantum dots in each layer) and the negatively charged quantum dots.
Further, in the step (2), the quantum dots are selected from one of cadmium sulfide (CdS), cadmium selenide (CdSe), cadmium telluride (CdTe) and zinc sulfide-coated cadmium selenide (CdSe @ ZnS) quantum dots.
Further, in the step (2), a layer of positive polyelectrolyte is deposited outside the quantum dots on the outermost layer to facilitate the subsequent grafting of the detection molecules, wherein the detection molecules are selected from one of DNA, RNA and protein.
The invention also provides a composite quantum dot coding microsphere based on the natural spiny pollen, which is prepared by adopting the preparation method.
The invention also provides a mixed type composite quantum dot coding microsphere based on the natural spiny pollen, the mixed type composite quantum dot coding microsphere is formed by mixing composite quantum dot coding microspheres with different specificities, the composite quantum dot coding microsphere is prepared by adopting the preparation method, the natural spiny pollen is used as a substrate, and quantum dots with different layers, different types or different sizes are adsorbed on the surface layer by layer.
Compared with the prior art, the invention has the beneficial effects that:
1) the natural spiny pollen with good monodispersity is used as the coding substrate, the unique and complex porous and spiny surface structure provides high porosity and large specific surface area, the adsorption quantity of target molecules can be increased, the detection fluorescence signal is obviously enhanced, the detection limit is greatly reduced, and the detection sensitivity is improved; meanwhile, the natural spiny pollen such as sunflower pollen has extremely wide source and huge quantity, and can be used for large-scale mass production and greatly reduce the cost;
2) the method adopts a polyelectrolyte layer-by-layer adsorption method, so that the outer surface of a pollen substrate adsorbs quantum dots with different layers, different types or different sizes layer by layer, the method is simple and easy to implement, and can uniformly combine multiple types of quantum dots;
3) the invention adopts the combination of quantum dots with different types and different layers to realize the coding form of multiple characteristic emission peaks and intensity, greatly expands the coding amount, has simple decoding method for the prepared composite quantum dot coding microsphere, can simultaneously excite the quantum dots with multiple characteristic emission peaks by using light with one wavelength, can simultaneously acquire the type and concentration information of target molecules during decoding, has simple, convenient and quick detection process and no cross interference during detection, and can meet the requirements of simultaneously detecting multiple indexes and high-flux detection;
4) the composite quantum dot coding microsphere prepared by the invention has wide application range, and can realize the fixation of various functional groups in a polyelectrolyte adsorption mode, thereby providing the condition of chemical coupling with downstream molecules.
Drawings
FIG. 1 is a flow chart of a preparation method of the composite quantum dot encoding microsphere based on natural spiny pollen, wherein 1 is the natural spiny pollen, 2 is a pollen substrate after autofluorescence is eliminated, 3 is a quantum dot, and 4 is the prepared composite quantum dot encoding microsphere;
FIG. 2 is a fluorescent representation of natural spiny pollen and a pollen substrate, wherein a is a natural spiny pollen image shot under a laser confocal fluorescence microscope bright field, b-d are autofluorescence images of natural spiny pollen under the excitation of ultraviolet light, blue light and green light in sequence, e is a pollen substrate image shot under the laser confocal fluorescence microscope bright field, and f-h are fluorescence images of the pollen substrate after natural spiny pollen carbonization treatment under the excitation of ultraviolet light, blue light and green light in sequence;
FIG. 3 is an emission spectrum of the sunflower pollen-based composite quantum dot-encoded microsphere of the present invention, wherein FIG. A is an emission spectrum of the sunflower pollen-based composite quantum dot-encoded microsphere prepared in example 1 (the microsphere is encoded by CdTe 465: CdTe 513: CdTe626= 1:2: 3) and its quantum dots corresponding to the number of layers (one CdTe 465, two CdTe layers 513, three CdTe layers 626), a1 is a fluorescence spectrum of the sunflower pollen-based composite quantum dot-encoded microsphere prepared in example 1, B1 is a fluorescence emission spectrum of one CdTe layer of quantum dots, c1 is a fluorescence emission spectrum of two CdTe layers 513, d1 is a fluorescence emission spectrum of three CdTe layers 626 quantum dots, and FIG. B is a composite ragweed pollen-based quantum dot-encoded microsphere prepared in example 2 (the microsphere is encoded by CdSe 460: CdSe620= 1:2: 1) and its quantum dots corresponding to the number of layers (CdSe 460: 525), Two layers of CdSe 525 and one layer of CdSe 620), a2 is the fluorescence emission spectrum of the ragweed pollen-based composite quantum dot-encoded microsphere prepared in example 2, b2 is the fluorescence emission spectrum of one layer of quantum dot CdSe460, c2 is the fluorescence emission spectrum of two layers of quantum dot CdSe 525, and d2 is the fluorescence emission spectrum of one layer of quantum dot CdSe 620;
fig. 4 is a characterization image of a stereomicroscope and a field emission scanning electron microscope of dried sunflower pollen grains and sunflower pollen-based composite quantum dot-encoded microspheres (encoded as CdSe620= 1) of example 4, wherein fig. a is a stereomicroscope image of dried sunflower pollen grains, and fig. b, c are scanning electron microscope images of dried sunflower pollen grains; fig. d is a body microscope image of the sunflower pollen-based composite quantum dot encoded microsphere (encoded as CdSe620= 1), and fig. e and f are scanning electron microscope images of the sunflower pollen-based composite quantum dot encoded microsphere (encoded as CdSe620= 1);
fig. 5 is a fluorescent image of different sections of sunflower pollen-based composite quantum dot-encoded microspheres (encoded as CdSe620= 1) scanned by a laser scanning confocal microscope in example 4;
fig. 6 is a fluorescent characterization image of the sunflower pollen-based composite quantum dot-encoded microsphere (encoded as CdSe460= 1), the sunflower pollen-based composite quantum dot-encoded microsphere (encoded as CdSe 525= 1), the sunflower pollen-based composite quantum dot-encoded microsphere (encoded as CdSe620= 1) under a laser scanning confocal fluorescent microscope in example 4, wherein fig. 6a to c are fluorescent images of the sunflower pollen-based composite blue quantum dot-encoded microsphere (encoded as CdSe460= 1) at different magnifications, fig. 6d to f are fluorescent images of the sunflower pollen-based composite green quantum dot-encoded microsphere (encoded as CdSe 525= 1) at different magnifications, and fig. 6g to i are fluorescent images of the sunflower pollen-based composite red quantum dot-encoded microsphere (encoded as CdSe620= 1) at different magnifications;
fig. 7 is a fluorescent image of sunflower pollen-based hybrid composite quantum dot-encoded microspheres (encoded as CdSe620=1, CdSe 525=1, CdSe460= 1) under simultaneous excitation of an ultraviolet light source in example 4, wherein CdSe620 refers to sunflower pollen-based composite quantum dot-encoded microspheres (encoded as CdSe620= 1), CdSe 525 refers to sunflower pollen-based composite quantum dot-encoded microspheres (encoded as CdSe 525= 1), and CdSe460 refers to sunflower pollen-based composite quantum dot-encoded microspheres (encoded as CdSe460= 1);
fig. 8 is a schematic diagram of a biological detection application process of the composite quantum dot encoding microsphere prepared in example 5, wherein 1 is the prepared composite quantum dot encoding microsphere, 2 is probe DNA, 3 is target DNA, and 4 is labeled DNA for modifying green fluorescent dye carboxylic acid fluorescein (FAM, with a characteristic peak position of 520 nm);
fig. 9 is a graph of the fluorescence intensity of the detected FAM versus the target DNA concentration when the sunflower pollen-based composite quantum dot-encoded microspheres (coded as CdSe460= 1) and the conventional bioassay product Glass Beads (Glass Beads) of example 5 were used for DNA detection, and fig. i and ii are fluorescence images of the sunflower pollen-based composite quantum dot-encoded microspheres (coded as CdSe460= 1) and the conventional bioassay product Glass Beads (Glass Beads) with the surface bound with the hybridization sandwich structure DNA under the modified FAM condition, respectively, at the time of DNA hybridization assay experiment;
fig. 10 is a fluorescence spectrum obtained by using three sunflower pollen-based composite quantum dot-encoded microspheres (CdSe 460: CdSe620= 2:1, CdSe 460: CdSe620= 1:2, respectively) in example 5 for multiplex, simultaneous DNA detection, wherein, a is an emission spectrum of a sunflower pollen-based composite quantum dot-encoded microsphere (CdSe 460: CdSe620= 2: 1) without target signal under the condition of marking unmodified FAM of DNA, and b-d are emission spectra of three sunflower pollen-based composite quantum dot-encoded microspheres (CdSe 460: CdSe620= 2:1, CdSe 460: CdSe620= 1:2, respectively) with hybrid sandwich structure DNA under the condition of combining modified FAM on the surface;
fig. 11 is a fluorescence emission spectrum of the natural sunflower pollen and sunflower pollen-based composite quantum dot-encoded microspheres (encoded as CdSe @ ZnS565= 1) of example 6;
fig. 12 is a graph of fluorescence intensity as a function of time for sunflower pollen-based composite quantum dot-encoded microspheres (encoded as CdSe460= 1) after DNA binding in example 5.
Detailed Description
In order to make those skilled in the art better understand the technical solution of the present invention, the following detailed description of the embodiments of the present invention is provided with reference to the accompanying drawings.
The experimental procedures used in the examples below are, unless otherwise specified, conventional procedures and the reagents, methods and equipment used are, unless otherwise specified, conventional in the art.
The invention provides a preparation method of a composite quantum dot coding microsphere based on natural spiny pollen, which has a process flow as shown in figure 1 and comprises the following steps:
(1) soaking thorny-shaped pollen in absolute ethyl alcohol for oscillation to obtain pollen dispersion liquid with fully stripped pollen grains, drying the pollen dispersion liquid to disperse and screen out dry pollen grains with clear grains, calcining the dry pollen grains at the high temperature of 300 ℃, and continuously introducing nitrogen for 12 hours to completely carbonize fluorescent organic substances so as to eliminate autofluorescence;
(2) the method comprises the steps of sequentially soaking a pollen substrate in a negative polyelectrolyte solution and a positive polyelectrolyte solution to obtain a microsphere substrate with a positively charged surface, and adsorbing quantum dots with different layers, different types or different sizes layer by layer on the outer surface of the microsphere substrate with the positively charged surface by utilizing a polyelectrolyte layer-by-layer deposition technology, so as to obtain the composite quantum dot coding microsphere taking natural multi-thorn-shaped pollen as the substrate.
Wherein the thorny pollen is selected from one of sunflower pollen, ragweed pollen and chrysanthemum pollen.
Wherein the positive polyelectrolyte is selected from one of polyacrylamide hydrochloride (PAH) and Polyethyleneimine (PEI).
Wherein, the negative polyelectrolyte is selected from one of sodium polystyrene sulfonate (PSS) and polyacrylic acid (PAA).
Further, the preparation method of the positive polyelectrolyte solution or the negative polyelectrolyte solution comprises the step of dissolving the positive polyelectrolyte or the negative polyelectrolyte solid in 0.5 mol/L NaCl solution to finally obtain the positive polyelectrolyte solution or the negative polyelectrolyte solution with the concentration of 1mg/m L.
Further, in the step (2), the positively charged microsphere substrate is soaked in the negatively charged quantum dot solution, the quantum dots are adsorbed on the outer surface of the microsphere substrate, and then the polyelectrolyte layers formed by sequentially adsorbing the positive-negative-positive polyelectrolyte layer (the positive polyelectrolyte solution, the negative polyelectrolyte solution and the positive polyelectrolyte solution are separated from each other by the polyelectrolyte layer) and the negatively charged quantum dots are adsorbed by electrostatic acting force between the polyelectrolyte layers and the negatively charged quantum dots, wherein the number of the quantum dots is different, and the quantum dots are different in type or size.
Further, the quantum dots are selected from one of cadmium sulfide (CdS), cadmium selenide (CdSe), cadmium telluride (CdTe), and zinc sulfide-coated cadmium selenide (CdSe @ ZnS) quantum dots.
Further, in the step (2), a layer of positive polyelectrolyte is deposited outside the quantum dots on the outermost layer to facilitate grafting of a subsequent detection molecule, wherein the subsequent detection molecule is selected from one of DNA, RNA and protein.
The preparation method of the natural spiny pollen-based composite quantum dot coding microsphere provided by the invention takes the natural spiny pollen as a coding substrate, has wide material source, can be used for large-scale mass production and greatly reduces the cost, adopts a polyelectrolyte layer-by-layer adsorption method to ensure that the outer surface of the pollen substrate adsorbs quantum dots with different layers, different types or different sizes layer by layer, is simple and feasible, can uniformly combine multiple types of quantum dots, has good coding stability for the prepared composite quantum dot coding microsphere, adopts the combination of the quantum dots with different types and different layers to realize the coding form of multiple characteristic emission peaks and intensity, and greatly expands the coding amount.
The composite quantum dot coding microsphere based on the natural spiny pollen can be prepared by the preparation method, the natural spiny pollen is used as a substrate, and quantum dots with different layers, different types or different sizes are adsorbed on the surface layer by layer.
The preparation method can also be used for preparing the mixed type composite quantum dot coding microspheres based on the natural spiny pollen, the mixed type composite quantum dot coding microspheres are formed by mixing composite coding microspheres with different specificities, the composite coding microspheres take the natural spiny pollen as a substrate, and quantum dots with different layers, different types or different sizes are adsorbed on the surface layer by layer.
The composite quantum dot coding microsphere or the mixed composite quantum dot coding microsphere based on the natural multi-thorn-shaped pollen takes the multi-thorn-shaped pollen as a coding substrate, the porous and thorn-shaped surface structures of the composite quantum dot coding microsphere or the mixed composite quantum dot coding microsphere have fluorescence enhancement benefits, the detection sensitivity is improved, the quantum dots adsorbed on the surface of the pollen substrate layer by layer adopt combinations of quantum dots of different types and different layers, the coding amount is greatly expanded, the coding stability is good, the decoding method of the composite quantum dot coding microsphere is simple, the quantum dots with various characteristic emission peaks can be excited simultaneously only by light with one wavelength, the type and concentration information of target molecules can be obtained simultaneously during decoding, the detection process is simple, convenient and rapid, no cross interference exists during detection, and the requirements of simultaneous detection of multiple indexes and high-flux detection can be met.
The following are examples:
example 1
A sunflower pollen-based composite quantum dot coding microsphere is prepared by the following steps:
(1) method for preparing sunflower pollen substrate without autofluorescence by carbonization
Soaking sunflower pollen in absolute ethyl alcohol, keeping shaking for 20 min to obtain dispersion with fully stripped pollen grains, heating the dispersion on a hot table at 100 deg.C until the ethanol is completely volatilized, and screening out dry sunflower pollen grains with clear grains; placing the dried sunflower pollen granules in a muffle furnace, calcining at a high temperature of 300 ℃ and continuously introducing nitrogen for 12 hours to obtain a positively charged sunflower pollen substrate after the fluorescent organic substance is completely carbonized, as shown in figure 1;
the natural sunflower pollen and the carbonized sunflower pollen substrate are subjected to fluorescence characterization by using a laser scanning confocal fluorescence microscope, and FIG. 2 is the fluorescence characterization of the natural sunflower pollen and the natural sunflower pollen substrate, wherein, the picture a is a natural spiny pollen image shot under a laser confocal fluorescence microscope bright field, the images b-d are autofluorescence images of natural multi-thorn pollen under the excitation of ultraviolet light, blue light and green light in sequence, the picture e is the pollen substrate image shot under the light field of a laser confocal fluorescence microscope, the pictures f-h are the fluorescence images of the pollen substrate after the carbonization treatment of the natural thorny pollen under the excitation of ultraviolet light, blue light and green light in sequence, compared with the fluorescent images of the natural sunflower pollen and the sunflower pollen substrate, the carbonized sunflower pollen substrate has better spine structure retention degree and completely removed autofluorescence; the Zeta potential of the sunflower pollen substrate is measured by using a dynamic light scattering instrument, and the result shows that the prepared sunflower pollen substrate has positive charge on the surface and the potential mean value is +27.1 mV;
(2) sunflower pollen-based composite quantum dot coding microsphere prepared by polyelectrolyte layer-by-layer adsorption method
Respectively dissolving solid particles of PAH and PSS in 0.5 mol/L NaCl solution to prepare PAH solution and PSS solution with the concentration of 1mg/m L, soaking the sunflower pollen substrate without autofluorescence in the PSS solution, standing for adsorption for 20 minutes, centrifuging, washing with ultrapure water for three times to wash off redundant PSS solution to obtain pollen microspheres with negative surface, soaking the pollen microspheres with negative surface in the PAH solution, standing for adsorption for 20 minutes, centrifuging, washing with ultrapure water for three times to obtain the sunflower pollen substrate with positive surface, soaking the pollen substrate in blue quantum dot CdTe 465 aqueous solution modified by thioglycolic acid, standing for 60 minutes, centrifuging, washing with ultrapure water for three times, referring to the polyelectrolyte adsorption step, adsorbing positive-negative-positive CdTe microsphere layers (sequentially adsorbing PAH polyelectrolyte solution, PAH solution and PAH solution) on the outer surface of the blue quantum dot, then adsorbing a first green dot CdTe layer on the outer surface of the CdTe-negative-positive CdTe microsphere layer, adsorbing a green dot layer 513, and continuously adsorbing a second green microsphere layer (sequentially), and depositing the green encoded dots 626 by adopting a polyelectrolyte coding technology to obtain a composite coding layer with different colors of the sunflower pollen with different colors, wherein the code peaks are 1mg/m L;
the prepared sunflower pollen-based composite quantum dot coded microspheres (code of CdTe 465: CdTe 513: CdTe626= 1:2: 3) and quantum dots (one layer of CdTe 465, two layers of CdTe 513, and three layers of CdTe 626) corresponding to the number of layers are subjected to emission spectrum measurement under the same ultraviolet excitation light, and fluorescence emission spectrograms of the prepared sunflower pollen-based composite quantum dot coded microspheres are measured by using an optical fiber spectrometer, as shown in FIG. 3A, wherein a1 is the fluorescence emission spectrogram of the sunflower pollen-based composite quantum dot coded microspheres prepared in the embodiment, b1 is the fluorescence emission spectrogram of one layer of CdTe 465, c1 is the fluorescence emission spectrogram of two layers of CdTe 513, d1 is the fluorescence emission spectrogram of three layers of CdTe, and as can be seen from the figure, the fluorescence intensity of the quantum dots is not obviously reduced after the quantum dots are modified on a pollen substrate.
Example 2
A ragweed pollen-based composite quantum dot coding microsphere is prepared by the following steps:
(1) method for preparing ragweed pollen base without autofluorescence by carbonization
Soaking ragweed pollen in absolute ethyl alcohol, keeping shaking for 20 min to obtain dispersion liquid with fully stripped pollen grains, heating the dispersion liquid on a hot bench at 100 deg.C until the ethanol is completely volatilized, and sieving out dry ragweed pollen grains with clear grains; placing the dried ragweed pollen grains in a muffle furnace, calcining at the high temperature of 300 ℃, and continuously introducing nitrogen for 12 hours to obtain a positively charged ragweed pollen substrate after the fluorescent organic substance is completely carbonized;
(2) ragweed pollen-based composite quantum dot encoding microsphere prepared by polyelectrolyte layer-by-layer adsorption method
Soaking the ragweed pollen substrate without autofluorescence in a PAA solution, standing and adsorbing for 20 minutes, centrifuging, washing with ultrapure water for three times to wash off redundant PAA solution, soaking pollen microspheres in the PAH solution, standing and adsorbing for 20 minutes, centrifuging, washing with ultrapure water for three times to obtain a ragweed pollen substrate with a positive surface, and preparing the ragweed pollen substrate based on the ragweed pollen by a polyelectrolyte layer-by-layer adsorption method in example 1, wherein blue quantum dots CdSe460, first layer green quantum dots CdSe 525, second layer green quantum dots CdSe 525 and red quantum dots CdSe620 modified by thioglycolic acid are adsorbed layer by a polyelectrolyte isolation layer (sequentially adsorbing the PAH solution, the PAA solution and the PAH solution), so as to obtain the ragweed pollen-based composite quantum dot coding microspheres, wherein the coding microspheres adopt different colors (characteristic emission peaks) and different intensities (layer number of quantum dots) to carry out composite coding, and the final obtained code CdSe is that the ratio of 1:2: 525;
the prepared ragweed pollen-based composite quantum dot coded microspheres (coded as CdSe 460: CdSe 525: CdSe620= 1:2: 1) and quantum dots (one layer of CdSe460, two layers of CdSe 525, one layer of CdSe 620) corresponding to the number of layers are subjected to emission spectrum measurement under the same ultraviolet excitation light, and the fluorescence emission spectra of the ragweed pollen-based composite quantum dot coded microspheres are measured by using a fiber optic spectrometer, as shown in fig. 3B, wherein a2 is the fluorescence emission spectrum of the sunflower pollen-based composite quantum dot coded microspheres prepared in the present embodiment, B2 is the fluorescence emission spectrum of the quantum dots, one layer of CdSe460, c2 is the fluorescence emission spectrum of the two layers of quantum dots 525, and d2 is the fluorescence emission spectrum of the quantum dots, one layer of CdSe620, and as can be seen from the figure, the fluorescence intensity of the quantum dots is not obviously reduced after being modified on the pollen substrate.
Example 3
A mixed type composite quantum dot coding microsphere based on chrysanthemum pollen is prepared by the following steps:
(1) soaking chrysanthemum pollen in absolute ethyl alcohol, keeping shaking for 20 minutes to obtain a dispersion liquid with fully stripped pollen grains, heating the dispersion liquid on a hot bench at 100 ℃ until the ethanol is fully volatilized, keeping the pollen in a dry state, and screening out dry ragweed pollen grains with clear grains; placing the dried chrysanthemum pollen grains in a muffle furnace, calcining at the high temperature of 300 ℃, and continuously introducing nitrogen for 12 hours to obtain a positively charged chrysanthemum pollen substrate after the fluorescent organic substance is completely carbonized;
(2) mixed type composite quantum dot coding microsphere based on chrysanthemum pollen prepared by polyelectrolyte layer-by-layer adsorption method
The method comprises the steps of respectively dissolving PEI and PSS solid particles in 0.5 mol/L NaCl solution to prepare PEI solution and PSS solution with the concentration of 1mg/m L, soaking the chrysanthemum pollen substrate without autofluorescence in the PSS solution, standing for adsorption for 20 minutes, centrifuging, washing with ultrapure water for three times to wash off redundant PSS solution, soaking pollen microspheres in PEI solution, standing for adsorption for 20 minutes, centrifuging, washing with ultrapure water for three times to obtain a chrysanthemum pollen substrate with a positively charged surface, and according to the polyelectrolyte layer-by-layer adsorption method of example 1, adsorbing a first blue quantum dot 540, a second blue quantum dot 540 and a third blue quantum dot CdS 540 layer by layer through polyelectrolyte isolating layers (sequentially adsorbing PEI solution, PSS solution and CdS solution) to obtain the chrysanthemum pollen substrate with the coded CdS 540, the chrysanthemum pollen-based composite quantum dot coded microsphere with the CdS 540, the first quantum dot 580, the second quantum dot 580 and the third dot CdS 540, preparing the composite quantum dot-based on chrysanthemum pollen, wherein the coded microsphere is based on the chrysanthemum pollen substrate with the coded CdS 540= 3, and according to the polyelectrolyte layer-by-layer adsorption method, adsorbing the composite quantum dot coded microsphere with the coded quantum dots 540, the chrysanthemum pollen substrate 540, the coded microsphere with the characteristics of the chrysanthemum pollen-based on the mixed quantum dot coded quantum dot, namely, the chrysanthemum pollen-coded microsphere with the chrysanthemum pollen substrate with the chrysanthemum pollen-coded quantum dot coded quantum dots with the characteristics of the chrysanthemum-coded quantum dots with the chrysanthemum quantum dots 540, the chrysanthemum-coded quantum dots with the characteristics of the chrysanthemum-coded quantum dots with the;
in the embodiment, a layer of PEI solution is deposited on the outer side of the quantum dot on the outermost layer of each composite quantum dot coding microsphere, and the PEI solution can be used for detecting various molecules to be detected, wherein the molecules are selected from one of DNA, RNA and protein.
Example 4: characterization experiment of composite quantum dot encoding microspheres based on natural spiny pollen
4.1) preparing dried sunflower pollen grains, sunflower pollen-based composite quantum dot-encoded microspheres (encoded as CdSe620= 1), sunflower pollen-based composite quantum dot-encoded microspheres (encoded as CdSe460= 1), sunflower pollen-based composite quantum dot-encoded microspheres (encoded as CdSe 525= 1);
the preparation method comprises the following steps of (1) soaking sunflower pollen in absolute ethyl alcohol for oscillation for 20 minutes to obtain a dispersion liquid with fully peeled pollen grains, placing the dispersion liquid on a hot table, heating to 100 ℃ until the ethyl alcohol is completely volatilized, enabling the pollen to be in a dry state, and obtaining dry sunflower pollen grains with distinct particles, placing the dry sunflower pollen grains with partially dried sunflower pollen grains in a muffle furnace, calcining at a high temperature of 300 ℃ and continuously introducing nitrogen for 12 hours to obtain a sunflower pollen base without autofluorescence, sequentially soaking the sunflower pollen base in a PSS solution and a PAH solution with the concentration of 1mg/m L, and soaking the sunflower pollen base with positive surface charges in quantum dot CdSe620 to obtain a composite quantum dot encoding microsphere with a surface modified by a layer of red quantum dot CdSe620 based on the natural sunflower pollen, namely the composite quantum dot encoding microsphere with the sunflower pollen based on the sunflower pollen (the encoding is CdSe620= 1), similarly soaking the sunflower pollen base with positive surface modified by a quantum dot encoding microsphere with the natural sunflower pollen in the quantum dot CdSe620 to obtain a composite quantum dot encoding microsphere with the sunflower pollen based on the natural sunflower pollen =1, and soaking the sunflower pollen base in the composite quantum dot encoding microsphere based on the sunflower pollen (the sunflower pollen encoding 460= sunflower pollen) to obtain the natural sunflower pollen encoding microsphere with positive surface encoding quantum dot).
4.2) carrying out characterization by a body microscope and a field emission scanning electron microscope on the dried sunflower pollen grains prepared in the step 4.1) and the composite quantum dot encoding microspheres (coded as CdSe620= 1), wherein the results are shown in a figure 4; as can be seen from fig. 4d-f, the diameter of the sunflower pollen-based composite quantum dot encoded microsphere (encoded as CdSe620= 1) is within the range of 20-30 μm, which is reduced by about 10 μm compared with the size of the natural sunflower pollen (fig. 4 a-c), which is the result of high-temperature calcination at 300 ℃, although the diameter of the composite quantum dot encoded microsphere is reduced, the retention degree of the pollen thorn-shaped structure is better, and at the same time, it can be obviously observed that the quantum dot CdSe620 is uniformly distributed on the surface of the pollen substrate, which indicates that the natural spiny pollen is used as the encoded substrate, the unique and complex porous and spiny surface structure thereof provides high porosity and large specific surface area, and multiple types of quantum dots can be uniformly combined by adopting a polyelectrolyte layer-by-layer adsorption method;
4.3) performing laser scanning confocal microscope scanning on the sunflower pollen-based composite quantum dot coded microspheres (coded as CdSe620= 1) prepared in the step 4.1) to obtain fluorescence images of different sections, wherein as shown in fig. 5, the red quantum dot CdSe620 (bright white part in the figure) covers the whole pollen surface and is only distributed on the surface, which indicates that the spiny pollen with good monodispersity is adopted as a coding substrate, the unique and complex porous and spiny surface structure provides high porosity and large specific surface area, and an effective quantum dot coating can be obtained by a polyelectrolyte layer-by-layer adsorption method, so that the adsorption quantity of target molecules can be increased, the detection fluorescence signal can be obviously enhanced, the detection limit can be greatly reduced, and the detection sensitivity can be improved;
4.4) performing fluorescence characterization under a laser scanning confocal fluorescence microscope on the sunflower pollen-based composite quantum dot coded microspheres (coded as CdSe460= 1), the sunflower pollen-based composite quantum dot coded microspheres (coded as CdSe 525= 1), and the sunflower pollen-based composite quantum dot coded microspheres (coded as CdSe620= 1) prepared in step 4.1), wherein fig. 6a-c are fluorescence images of the sunflower pollen-based composite quantum dot coded microspheres (coded as CdSe460= 1), blue quantum dot CdSe460 (bright part in the figure) covers the whole pollen surface and is only distributed on the surface, fig. b, c are respectively graphs a under different magnifications, and fig. 6d-f are fluorescence images of the sunflower pollen-based composite quantum dot coded microspheres (coded as CdSe 525= 1), green quantum dots CdSe 525 (bright part in the figure) covers the whole pollen surface and only distributes on the surface, e, f are respectively the pictures d under different magnifications, 6g-i are the fluorescence images of the sunflower pollen-based composite quantum dot coding microsphere (coded as CdSe620= 1), red quantum dots CdSe620 (bright part in the figure) covers the whole pollen surface and only distributes on the surface, h, i are respectively the pictures g under different magnifications, and at the same time, stronger fluorescence can be observed at the top of the pollen surface spine;
mixing a sunflower pollen-based composite quantum dot coded microsphere (coded as CdSe460= 1), a sunflower pollen-based composite quantum dot coded microsphere (coded as CdSe 525= 1) and a sunflower pollen-based composite quantum dot coded microsphere (coded as CdSe620= 1) to obtain three different specificity sunflower pollen-based mixed type composite quantum dot coded microspheres, namely, a combination of composite quantum dot coded microspheres with quantum dot codes of CdSe460=1, CdSe 525=1 and CdSe620=1 respectively; the mixed type composite quantum dot coded microspheres based on sunflower pollen (the codes are CdSe460=1, CdSe 525=1 and CdSe620=1 respectively) are subjected to fluorescence characterization under a laser scanning confocal fluorescence microscope, as shown in FIG. 7, under the excitation of the same ultraviolet light source, quantum dots with multiple characteristic emission peaks can be excited simultaneously, and coded microspheres with three colors of red (the code is CdSe620= 1), green (the code is CdSe 525= 1) and blue (the code is CdSe460= 1) can be detected simultaneously, which indicates that the decoding method of the composite quantum dot coded microspheres or mixed type composite quantum dot coded microspheres based on natural spiny pollen prepared by the invention is simple, and the quantum dots with multiple characteristic emission peaks can be excited simultaneously only by light with one wavelength.
Example 5: DNA hybridization assay
(1) Taking the sunflower pollen-based composite quantum dot-coded microsphere prepared in example 4 (coded as CdSe460= 1) as an example, probe DNA is immobilized on the composite quantum dot-coded microsphere by firstly performing amine functionalization on the 5 'end of the probe DNA and labeling the 5' end of the labeled DNA with green fluorescent dye carboxylic acid fluorescein (FAM with a characteristic peak position of 520 nm), a hybridization sandwich structure is shown in FIG. 8, the prepared composite quantum dot-coded microsphere with carboxyl is added into a 50 m L ready-prepared 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride EDC (4.2 mg/m L) solution, and simultaneously, the active intermediate is stabilized with a 50 m L N-hydroxysuccinimide NHS (5 mg/m L) solution, then the 1m L amine-functionalized probe DNA solution is added into the mixture, and the sample is incubated at room temperature for 2 hours while continuously oscillating at 4 ℃ under the condition of room temperature, the carboxyl-coded microsphere with amino acid is capable of being immobilized with the sunflower pollen-coded microsphere after incubation with the amino acid, the amino acid-coded microsphere is subjected to a hybridization reaction under the room temperature scanning microscope, the hybridization probe DNA buffer is immobilized on the sunflower pollen-coded microsphere solution, the sunflower pollen-coded microsphere after the hybridization probe is incubated for three times, the hybridization probe DNA, the hybridization sandwich structure, the hybridization probe DNA is subjected to form a fluorescent probe DNA buffer solution, the hybridization sandwich structure, the fluorescent probe DNA, the fluorescent dye-coded microsphere is immobilized under the fluorescent dye, the fluorescent probe DNA buffer solution, the fluorescent dye-coded microsphere is immobilized under the fluorescent probe, the fluorescent probe DNA buffer solution, the fluorescent probe is immobilized under the fluorescent probe, the fluorescent probe is immobilized under the fluorescent sandwich structure under the fluorescent probe DNA buffer structure under the fluorescent probe, the fluorescent sandwich structure under the conditions of sunflower.
Referring to the above steps, the performance of sunflower pollen-based composite quantum dot-encoded microspheres (encoded as CdSe460= 1) and Glass Beads in DNA hybridization assay experiments was compared using traditional bioassay product Glass Beads (Glass Beads) as controls, with other experimental parameter conditions being identical. And (3) performing fluorescent image and emission spectrum characterization on Glass Beads of the hybridization sandwich structure DNA with the surface combined with the modified FAM by using a laser scanning confocal microscope and a fiber optic spectrometer. As shown in fig. 9i and 9ii, the green fluorescence of the labeled dye FAM can be detected by the fluorescence image, and the fluorescence intensity emitted by the sunflower pollen-based composite quantum dot-encoded microsphere (encoded by CdSe460= 1) is much higher than that of Glass Beads. As shown in fig. 9, the fluorescence curve proves that the sunflower pollen-based composite quantum dot-encoded microspheres (encoded by CdSe460= 1) can detect a wider DNA concentration range than Glass Beads, and further the detection limit of the Glass Beads is 1090pM, while the quantum dot-encoded microspheres prepared by the invention have the detection limit of only 9.7 pM, so that the spiny pollen is used as an encoding substrate, and the unique and complex porous and spiny surface structure thereof provides high porosity and large specific surface area, increases the adsorption amount of target molecules, further significantly enhances the detection fluorescence signal, greatly reduces the detection limit, and improves the detection sensitivity.
(2) Determination of multiple DNA target sequences
Referring to the preparation method of example 3, mixed type composite quantum dot-coded microspheres based on sunflower pollen (coded as CdSe 460: CdSe620= 2:1, 1:1, 1: 2) were prepared, referring to the above step (1) of this example, different probe DNAs (1 nmol/L) were immobilized on three sunflower pollen-based composite quantum dot-coded microspheres (coded as CdSe 460: CdSe620= 2:1, CdSe 460: CdSe620= 1:2, respectively), different carrier-probe conjugates were mixed at room temperature and four target DNA molecules with different sequences were added at equimolar ratio, the mixture was incubated continuously for 6 hours at 37 ℃ in the dark to obtain mixed type composite quantum dot-coded microspheres with hybrid sandwich structure DNA bound on the surface (coded as CdSe 460: CdSe 620: 1, 1:1, 1: 2), then a spectrometer was used to detect the reflected spectral signals on different composite quantum dot-coded microspheres (coded as CdSe 460: 620: 1, 1:2, 1: 2), and the combined emission spectrum of the mixed type coded quantum dot-coded microspheres was obtained, and the detection results were obtained by washing the same time, the detection results of the multiple CdSe-coded quantum dot-coded composite quantum dot-coded microsphere, the detection results of the multiple quantum dot-coded peaks of the emission spectrum of the emission peaks of the different composite quantum dots were obtained under the same type of the same fluorescent quantum dots can be clearly observed under the same spectrum of the same type of the same fluorescent nanoparticles, the same type of the same fluorescent quantum dots can be detected, the same type of the same fluorescent quantum dots, the same type of the fluorescent quantum dots can be detected, the same type of the fluorescent quantum dots, the same type of the fluorescent nanoparticles, the same type of.
Example 6: emission spectra and stability characterization of quantum dot encoding
The reference is the preparation method of the composite quantum dot coding microsphere described in the embodiment 1, the natural sunflower pollen is used as a substrate, and the polyelectrolyte layer-by-layer deposition technology is adopted, modifying a layer of CdSe @ ZnS565 quantum dots on the surface of a sunflower pollen substrate so as to prepare a sunflower pollen-based composite quantum dot coding microsphere (the code is CdSe @ ZnS565= 1), the excitation spectrum and the emission spectrum of the natural sunflower pollen and the composite quantum dot coded microsphere (coded as CdSe @ ZnS565= 1) based on the sunflower pollen are measured by a fluorescence spectrophotometer, and the results are shown in FIG. 11, the sunflower pollen-based composite quantum dot coded microsphere (coded as CdSe @ ZnS565= 1) (marked as the composite quantum dot coded microsphere in the figure) can be observed to have a wide excitation spectrum and a narrow emission spectrum, and compared with natural sunflower pollen, the fluorescence of the composite quantum dot coded microsphere has a strong optical coding advantage;
in the DNA hybridization assay experiment of example 5, the change of fluorescence intensity of the sunflower pollen-based composite quantum dot-encoded microsphere (encoded as CdSe460= 1) after DNA binding was detected by fiber optic spectrometer, and as a result, as shown in fig. 12, it can be seen that the decrease of the fluorescence intensity of the quantum dot is small over a long period of time; the composite coding microsphere based on natural spiny pollen prepared by the invention has better coding stability in the application process.
The above is only a preferred embodiment of the present invention, and the protection scope of the present invention is not limited to the above-mentioned embodiments, and all technical solutions belonging to the idea of the present invention belong to the protection scope of the present invention. It should be noted that modifications and embellishments within the scope of the invention may be made by those skilled in the art without departing from the principle of the invention.
Claims (10)
1. A preparation method of a composite quantum dot coding microsphere based on natural spiny pollen is characterized by comprising the following steps:
(1) carrying out dispersive screening on the thorny-shaped pollen, and carrying out carbonization treatment on the screened pollen grains to obtain a pollen substrate after autofluorescence is eliminated;
(2) the method comprises the steps of sequentially soaking a pollen substrate in a negative polyelectrolyte solution and a positive polyelectrolyte solution to obtain a microsphere substrate with a positively charged surface, and adsorbing quantum dots with different layers, different types or different sizes layer by layer on the outer surface of the microsphere substrate with the positively charged surface by utilizing a polyelectrolyte layer-by-layer deposition technology, so as to obtain the composite quantum dot coding microsphere taking natural multi-thorn-shaped pollen as the substrate.
2. The method of claim 1, wherein: in the step (1), thorny pollen is soaked in absolute ethyl alcohol and vibrated to obtain pollen dispersion liquid with fully stripped pollen grains, the pollen dispersion liquid is dried to disperse and screen out dry pollen grains with clear grains, the dry pollen grains are calcined at the high temperature of 300 ℃ and continuously introduced with nitrogen for 12 hours, so that fluorescent organic substances are completely carbonized to eliminate autofluorescence.
3. The method of claim 2, wherein: the thorny pollen is one selected from sunflower pollen, ragweed pollen and chrysanthemum pollen.
4. The method of claim 1, wherein: the positive polyelectrolyte is selected from one of polyacrylamide hydrochloride and polyethyleneimine, and the negative polyelectrolyte is selected from one of sodium polystyrene sulfonate and polyacrylic acid.
5. The method according to claim 4, wherein the positive or negative polyelectrolyte solution is prepared by dissolving a solid of the positive or negative polyelectrolyte in 0.5 mol/L NaCl solution to obtain a solution of the positive or negative polyelectrolyte having a concentration of 1mg/m L.
6. The method of claim 5, wherein: in the step (2), the microsphere substrate with positive electricity is soaked in the quantum dot solution with negative electricity, the quantum dots are adsorbed on the outer surface of the microsphere substrate, and quantum dots with different layers, different types or different sizes are adsorbed layer by layer through the electrostatic acting force between the positive-negative-positive polyelectrolyte layer and the quantum dots with negative electricity.
7. The method of claim 6, wherein: in the step (2), the quantum dots are selected from one of cadmium sulfide, cadmium selenide, cadmium telluride and cadmium selenide quantum dots coated by zinc sulfide.
8. The method of claim 7, wherein: in the step (2), a layer of positive polyelectrolyte is continuously deposited outside the quantum dots on the outermost layer so as to facilitate the grafting of subsequent detection molecules, wherein the detection molecules are selected from one of DNA, RNA and protein.
9. The composite quantum dot coding microsphere based on natural spiny pollen is characterized by being prepared by the preparation method of any one of claims 1 to 8, wherein the natural spiny pollen is used as a substrate, and quantum dots with different layers, different types or different sizes are adsorbed on the surface layer by layer.
10. The mixed type composite quantum dot coding microsphere based on natural spiny pollen is characterized in that the mixed type composite quantum dot coding microsphere is formed by mixing composite quantum dot coding microspheres with different specificities, the composite quantum dot coding microsphere is prepared by adopting the preparation method of any one of claims 1-8, the natural spiny pollen is used as a substrate, and quantum dots with different layers, different types or different sizes are adsorbed on the surface layer by layer.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202010303050.2A CN111410951B (en) | 2020-04-17 | 2020-04-17 | Composite quantum dot encoding microsphere based on natural spiny pollen and preparation method thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202010303050.2A CN111410951B (en) | 2020-04-17 | 2020-04-17 | Composite quantum dot encoding microsphere based on natural spiny pollen and preparation method thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN111410951A true CN111410951A (en) | 2020-07-14 |
| CN111410951B CN111410951B (en) | 2020-11-24 |
Family
ID=71489930
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202010303050.2A Active CN111410951B (en) | 2020-04-17 | 2020-04-17 | Composite quantum dot encoding microsphere based on natural spiny pollen and preparation method thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN111410951B (en) |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113189698A (en) * | 2021-05-11 | 2021-07-30 | 河南工业大学 | Preparation method of multifunctional diamagnetic optical fiber doped with composite quantum dots |
| CN113218920A (en) * | 2021-02-04 | 2021-08-06 | 安徽师范大学 | Preparation method of fluorescent carbon nano-microspheres based on chrysanthemum and preparation method of fluorescent carbon nano-microspheres based on chrysanthemum2+And detection of captopril |
| CN113611228A (en) * | 2021-08-05 | 2021-11-05 | 深圳市电通材料技术有限公司 | Fluorescent texture identification based on multi-emission peak wavelength fluorescent powder particles |
| CN114788797A (en) * | 2022-03-17 | 2022-07-26 | 南京深呼吸生物科技有限公司 | Smearing type color additive based on composite assembly particles and preparation method thereof |
| CN117310147A (en) * | 2023-07-06 | 2023-12-29 | 中国人民解放军陆军军医大学第二附属医院 | Application of biological microsphere in DNA selective adsorption and microRNA detection |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1884430A (en) * | 2006-06-30 | 2006-12-27 | 上海师范大学 | Fluorescent carbon nanometer tube and its preparation method and application |
| CN101216415A (en) * | 2008-01-08 | 2008-07-09 | 东南大学 | Photon crystal composite encoding microsphere and preparation method |
-
2020
- 2020-04-17 CN CN202010303050.2A patent/CN111410951B/en active Active
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1884430A (en) * | 2006-06-30 | 2006-12-27 | 上海师范大学 | Fluorescent carbon nanometer tube and its preparation method and application |
| CN101216415A (en) * | 2008-01-08 | 2008-07-09 | 东南大学 | Photon crystal composite encoding microsphere and preparation method |
Non-Patent Citations (3)
| Title |
|---|
| JUAN LI等: "Colloidal crystal beads coated with multicolor CdTe quantum dots: microcarriers for optical encoding and fluorescence enhancement", 《J. MATER. CHEM.》 * |
| PING YANG等: "Phenol Formaldehyde Resin Nanoparticles Loaded with CdTe Quantum Dots: A Fluorescence Resonance Energy Transfer Probe for Optical Visual Detection of Copper(II) Ions", 《ACS NANO》 * |
| ZHANG BO等: "Polyethyleneimine-interlayered silica-core quantum dot-shell nanocomposites for sensitive detection of Salmonella typhimurium via a lateral flow immunoassay", 《RSC ADVANCES》 * |
Cited By (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113218920A (en) * | 2021-02-04 | 2021-08-06 | 安徽师范大学 | Preparation method of fluorescent carbon nano-microspheres based on chrysanthemum and preparation method of fluorescent carbon nano-microspheres based on chrysanthemum2+And detection of captopril |
| CN113218920B (en) * | 2021-02-04 | 2022-12-23 | 安徽师范大学 | Fluorescent carbon based on chrysanthemum preparation method of nano-microsphere and para-Hg 2+ Detection of Subtopril |
| CN113189698A (en) * | 2021-05-11 | 2021-07-30 | 河南工业大学 | Preparation method of multifunctional diamagnetic optical fiber doped with composite quantum dots |
| CN113611228A (en) * | 2021-08-05 | 2021-11-05 | 深圳市电通材料技术有限公司 | Fluorescent texture identification based on multi-emission peak wavelength fluorescent powder particles |
| CN114788797A (en) * | 2022-03-17 | 2022-07-26 | 南京深呼吸生物科技有限公司 | Smearing type color additive based on composite assembly particles and preparation method thereof |
| CN114788797B (en) * | 2022-03-17 | 2024-01-12 | 南京深呼吸生物科技有限公司 | Smearing type color additive based on composite assembly particles and preparation method thereof |
| CN117310147A (en) * | 2023-07-06 | 2023-12-29 | 中国人民解放军陆军军医大学第二附属医院 | Application of biological microsphere in DNA selective adsorption and microRNA detection |
| CN117310147B (en) * | 2023-07-06 | 2024-07-02 | 中国人民解放军陆军军医大学第二附属医院 | Application of biological microsphere in DNA selective adsorption and microRNA detection |
Also Published As
| Publication number | Publication date |
|---|---|
| CN111410951B (en) | 2020-11-24 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN111410951B (en) | Composite quantum dot encoding microsphere based on natural spiny pollen and preparation method thereof | |
| Smith et al. | Bioconjugated silica-coated nanoparticles for bioseparation and bioanalysis | |
| Trau et al. | Novel colloidal materials for high‐throughput screening applications in drug discovery and genomics | |
| Jun et al. | Protein separation and identification using magnetic beads encoded with surface-enhanced Raman spectroscopy | |
| CN110791286B (en) | Perovskite fluorescent microsphere and preparation method and application thereof | |
| EP1490691B1 (en) | Luminescent, spheroid, non-autofluorescent silica gel particles having variable emission intensities and frequencies | |
| Cao et al. | Preparation of silica encapsulated quantum dot encoded beads for multiplex assay and its properties | |
| Gao et al. | Quantum dot-encoded beads | |
| Amiri et al. | Application of ratiometric fluorescence sensor-based microwave-assisted synthesized CdTe quantum dots and mesoporous structured epitope-imprinted polymers for highly efficient determination of tyrosine phosphopeptide | |
| US7875255B2 (en) | Nanoparticles comprising calcium phosphate and ethylene imine compositions and methods of production thereof | |
| CN1826171A (en) | Colorable microspheres for DNA and protein microarray | |
| CN106957644A (en) | A kind of fingerprint detection probe based on long-persistence nano material and preparation method thereof and the application in latent fingerprint detection | |
| Wang et al. | Quantum dots integrated biomass pollens as functional multicolor barcodes | |
| Zhu et al. | Encoding through the host–guest structure: construction of multiplexed fluorescent beads | |
| Zhang et al. | Polymers mediate a one-pot route for functionalized quantum dot barcodes with a large encoding capacity | |
| Liu et al. | Microfluidic generation of uniform quantum dot-encoded microbeads by gelation of alginate | |
| Gu et al. | A membrane biosensor based on colour differences encoded upconversion silica nanoparticles for multiplex nucleic acids detection | |
| Zhang et al. | Facile preparation of polydiacetylene-based uniform porous fluorescent microspheres for potential immunoassay applications | |
| Li et al. | Colloidal crystal beads coated with multicolor CdTe quantum dots: microcarriers for optical encoding and fluorescence enhancement | |
| Ren et al. | DNA-mediated anisotropic silica coating of upconversion nanoparticles | |
| Ma et al. | Synthesis and application of quantum dot-tagged fluorescent microbeads | |
| Li et al. | Rewritable magnetic fluorescence-encoded microspheres: preparation, characterization, and recycling | |
| CN108752522B (en) | Preparation of quantum dot biosensor and detection method of bovine hemoglobin | |
| Halabi et al. | Magnetic Silica‐Coated Fluorescent Microspheres (MagSiGlow) for Simultaneous Detection of Tumor‐Associated Proteins | |
| Wang et al. | Rhodamine B doped silica nanoparticle labels for protein microarray detection |
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 | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |