Up-conversion fluorescent microsphere and preparation method thereof
The upconverted fluorescent microspheres are prepared by microfluidic control methods, which solves the problems of uneven luminescence distribution, limited load capacity and cumbersome process in the prior art, and achieves the effects of uniform luminescence, stable performance and high load capacity.
Patent Information
- Application Number
- CN202411728609.0
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2024-11-28
- Publication Date
- 2025-02-25
AI Technical Summary
In the prior art, when the upconverted fluorescent microspheres are prepared by swelling method, the luminescence distribution is uneven, the loading of the upconverted nanoparticles is limited, and the process is cumbersome and time-consuming.
Upconverted fluorescent microspheres were prepared by microfluidic control method. By using interfacial tension and flow shear force on the microfluidic chip, the upconverted nanoparticle-polymer organic solution was cut into tiny dispersed droplets to achieve uniform distribution of upconverted nanoparticles.
The upconverted fluorescent microspheres have uniform luminescence distribution and stable fluorescence performance, and the loading of upconverted nanoparticles is improved, with simple process and good repeatability.
Smart Images
Figure CN119505906A_ABST
Abstract
Description
Technical Field
[0001] The invention relates to the technical field of up-conversion fluorescent microspheres, and in particular to up-conversion fluorescent microspheres and a preparation method thereof. Background Art
[0002] Upconversion fluorescent nanoparticles are materials that emit visible and ultraviolet bands when excited by near-infrared light. This type of material solves the shortcomings of traditional fluorescent materials, such as resistance to photobleaching, shallow penetration depth of excitation light, and strong interference from sample autofluorescence. Unlike traditional fluorescent materials such as organic dyes and quantum dots, upconversion fluorescent nanoparticles can produce a characteristic fluorescence emission spectrum with a very narrow peak width when excited by 980nm near-infrared light. The wavelength range of the emission peak can be precisely adjusted in the near-infrared, visible and ultraviolet bands by adjusting the type and / or concentration of doped rare earth ions.
[0003] Since biological molecules themselves do not have upconversion optical properties, there is almost no background fluorescence noise when using upconversion fluorescent nanoparticles for fluorescent labeling, which makes upconversion fluorescent materials very promising to become a type of fluorescent probe that provides high-sensitivity detection for trace target molecules. In addition, since the core of upconversion fluorescent materials is synthesized from biologically inert elements and the rare earth ions doped therein are slightly toxic, upconversion fluorescent materials have great advantages in safety compared to quantum dots using highly toxic metal ions such as cadmium-selenium ion pairs. These characteristics give upconversion nanoparticles unique advantages in the preparation of stable, highly sensitive, safe and large-scale microspheres for use in the biomedical field.
[0004] In the related art, upconversion fluorescent microspheres are mainly obtained by encapsulating upconversion nanoparticles in polystyrene microspheres through a swelling method. The volume of polystyrene microspheres swells in a specific solvent, and micropores appear in the microspheres after expansion, and the upconversion nanoparticles diffuse into these micropores; then these microspheres loaded with nanoparticles are placed in a specific solvent to shrink the volume of the microspheres to their original size, so that the upconversion nanoparticles are stably encapsulated in the microspheres. However, in the upconversion fluorescent microspheres obtained by this method, the upconversion nanoparticles are mainly distributed on the surface of the microspheres, which makes the luminescence distribution of the microspheres uneven and greatly limits the loading amount of the upconversion nanoparticles; in addition, this method based on the swelling of polystyrene microspheres is cumbersome and time-consuming.
[0005] Therefore, improving the preparation method of upconversion fluorescent microspheres in the related art and providing a new preparation method of upconversion fluorescent microspheres have become technical problems that need to be solved urgently by those skilled in the art. Summary of the invention
[0006] The invention discloses an upconversion fluorescent microsphere and a preparation method thereof, so as to solve the technical problems that when the upconversion fluorescent microsphere is prepared by a swelling method in the related art, the obtained upconversion fluorescent microsphere has uneven luminescence distribution, the loading amount of upconversion nanoparticles is limited, and the process is complicated and time-consuming.
[0007] In order to solve the above problems, the present invention adopts the following technical solutions: The first aspect of the present invention provides a method for preparing upconversion fluorescent microspheres.
[0008] The method for preparing upconversion fluorescent microspheres of the present invention adopts a microfluidic method to prepare upconversion fluorescent microspheres, and the method for preparing upconversion fluorescent microspheres comprises the following steps: preparing a first surfactant aqueous solution, with the first surfactant aqueous solution serving as a continuous phase; Mixing upconversion nanoparticles, high molecular polymers, and an organic solution to obtain an upconversion nanoparticle-high molecular polymer organic solution, wherein the upconversion nanoparticle-high molecular polymer organic solution is a dispersed phase; preparing a second surfactant aqueous solution, and using the second surfactant aqueous solution as a collection phase; The continuous phase and the dispersed phase are introduced into a microfluidic chip, and under the action of interfacial tension and flow shear force, the dispersed phase is cut into tiny dispersed droplets to obtain upconversion fluorescent microspheres, and the obtained upconversion fluorescent microspheres are collected by the collecting phase located at the outlet of the microfluidic chip.
[0009] According to an optional embodiment, when the continuous phase and the dispersed phase are passed into the microfluidic chip, the flow rate ratio of the continuous phase to the dispersed phase is 1:1 to 50:1.
[0010] According to an optional embodiment, preparing the first surfactant aqueous solution comprises the following steps: The first surfactant is mixed with deionized water, and then ultrasonically mixed for later use.
[0011] According to an optional embodiment, the first surfactant is one or more of a sodium alkylbenzene sulfonate surfactant, a Triton-X surfactant, a polyvinyl alcohol surfactant and a Tween surfactant, and the mass fraction of the first surfactant aqueous solution is 1% to 20%.
[0012] According to an optional embodiment, the preparation of the upconversion nanoparticle-polymer organic solution comprises the following steps: The upconversion nanoparticles, the high molecular polymer and the organic solution are mixed, and ultrasonically mixed and then set aside; Alternatively, the upconversion nanoparticles are dissolved in an organic solution to obtain an upconversion nanoparticle organic solution, and then the upconversion nanoparticle organic solution is mixed with a high molecular polymer, or the upconversion nanoparticle organic solution is mixed with the high molecular polymer and the organic solution, and then ultrasonically mixed for later use; Alternatively, the polymer is dissolved in an organic solution to obtain a polymer organic solution, and then the polymer organic solution is mixed with upconversion nanoparticles, or the polymer organic solution is mixed with upconversion nanoparticles and the organic solution, and then ultrasonically mixed for later use.
[0013] According to an optional embodiment, when preparing upconversion nanoparticle-polymer organic solution, the amounts of upconversion nanoparticles, polymer and organic solution meet the following requirements: the mass of the polymer accounts for 1% to 20% of the mass of the dispersed phase; the mass of the upconversion nanoparticles accounts for 0.1 to 10% of the mass of the polymer.
[0014] According to an optional embodiment, the upconversion nanoparticles are NaYF4:Yb,Er nanoparticles or NaYF4:Yb,Tm nanoparticles; The high molecular polymer is one or more of polymethyl methacrylate, polymethacrylic acid, polyacrylic acid, polystyrene, polyglycolic acid, polyisopropylacrylamide or derivatives thereof; The organic solvent is one or more of dichloromethane, chloroform, tetrahydrofuran, toluene, cyclohexane, and N,N'-dimethylacrylamide.
[0015] According to an optional embodiment, preparing the second surfactant aqueous solution comprises the following steps: The second surfactant is mixed with deionized water, and then ultrasonically mixed for later use.
[0016] According to an optional embodiment, the second surfactant is one or more of a sodium alkylbenzene sulfonate surfactant, a Triton-X surfactant, a polyvinyl alcohol surfactant and a Tween surfactant, and the mass fraction of the second surfactant aqueous solution is 1% to 20%.
[0017] According to an optional embodiment, the method for preparing upconversion fluorescent microspheres further comprises the step of drying the collection phase in which the upconversion fluorescent microspheres are collected, and drying the collection phase in which the upconversion fluorescent microspheres are collected comprises: The collection phase containing the up-conversion fluorescent microspheres is allowed to stand at room temperature to evaporate the organic solvent; The upconversion fluorescent microspheres were centrifuged, washed with pure water, and dried in an oven at 60°C.
[0018] The second aspect of the present invention provides an up-conversion fluorescent microsphere.
[0019] The upconversion fluorescent microspheres prepared by the preparation method of the upconversion fluorescent microspheres described in any technical solution of the present invention include upconversion nanoparticles and a polymer, the upconversion nanoparticles are embedded in the polymer, and the fluorescence intensity distribution of the upconversion fluorescent microspheres can be regulated based on the mass of the upconversion nanoparticles and the mass of the polymer, and when the mass of the upconversion nanoparticles accounts for 0.1-10% of the mass of the polymer, the fluorescence intensity fluctuation value of the upconversion fluorescent microspheres is within ±15%.
[0020] The technical solution adopted by the present invention can achieve the following beneficial effects: The preparation method of upconversion fluorescent microspheres of the present invention adopts a microfluidic method to prepare the upconversion fluorescent microspheres. At the microfluidic chip, under the action of interfacial tension and flow shear force, the dispersed phase is cut into tiny dispersed droplets and the upconversion fluorescent microspheres are obtained. The method can manipulate microfluids through microchannels, and can control the flow and mixing of fluids on a microscale, thereby achieving control of the size of the microspheres and the distribution of fluorescent substances. The upconversion fluorescent microspheres obtained by the microfluidic method of the present invention have good monodispersity, and the upconversion nanoparticles are evenly distributed in the microspheres, so that the obtained upconversion fluorescent microspheres have the advantages of uniform luminescence distribution and stable fluorescence performance; and the upconversion nanoparticles are not limited to being distributed on the surface of the microspheres, so the loading amount of the upconversion nanoparticles can also be increased.
[0021] On the other hand, the preparation method of the upconversion fluorescent microspheres of the present invention can realize the controllable fluorescence intensity of the upconversion fluorescent microspheres. Specifically, the fluorescence intensity distribution of the upconversion fluorescent microspheres can be obtained based on the quality of the upconversion nanoparticles and the quality of the high molecular polymer. The fluorescence intensity distribution of the obtained upconversion fluorescent microspheres is uniform and the fluorescence performance is stable. In addition, the preparation method of the upconversion fluorescent microspheres of the present invention also has the advantages of simple process and good repeatability.
[0022] That is, the present invention adopts a microfluidic method to prepare upconversion fluorescent microspheres, which solves the technical problems of uneven luminescence distribution of the upconversion fluorescent microspheres obtained when the upconversion fluorescent microspheres are prepared by the swelling method in the related art, limited loading capacity of upconversion nanoparticles, and complicated and time-consuming process. BRIEF DESCRIPTION OF THE DRAWINGS
[0023] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required for use in the embodiments or the description of the prior art will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying creative work.
[0024] Figure 1 is a schematic diagram of a microfluidic device according to an embodiment of the present application; Figure 2 is a SEM image of the upconversion fluorescent microspheres obtained in Example 1 of the present application; Figure 3 (a) to (c) are respectively the near-infrared fluorescence confocal images of the upconversion fluorescent microspheres obtained in Example 1 of the present application in dark field, bright field and composite field, Figure 3 (d) Yes Figure 3 (a) is a partial schematic diagram. Figure 3 (e) is along Figure 3 (d) Fluorescence intensity distribution diagram of the oblique line; Figure 4 This is a fluorescence confocal 3D layer scan of the upconversion fluorescent microspheres obtained in Example 2 of the present application; Figure 5 is a SEM image of the upconversion fluorescent microspheres obtained in Example 4 of the present application; Figure 6 is a SEM image of the upconversion fluorescent microspheres obtained in Example 6 of the present application; Figure 7 (a) to (d) are optical microscope photos of the upconversion fluorescent microspheres obtained in Examples 1 to 4 of the present application, respectively. Figure 7 The scale bars in (a) to (d) are 100 μm; Figure 8 (a) and (b) are optical microscope photos of the upconversion fluorescent microspheres obtained in Examples 5 and 6 of the present application, respectively. Figure 8 The scale bars in (a) and (b) are 100 μm; Fig. 9 This is a SEM image of the upconversion fluorescent microspheres obtained in the comparative example of the present application.
[0025] In the figure: 10, continuous phase storage tank; 20, dispersed phase storage tank; 30, microfluidic chip; 31, continuous phase inlet; 32, continuous phase channel; 33, dispersed phase inlet; 34, dispersed phase channel; 35, microsphere outlet; 36, microsphere channel; 40, collection phase storage tank. DETAILED DESCRIPTION
[0026] To make the purpose, technical solution and advantages of the present invention clearer, the technical solution of the present invention will be described in detail below. Obviously, the described embodiments are only part of the embodiments of the present invention, rather than all the embodiments. Based on the embodiments of the present invention, all other implementation methods obtained by ordinary technicians in this field without creative work belong to the scope of protection of the present invention.
[0027] The terms "first", "second", etc. in the specification and claims of the present application are used to distinguish similar objects, and are not used to describe a specific order or sequence. It should be understood that the data used in this way can be interchangeable under appropriate circumstances, so that the embodiments of the present application can be implemented in an order other than those illustrated or described here, and the objects distinguished by "first", "second", etc. are generally of one type, and the number of objects is not limited. For example, the first object can be one or more. In addition, "and / or" in the specification and claims represents at least one of the connected objects, and the character " / " generally indicates that the objects associated with each other are in an "or" relationship.
[0028] Figure 1 The microfluidic device used in the preparation method of upconversion fluorescent microspheres in this embodiment is shown. Figure 1 As shown, the microfluidic device includes a continuous phase storage tank 10, a dispersed phase storage tank 20, a microfluidic chip 30 and a collection phase storage tank 40. The microfluidic chip 30 is provided with a continuous phase inlet 31 and a continuous phase channel 32, and the continuous phase inlet 31 is connected to the continuous phase channel 32; the microfluidic chip 30 is provided with a dispersed phase inlet 33 and a dispersed phase channel 34, and the dispersed phase inlet 33 is connected to the dispersed phase channel 34. The dispersed phase channel 34 is cross-intersected with the continuous phase channel 32 on one side, so that the continuous phase flowing in from both sides can generate shear force on the dispersed phase flowing in a straight line and form microspheres. The microfluidic chip 30 is also provided with a microsphere outlet 35 and a microsphere channel 36, and the microsphere channel 36 is connected to the intersection of the dispersed phase channel 34 and the continuous phase channel 32, and the microsphere channel 36 is also connected to the microsphere outlet 35.
[0029] Continue to see Figure 1 The continuous phase storage tank 10 is connected to the continuous phase inlet 31 so that the continuous phase can be introduced into the microfluidic chip 30; the dispersed phase storage tank 20 is connected to the dispersed phase inlet 33 so that the dispersed phase can be introduced into the microfluidic chip 30; the collecting phase storage tank 40 is connected to the microsphere outlet 35 so that the formed microspheres can enter the collecting phase storage tank 40 for collection.
[0030] Furthermore, the connecting pipes between the continuous phase storage tank 10 and the continuous phase inlet 31, the connecting pipes between the dispersed phase storage tank 20 and the dispersed phase inlet 33, and the connecting pipes between the collection phase storage tank 40 and the microsphere outlet 35 are made of polytetrafluoroethylene plastic tubes and polyetheretherketone joints, which can prevent the dispersed phase from swelling the pipes.
[0031] When upconversion fluorescent microspheres are prepared using a microfluidic device, upconversion fluorescent microspheres of different size requirements can be prepared by controlling the length, width and depth of the continuous phase channel 32 and the dispersed phase channel 34 .
[0032] For example, in the microfluidic device used in this embodiment, the width of the continuous phase channel 32 and the dispersed phase channel 34 is 60 microns, the depth is 100 microns, and the length is not limited.
[0033] The preparation method of the upconversion fluorescent microspheres in this embodiment adopts a microfluidic method to prepare the upconversion fluorescent microspheres. In some embodiments, the preparation method of the upconversion fluorescent microspheres comprises the following steps: Step 100: Prepare a first surfactant aqueous solution, with the first surfactant aqueous solution as the continuous phase. Exemplarily, the first surfactant is mixed with deionized water, and then ultrasonically mixed for later use.
[0034] In the process of preparing upconversion fluorescent microspheres, the continuous phase can produce a shearing effect on the dispersed phase and promote the formation of microspheres. Further, the continuous phase is a first surfactant aqueous solution, and the first surfactant can reduce the interfacial tension between the aqueous phase and the oil phase, promoting the formation of stable microspheres; at the same time, the first surfactant can also form a protective film on the surface of the microspheres, which can reduce the leakage of upconversion nanoparticles from the microspheres, so as to further improve the stability of the upconversion fluorescent microspheres; secondly, the presence of the first surfactant can increase the dispersibility of the upconversion nanoparticles in the microspheres, reduce the aggregation of the upconversion nanoparticles, and thus improve the embedding efficiency of the upconversion nanoparticles; thirdly, the presence of the first surfactant can also transfer energy between upconversion nanoparticles, thereby improving the optical stability of the microspheres.
[0035] The first surfactant aqueous solution is used as the continuous phase, and the size and dispersibility of the microspheres can be precisely controlled by selecting the type and concentration of the first surfactant.
[0036] In some embodiments, the first surfactant is one or more of a sodium alkylbenzene sulfonate surfactant, a Triton-X surfactant, a polyvinyl alcohol surfactant, and a Tween surfactant. The mass fraction of the first surfactant aqueous solution is 1% to 20%. Exemplarily, the mass fraction of the first surfactant aqueous solution is 1%, 2%, 5%, 10%, 15%, and 20%.
[0037] Step 200: Mix upconversion nanoparticles, high molecular polymers, and an organic solution to obtain an upconversion nanoparticle-high molecular polymer organic solution, with the upconversion nanoparticle-high molecular polymer organic solution being a dispersed phase.
[0038] The dispersed phase includes upconversion nanoparticles, high molecular polymers and organic solutions, wherein the organic solution can be used as a solvent for the upconversion nanoparticles, and the organic solution is selected from substances with good solubility for the upconversion nanoparticles; the high molecular polymer is used to form microspheres, and when the upconversion nanoparticles are dissolved in the organic solution, the uniformity of the upconversion nanoparticles embedded in the high molecular polymer can be improved.
[0039] In some embodiments, the mixing order of up-conversion nanoparticles, high molecular polymers and organic solutions may not be limited. Exemplarily, the up-conversion nanoparticles, high molecular polymers and organic solutions may be mixed at the same time, and ultrasonic mixing may be used for standby. Exemplarily, the up-conversion nanoparticles may be first dissolved in an organic solution to obtain an up-conversion nanoparticle organic solution, and then the up-conversion nanoparticle organic solution is mixed with the high molecular polymer, or the up-conversion nanoparticle organic solution is mixed with the high molecular polymer and the organic solution, and ultrasonic mixing may be used for standby. Exemplarily, the high molecular polymer may be dissolved in an organic solution to obtain a high molecular polymer organic solution, and then the high molecular polymer organic solution is mixed with the up-conversion nanoparticles, or the high molecular polymer organic solution is mixed with the up-conversion nanoparticles and the organic solution, and ultrasonic mixing may be used for standby.
[0040] In some embodiments, the upconversion nanoparticles are NaYF4:Yb, Er nanoparticles or NaYF4:Yb, Tm nanoparticles. Not limited thereto, the upconversion nanoparticles may also be made of other fluorescent materials. By changing the type of upconversion nanoparticles, the fluorescence properties of the upconversion fluorescent microspheres can be adjusted.
[0041] In some embodiments, the high molecular polymer is one or more of polymethyl methacrylate, polymethacrylic acid, polyacrylic acid, polystyrene, polyglycolic acid, polyisopropylacrylamide or their derivatives. Not limited thereto, the high molecular polymer may also be other materials.
[0042] In some embodiments, the organic solvent is one or more of dichloromethane, chloroform, tetrahydrofuran, toluene, cyclohexane, and N,N'-dimethylacrylamide. Not limited thereto, the organic solvent may also be other substances that have good solubility for upconversion nanoparticles.
[0043] In some embodiments, the amounts of upconversion nanoparticles, high molecular polymer, and organic solution satisfy: the mass of the high molecular polymer accounts for 1% to 20% of the mass of the dispersed phase; the mass of the upconversion nanoparticles accounts for 0.1 to 10% of the mass of the high molecular polymer.
[0044] By adopting the method of the present application, the added upconversion nanoparticles can be almost completely coated with the high molecular polymer, and the loading amount of the upconversion nanoparticles is increased.
[0045] Step 300: Prepare a second surfactant aqueous solution, and use the second surfactant aqueous solution as the collection phase. Exemplarily, the second surfactant is mixed with deionized water, and then ultrasonically mixed for later use.
[0046] In the process of preparing upconversion fluorescent microspheres, the microspheres formed by shearing the continuous phase against the dispersed phase are collected by the second surfactant aqueous solution, which can prevent the formed upconversion fluorescent microspheres from contacting and merging with each other, thereby further maintaining the stability of the formed upconversion fluorescent microspheres; and is also conducive to the subsequent separation and purification of the upconversion fluorescent microspheres.
[0047] In some embodiments, the second surfactant is one or more of a sodium alkylbenzene sulfonate surfactant, a Triton-X surfactant, a polyvinyl alcohol surfactant, and a Tween surfactant. The mass fraction of the second surfactant aqueous solution is 1% to 20%. Exemplarily, the mass fraction of the second surfactant aqueous solution is 1%, 2%, 5%, 10%, 15%, and 20%.
[0048] In some embodiments, the first surfactant aqueous solution as the continuous phase and the second surfactant aqueous solution as the collecting phase may have the same or different surfactant types and / or concentrations, which are not limited herein.
[0049] Step 400: The continuous phase and the dispersed phase are introduced into the microfluidic chip. Under the action of interfacial tension and flow shear force, the dispersed phase is cut into tiny dispersed droplets and upconversion fluorescent microspheres are obtained. The upconversion fluorescent microspheres obtained are collected using a collection phase located at the outlet of the microfluidic chip.
[0050] In some embodiments, when the continuous phase and the dispersed phase are passed into the microfluidic chip, the flow rate ratio of the continuous phase to the dispersed phase is 1:1 to 50:1. By controlling the flow rate ratio of the continuous phase to the dispersed phase, the particle size of the obtained upconversion fluorescent microspheres can be adjusted. Exemplarily, by controlling the flow rate ratio of the continuous phase to the dispersed phase to be 1:1 to 50:1, the particle size of the obtained upconversion fluorescent microspheres can be maintained at 20 to 50 μm.
[0051] In some embodiments, the method for preparing upconversion fluorescent microspheres further comprises the step of drying the collection phase containing the upconversion fluorescent microspheres.
[0052] Exemplarily, drying the collection phase containing the upconversion fluorescent microspheres comprises: The collection phase containing the up-conversion fluorescent microspheres is allowed to stand at room temperature to evaporate the organic solvent; The upconversion fluorescent microspheres are centrifuged and washed with pure water to remove the surfactant on the surface of the upconversion fluorescent microspheres, and then the washed upconversion fluorescent microspheres are placed in a 60° C. oven to dry.
[0053] In this embodiment, by drying the collection phase containing the upconversion fluorescent microspheres, powdered upconversion fluorescent microspheres can be obtained so that the upconversion fluorescent microspheres can remain stable. It is known that the upconversion fluorescent microspheres can also be kept in the collection phase. The state of the upconversion fluorescent microspheres is determined based on actual applications.
[0054] The upconversion fluorescent microspheres obtained by the microfluidic method of the present invention have good monodispersity, and the upconversion nanoparticles are evenly distributed in the microspheres, so that the obtained upconversion fluorescent microspheres have the advantages of uniform luminescence distribution and stable fluorescence performance; and the upconversion nanoparticles are not limited to being distributed on the surface of the microspheres, so the loading amount of the upconversion nanoparticles can also be increased. In addition, the preparation method of the upconversion fluorescent microspheres of the present invention also has the advantages of simple process and good repeatability.
[0055] The upconversion fluorescent microspheres obtained by the microfluidic method of the present invention can directly encapsulate upconversion nanoparticles while preparing surface functionalized polymer microspheres, without the need to pre-prepare pure polymer microspheres and then couple them, and without the need for a synchronous polymerization process, so the preparation process is faster.
[0056] The present application also discloses upconversion fluorescent microspheres obtained by the above method. Specifically, the upconversion fluorescent microspheres include upconversion nanoparticles and polymers, the upconversion nanoparticles are embedded in the polymers, and the fluorescence intensity distribution of the upconversion fluorescent microspheres can be regulated based on the mass of the upconversion nanoparticles and the mass of the polymers, and when the mass of the upconversion nanoparticles accounts for 0.1-10% of the mass of the polymers, the fluorescence intensity fluctuation value of the upconversion fluorescent microspheres is within ±15%.
[0057] When the mass of the upconversion nanoparticles accounts for a larger proportion of the mass of the polymer, the upconversion nanoparticles are more evenly distributed in the polymer, and the fluorescence intensity fluctuation value of the upconversion fluorescent microspheres is smaller, specifically, the fluorescence intensity fluctuation value of the upconversion fluorescent microspheres can be made within ±5%. When the mass of the upconversion nanoparticles accounts for a smaller proportion of the mass of the polymer, the fluorescence intensity fluctuation value of the upconversion fluorescent microspheres is slightly larger, but overall, the fluorescence intensity fluctuation value of the upconversion fluorescent microspheres can be made within ±15%.
[0058] It can be seen that the fluorescence intensity of the upconversion fluorescent microspheres obtained in the present application is controllable. Specifically, the fluorescence intensity distribution of the upconversion fluorescent microspheres can be obtained based on the mass of the upconversion nanoparticles and the mass of the polymer. The fluorescence intensity distribution of the obtained upconversion fluorescent microspheres is uniform and the fluorescence performance is stable.
[0059] The following is combined with Figures 2 to 8 , the up-conversion fluorescent microspheres and the preparation method thereof provided in the present application are described in detail through specific examples.
[0060] The preparation method of upconversion fluorescent microspheres in this embodiment includes the following steps: To prepare the continuous phase, 0.5 g of Tween 80 was mixed with 10 g of deionized water, vortexed and ultrasonicated for 20 min for later use; To prepare the dispersed phase, 1 ml of a 10 mg / ml dichloromethane solution of NaYF4:Yb,Er nanoparticles was mixed with 200 mg of polystyrene and 2 ml of dichloromethane, vortexed and sonicated for 20 min for later use; Preparation of the collection phase: 1 g of sodium dodecylpropane sulfonate was mixed with 10 g of deionized water, vortexed and sonicated for 20 min for later use; As in the above-mentioned microfluidic device and its construction method, the continuous phase injection rate is 50 μl / min, and the dispersed phase injection rate is 5 μl / min; Collection and drying of upconversion fluorescent microspheres: After the microfluidic device is stabilized, the microspheres are collected for 30 minutes using the collection phase and allowed to stand at room temperature for 48 hours to completely evaporate the dichloromethane in the microspheres. The remaining microspheres are centrifuged at 3500 r / min for 5 minutes, washed three times with pure water, and dried in an oven at 60°C for 48 hours.
[0061] The SEM image of the upconversion fluorescent microspheres obtained by the above steps is as follows: Figure 2 As shown, from Figure 2 It can be seen that the up-conversion fluorescent microspheres obtained in this example have uniform particle sizes, and the particle size of the microspheres is about 35 μm.
[0062] Figure 3 (a) to (c) show the near-infrared fluorescence confocal images of the upconversion fluorescent microspheres of this embodiment, where (a), (b) and (c) are the dark field image, bright field image and composite image, respectively. Figure 3 As can be seen from (a) to (c), the upconversion fluorescent microspheres obtained in this embodiment have substantially the same green fluorescence exhibited by each microsphere, and the luminescence intensity within a single microsphere is highly consistent, that is, the luminescence intensity is uniform between each microsphere and for a single microsphere.
[0063] from Figure 3 (d) and (e) show that the microspheres Figure 3 The fluorescence intensity in the oblique direction in (d) is also uniformly distributed, which further indicates that the fluorescence intensity of each microsphere and inside the microsphere is uniformly distributed.
[0064] Further, from Figure 3(e) It can be seen that when the mass of the upconversion nanoparticles in this embodiment accounts for 5% of the mass of the high molecular weight polymer, the fluorescence intensity fluctuation value of the upconversion fluorescent microspheres is about ±10%, which is a small fluctuation, which further indicates that the upconversion fluorescent microspheres obtained in this embodiment have uniform luminescence intensity.
[0065] The preparation method of upconversion fluorescent microspheres in this embodiment includes the following steps: To prepare the continuous phase, 0.5 g of Triton X-100 was mixed with 10 g of deionized water, vortexed and ultrasonicated for 20 min for later use; To prepare the dispersed phase, 1 ml of a 10 mg / ml dichloromethane solution of NaYF4:Yb,Er nanoparticles was mixed with 200 mg of polystyrene and 2 ml of dichloromethane, vortexed and sonicated for 20 min for later use; Preparation of the collection phase: 1 g of sodium dodecylpropane sulfonate was mixed with 10 g of deionized water, vortexed and sonicated for 20 min for later use; As in the above-mentioned microfluidic device and its construction method, the continuous phase injection rate is 50 μl / min, and the dispersed phase injection rate is 5 μl / min; Collection and drying of upconversion fluorescent microspheres: After the microfluidic device is stabilized, the microspheres are collected for 30 minutes using the collection phase and allowed to stand at room temperature for 48 hours to completely evaporate the dichloromethane in the microspheres. The remaining microspheres are centrifuged at 3500 r / min for 5 minutes, washed three times with pure water, and dried in an oven at 60°C for 48 hours.
[0066] The optical microscope photo of the upconversion fluorescent microspheres obtained by the above steps is as follows: Figure 7 (b) As shown in Figure 7 (b) It can be seen that the up-conversion fluorescent microspheres obtained in this example have uniform particle sizes, which are about 25 μm.
[0067] Figure 4 The fluorescence confocal 3D layer scanning images of the upconversion fluorescent microspheres in this embodiment are shown, wherein (a) to (i) are the luminescence images of different cross sections of the microspheres. Figure 4 It can be seen that the upconversion fluorescent microspheres exhibit uniform green fluorescence intensity on different cross sections, and the green fluorescence between cross sections is basically consistent. Therefore, the upconversion fluorescent microsphere particles obtained in this embodiment have uniform luminescence intensity in the 3D dimension inside the microspheres.
[0068] It should be noted that Figure 4(a), (h) and (i) are the detection results of the cross-sections near the upper and lower ends of the sphere. Since the cross-sectional area near the upper and lower ends of the sphere is smaller, the luminous intensity detected is slightly dim, but it can still be seen that the cross-sections near the upper and lower ends of the sphere are also uniformly illuminated, without any uneven luminescence such as no light, scattered light spots or clusters.
[0069] The preparation method of upconversion fluorescent microspheres in this embodiment includes the following steps: For the preparation of the continuous phase, 0.5 g of PVA was mixed with 10 g of deionized water, vortexed and ultrasonicated for 20 min for later use; To prepare the dispersed phase, 1 ml of a 10 mg / ml dichloromethane solution of NaYF4:Yb,Er nanoparticles was mixed with 200 mg of polystyrene and 2 ml of dichloromethane, vortexed and sonicated for 20 min for later use; Preparation of the collection phase: 1 g of sodium dodecylpropane sulfonate was mixed with 10 g of deionized water, vortexed and sonicated for 20 min for later use; As in the above-mentioned microfluidic device and its construction method, the continuous phase injection rate is 50 μl / min, and the dispersed phase injection rate is 5 μl / min; Collection and drying of upconversion fluorescent microspheres: After the microfluidic device is stabilized, the microspheres are collected for 30 minutes using the collection phase and allowed to stand at room temperature for 48 hours to completely evaporate the dichloromethane in the microspheres. The remaining microspheres are centrifuged at 3500 r / min for 5 minutes, washed three times with pure water, and dried in an oven at 60°C for 48 hours.
[0070] The optical microscope photo of the upconversion fluorescent microspheres obtained by the above steps is as follows: Figure 7 (c), from Figure 7 (c) It can be seen that the upconversion fluorescent microspheres obtained in this example have uniform particle size, which is about 23 μm.
[0071] The preparation method of upconversion fluorescent microspheres in this embodiment includes the following steps: To prepare the continuous phase, 0.5 g of sodium dodecylbenzene sulfonate was mixed with 10 g of deionized water, vortexed and then sonicated for 20 min for later use; To prepare the dispersed phase, 1 ml of a 1 mg / ml NaYF4:Yb,Er nanoparticle dichloromethane solution was mixed with 200 mg of polystyrene and 2 ml of dichloromethane, vortexed evenly, and then sonicated for 20 min for later use; Preparation of the collection phase: 1 g of sodium dodecylpropane sulfonate was mixed with 10 g of deionized water, vortexed and sonicated for 20 min for later use; As in the above-mentioned microfluidic device and its construction method, the continuous phase injection rate is 50 μl / min, and the dispersed phase injection rate is 5 μl / min; Collection and drying of upconversion fluorescent microspheres: After the microfluidic device is stabilized, the microspheres are collected for 30 minutes using the collection phase and allowed to stand at room temperature for 48 hours to completely evaporate the dichloromethane in the microspheres. The remaining microspheres are centrifuged at 3500 r / min for 5 minutes, washed three times with pure water, and dried in an oven at 60°C for 48 hours.
[0072] The SEM image of the upconversion fluorescent microspheres obtained by the above steps is as follows: Figure 5 As shown, from Figure 5 It can be seen that the up-conversion fluorescent microspheres obtained in this example have uniform particle sizes, and the particle size of the microspheres is about 23 μm.
[0073] The preparation method of upconversion fluorescent microspheres in this embodiment includes the following steps: To prepare the continuous phase, 0.5 g of sodium dodecylbenzene sulfonate was mixed with 10 g of deionized water, vortexed and then sonicated for 20 min for later use; Preparation of dispersed phase: 1.2 ml of 1 mg / ml NaYF4:Yb,Er nanoparticles in chloroform solution was mixed with 90 mg of polymethyl methacrylate and 1.8 ml of chloroform, vortexed and sonicated for 20 min for later use; Preparation of the collection phase: 1 g of sodium dodecylpropane sulfonate was mixed with 10 g of deionized water, vortexed and sonicated for 20 min for later use; As in the above-mentioned microfluidic device and its construction method, the continuous phase injection rate is 30 μl / min, and the dispersed phase injection rate is 5 μl / min; Collection and drying of upconversion fluorescent microspheres: After the microfluidic device is stabilized, the microspheres are collected for 30 minutes using the collection phase and allowed to stand at room temperature for 48 hours to completely evaporate the dichloromethane in the microspheres. The remaining microspheres are centrifuged at 3500 r / min for 5 minutes, washed three times with pure water, and dried in an oven at 60°C for 48 hours.
[0074] The optical microscope photo of the upconversion fluorescent microspheres obtained by the above steps is as follows: Figure 8 As shown in (a), from Figure 8 (a) It can be seen that the upconversion fluorescent microspheres obtained in this example have uniform particle size, which is about 45 μm.
[0075] The preparation method of upconversion fluorescent microspheres in this embodiment includes the following steps: To prepare the continuous phase, 0.5 g of sodium dodecylbenzene sulfonate was mixed with 10 g of deionized water, vortexed and then sonicated for 20 min for later use; To prepare the dispersed phase, 1.2 ml of a 1 mg / ml chloroform solution of NaYF4:Yb,Er nanoparticles was mixed with 90 mg of polymethyl methacrylate, 30 mg of poly(methyl methacrylate-co-methacrylic acid), and 1.8 ml of chloroform, vortexed until uniform, and then sonicated for 20 min for later use; Preparation of the collection phase: 1 g of sodium dodecylpropane sulfonate was mixed with 10 g of deionized water, vortexed and sonicated for 20 min for later use; As in the above-mentioned microfluidic device and its construction method, the continuous phase injection rate is 30 μl / min, and the dispersed phase injection rate is 5 μl / min; Collection and drying of upconversion fluorescent microspheres: After the microfluidic device is stabilized, the microspheres are collected for 30 minutes using the collection phase and allowed to stand at room temperature for 48 hours to completely evaporate the dichloromethane in the microspheres. The remaining microspheres are centrifuged at 3500 r / min for 5 minutes, washed three times with pure water, and dried in an oven at 60°C for 48 hours.
[0076] The SEM image of the upconversion fluorescent microspheres obtained by the above steps is as follows: Figure 6 As shown, from Figure 6 It can be seen that the up-conversion fluorescent microspheres obtained in this example have uniform particle sizes, which are about 25 μm.
[0077] In addition, in this embodiment, polymethyl methacrylate and poly (methyl methacrylate-co-methacrylic acid) are used together as microsphere matrix polymers to directly form surface functionalized microspheres. It can be seen that when the upconversion fluorescent microspheres are obtained by the microfluidic method, the upconversion nanoparticles can be directly encapsulated in the surface functionalized polymer microspheres while preparing the surface functionalized polymer microspheres, without the need to prepare pure polymer microspheres in advance and then couple them, and without the need for a synchronous polymerization process, and the preparation process is faster.
[0078] The present application also examined the effect of using different surfactant solutions as continuous phases on the dispersibility and particle size of upconversion fluorescent microspheres. Figure 7 (a) to (d) in the figure show the microscopic images of the upconversion fluorescent microspheres obtained in Examples 1 to 4, respectively. Figure 7 It can be seen that when the aqueous solution of Tween80 is used as the continuous phase, the particle size of the obtained upconversion fluorescent microspheres is larger, and when the aqueous solution of TritonX-100, PVA and sodium dodecylbenzenesulfonate is used as the continuous phase, the particle size of the obtained upconversion fluorescent microspheres is smaller; in addition, when the aqueous solution of PVA and sodium dodecylbenzenesulfonate is used as the continuous phase, the uniformity of the dispersion of the obtained upconversion fluorescent microspheres is better than that using the aqueous solution of Tween80 and the aqueous solution of TritonX-100 as the continuous phase.
[0079] Therefore, the continuous phase of the present application is preferably an aqueous solution of a PVA-based surfactant or an aqueous solution of a sodium dodecylbenzene sulfonate-based surfactant.
[0080] The present application also examines the effect of using different polymers as dispersants on the dispersibility and particle size of upconversion fluorescent microspheres. Figure 8 (a) and (b) in the figure respectively show the microscopic images of the upconversion fluorescent microspheres obtained in Examples 5 and 6. Figure 8 It can be seen that when polymethyl methacrylate is used as the dispersed phase, the particle size of the obtained upconversion fluorescent microspheres is larger, and when polymethyl methacrylate and poly(methyl methacrylate-co-methacrylic acid) are used as the dispersed phase, the particle size of the obtained upconversion fluorescent microspheres is smaller; in addition, when polymethyl methacrylate and poly(methyl methacrylate-co-methacrylic acid) are used as the dispersed phase, the uniformity of the dispersion of the obtained upconversion fluorescent microspheres is better than that when polymethyl methacrylate is used as the dispersed phase.
[0081] Therefore, the high molecular weight polymer of the dispersed phase in the present application is preferably a block copolymer.
[0082] Comparative Example The preparation method of the upconversion fluorescent microspheres in this comparative example comprises the following steps: To prepare the continuous phase, 0.5 g of Tween 80 was mixed with 10 g of deionized water, vortexed and ultrasonicated for 20 min for later use; To prepare the dispersed phase, 1 ml of a 10 mg / ml dichloromethane solution of NaYF4:Yb,Er nanoparticles was mixed with 200 mg of polystyrene and 2 ml of dichloromethane, vortexed and sonicated for 20 min for later use; Preparation of the collection phase: 1 g of sodium dodecylpropane sulfonate was mixed with 10 g of deionized water, vortexed and sonicated for 20 min for later use; As in the above-mentioned microfluidic device and its construction method, the continuous phase injection rate is 300 μl / min, and the dispersed phase injection rate is 5 μl / min; Collection and drying of upconversion fluorescent microspheres: After the microfluidic device is stabilized, the microspheres are collected for 30 minutes using the collection phase and allowed to stand at room temperature for 48 hours to completely evaporate the dichloromethane in the microspheres. The remaining microspheres are centrifuged at 3500 r / min for 5 minutes, washed three times with pure water, and dried in an oven at 60°C for 48 hours.
[0083] The SEM image of the upconversion fluorescent microspheres obtained by the above steps is as follows: Fig. 9 As shown, from Fig. 9 It can be seen that the upconversion fluorescent microspheres obtained in this comparative example are large and small, and the particle size distribution is uneven. The largest particle size of the microsphere can reach about 60 μm, and the smallest particle size of the microsphere is less than 10 μm.
[0084] It should be noted that, in this article, the terms "include", "comprises" or any other variations thereof are intended to cover non-exclusive inclusion, so that a process, method, article or device including a series of elements includes not only those elements, but also other elements not explicitly listed, or also includes elements inherent to such process, method, article or device. In the absence of further restrictions, an element defined by the sentence "comprises a ..." does not exclude the existence of other identical elements in the process, method, article or device including the element.
[0085] In addition, it should be noted that the scope of the methods and devices in the embodiments of the present application is not limited to performing functions in the order shown or discussed, and may also include performing functions in a substantially simultaneous manner or in a reverse order according to the functions involved. For example, the described methods may be performed in an order different from that described, and various steps may be added, omitted, or combined. In addition, features described with reference to certain examples may be combined in other examples.
[0086] The above description is only a specific implementation mode of the present invention, but the protection scope of the present invention is not limited thereto. Any technician familiar with the technical field can easily think of changes or substitutions within the technical scope disclosed by the present invention, which should be covered by the protection scope of the present invention.
Claims
1. A method for preparing upconversion fluorescent microspheres, characterized in that: The upconversion fluorescent microspheres are prepared by a microfluidic method, and the preparation method of the upconversion fluorescent microspheres includes the following steps: preparing a first surfactant aqueous solution, with the first surfactant aqueous solution serving as a continuous phase; Mixing upconversion nanoparticles, high molecular polymers, and an organic solution to obtain an upconversion nanoparticle-high molecular polymer organic solution, wherein the upconversion nanoparticle-high molecular polymer organic solution is a dispersed phase; preparing a second surfactant aqueous solution, and using the second surfactant aqueous solution as a collection phase; The continuous phase and the dispersed phase are introduced into a microfluidic chip, and under the action of interfacial tension and flow shear force, the dispersed phase is cut into tiny dispersed droplets to obtain upconversion fluorescent microspheres, and the obtained upconversion fluorescent microspheres are collected by the collecting phase located at the outlet of the microfluidic chip.
2. The method for preparing upconversion fluorescent microspheres according to claim 1, characterized in that: When the continuous phase and the dispersed phase are passed into the microfluidic chip, the flow rate ratio of the continuous phase to the dispersed phase is 1:1 to 50:
1.
3. The method for preparing upconversion fluorescent microspheres according to claim 1 or 2, characterized in that: The preparation of the first surfactant aqueous solution comprises the following steps: The first surfactant is mixed with deionized water, and then ultrasonically mixed for later use.
4. The method for preparing upconversion fluorescent microspheres according to claim 3, characterized in that: The first surfactant is one or more of a sodium alkylbenzene sulfonate surfactant, a Triton-X surfactant, a polyvinyl alcohol surfactant and a Tween surfactant, and the mass fraction of the first surfactant aqueous solution is 1% to 20%.
5. The method for preparing upconversion fluorescent microspheres according to claim 1 or 2, characterized in that: The preparation of the upconversion nanoparticle-polymer organic solution comprises the following steps: The upconversion nanoparticles, the high molecular polymer and the organic solution are mixed, and ultrasonically mixed and then set aside; Alternatively, the upconversion nanoparticles are dissolved in an organic solution to obtain an upconversion nanoparticle organic solution, and then the upconversion nanoparticle organic solution is mixed with a high molecular polymer, or the upconversion nanoparticle organic solution is mixed with the high molecular polymer and the organic solution, and then ultrasonically mixed for later use; Alternatively, the polymer is dissolved in an organic solution to obtain a polymer organic solution, and then the polymer organic solution is mixed with upconversion nanoparticles, or the polymer organic solution is mixed with upconversion nanoparticles and the organic solution, and then ultrasonically mixed for later use.
6. The method for preparing upconversion fluorescent microspheres according to claim 5, characterized in that: When preparing upconversion nanoparticle-polymer organic solution, the amounts of upconversion nanoparticles, polymer and organic solution meet the following requirements: the mass of polymer accounts for 1%~20% of the mass of dispersed phase; the mass of upconversion nanoparticles accounts for 0.1~10% of the mass of polymer.
7. The method for preparing upconversion fluorescent microspheres according to claim 5, characterized in that: The up-conversion nanoparticles are NaYF4:Yb, Er nanoparticles or NaYF4:Yb, Tm nanoparticles; The high molecular polymer is one or more of polymethyl methacrylate, polymethacrylic acid, polyacrylic acid, polystyrene, polyglycolic acid, polyisopropylacrylamide or derivatives thereof; The organic solvent is one or more of dichloromethane, chloroform, tetrahydrofuran, toluene, cyclohexane, and N,N'-dimethylacrylamide.
8. The method for preparing upconversion fluorescent microspheres according to claim 1 or 2, characterized in that: The preparation of the second surfactant aqueous solution comprises the following steps: The second surfactant is mixed with deionized water, and then ultrasonically mixed for later use.
9. The method for preparing upconversion fluorescent microspheres according to claim 8, characterized in that: The second surfactant is one or more of a sodium alkylbenzene sulfonate surfactant, a Triton-X surfactant, a polyvinyl alcohol surfactant, and a Tween surfactant, and the mass fraction of the second surfactant aqueous solution is 1% to 20%.
10. An upconversion fluorescent microsphere prepared by the preparation method of upconversion fluorescent microspheres according to any one of claims 1 to 9, characterized in that: The invention comprises up-conversion nanoparticles and a high molecular polymer, wherein the up-conversion nanoparticles are embedded in the high molecular polymer, and The fluorescence intensity distribution of the upconversion fluorescent microspheres can be regulated based on the mass of the upconversion nanoparticles and the mass of the high molecular polymer. When the mass of the upconversion nanoparticles accounts for 0.1-10% of the mass of the high molecular polymer, the fluorescence intensity fluctuation value of the upconversion fluorescent microspheres is within ±15%.