CN1693412A - Process for preparing fluorescent nano mciroball - Google Patents
Process for preparing fluorescent nano mciroball Download PDFInfo
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- CN1693412A CN1693412A CN 200510026434 CN200510026434A CN1693412A CN 1693412 A CN1693412 A CN 1693412A CN 200510026434 CN200510026434 CN 200510026434 CN 200510026434 A CN200510026434 A CN 200510026434A CN 1693412 A CN1693412 A CN 1693412A
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- polyelectrolyte
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Abstract
A process for preparing the fluorescent nanosphere includes such steps as using the microsphere of polymer of SiO2 as the kernel, alternatively assembling multiple layers of polyelectrolyte containing Cd or Zn ions to the surface of said microsphere layer by layer, introducing S, Se, or Te ions to it, and in-situ reaction.
Description
Technical field
The present invention relates to a kind of preparation method who is used for the fluorescent microsphere of biological detection, relate in particular to preparation method a kind of polymer-based carbon or silicon dioxide base, that have fluorescent nano mciroball.
Background technology
Fluorescent microsphere refers generally to microsphere surface and indicates the microballoon that fluorescent substance (comprise surface coat) or microsphere inner structure contain fluorescent substance (as embedding or polymerization).By means of fluorometric analysis, fluorescent microsphere can utilize the contained specific groups of this microsphere surface usually, be used to measure the existence that specificity bonded biomolecules is arranged with it as probe, be used to proofread and correct the fluorescent base detection system as correction agent, and be used to follow the trail of the fluid flow that contains fluorescent microsphere as tracer agent.
What traditional fluorescent substance adopted usually is organic fluorescent dye, for example, and fluorescein, rhodamine, phycoerythrin and similar material.But these fluorescence dyes are subject to the restriction of physics and chemical factor: (1) dyes in different colors it excite light wavelength different, two or more fluorescent markers that cause having different excitation wavelengths simultaneously need the compound excitation light source, and the method for using many fluorescence dyes has been increased cost and complicacy.(2) organic dye is being exposed to exciting light following time for a long time, and its fluorescence intensity can go down.This phenomenon depends on the time length that excites light intensity and illumination photobleaching.(3) to change into non-fluorescent substance be irreversible to organic dye.And the degraded product of dye substance is an organism, may influence the biological detection process.(4) there is the spectrum crossover between the different dyes.This part is because the wide emmission spectrum of organic dye and present crossover near the spectrum of tail region.Almost there is not low-molecular-weight dyestuff that big Stokes displacement and high fluorescence output (the Stokes displacement is defined as emission and absorbs the spacing of peak position) are arranged.
In order to overcome the above-mentioned deficiency that contains the organic dye fluorescent microsphere, in recent years, the fluorescent microsphere that contains the semiconductor nano grain has had bibliographical information.Their advantage is to have an excitation wavelength spectrum, and fluorescence and the emmission spectrum that can launch high quantum yield have big Stokes displacement.Chinese patent CN 1389539A (2003) discloses a kind ofly has the fluorescence nano grain of different functional groups to be compound to the nanoparticle fluorescent microsphere preparation method of inorganic silicon dioxide or organic polymer microballoon finishing.U.S. Pat 6,680,211 (2004) disclose a kind of swelling character that utilizes some polymkeric substance to have, and the fluorescence nano grain is filled in the swollen polymkeric substance hole, then go swelling to obtain the preparation method of the polymer-based carbon fluorescent microsphere of physically trapping.Yet these methods are difficult to accomplish uniform fluorescence nano grain content in uniform microballoon distribution and the microballoon, are unfavorable for quantitative fluorescence analysis accurately.
Utilize self-assembly layer by layer (layer-by-layer self-assembly) technology to construct functional microsphere and obtained considerable progress recently with nucleocapsid structure.The preparation of the hud typed microballoon of this class is a nuclear with charged organic or inorganic microballoon normally, the successively polyelectrolyte and/or the nanoparticle of adsorption zone opposite charges on its surface.The thickness of the shell of the hud typed microballoon that obtains can be controlled by the number of times that changes sorption cycle at an easy rate, and the size and shape of shell can be pre-determined by the yardstick of used nuclear simultaneously.The hud typed microballoon that obtains like this has and the special property of nuclear significant difference (as different chemical constitutions, satisfactory stability, high specific surface area, and different magnetic and optical property).U.S. Pat 6,833,192 (2004) have just adopted layer-by-layer, have alternately assembled multi-layer polyelectrolyte and/or nanoparticle on the crystal particles surface, have formed core-shell type complex particle, and wherein nanoparticle can be the fluorescence nano grain.
Usually the fluorescence nano grain adopts the II-VI family semiconductor nano grain of high fluorescence yield, has the nanoparticle of nucleocapsid structure as CdS, CdSe, ZnSe, CdTe and CdSe/CdS and CdSe/ZnS etc.Yet directly at these nanoparticles of aqueous phase self-assembly layer by layer, because of strong Coulomb repulsion effect has limited the nanoparticle adsorptive capacity.
Summary of the invention
At the deficiencies in the prior art, the invention provides a kind of new method for preparing fluorescent nano mciroball, to prepare the equally distributed fluorescent nano mciroball of fluorescence nano grain.
The method for preparing fluorescent nano mciroball that the present invention proposes is to be nuclear with polymkeric substance or silicon dioxide microsphere, adopt layer-by-layer, alternately assemble the multi-layer polyelectrolyte that complexing has cadmium ion or zine ion at microsphere surface, then introduce sulfonium ion, plasma selenium or tellurium ion, reaction in obtains uniform fluorescent nano mciroball.
The size of microballoon is at 50nm～10 μ m, and the size of fluorescence nano grain is at 1～15nm, and its fluorescence peak position is in 400～650nm scope.
The preparation method of fluorescent nano mciroball of the present invention comprises the steps:
1. microsphere surface self-assembly complexing layer by layer M
2+Multi-layer polyelectrolyte (M is the elements such as Cd, Zn of II family):
Selecting equally distributed polymkeric substance of monodispersed particle diameter or silicon dioxide microsphere for use is template core, and polymer microballoon can be selected for use as polystyrene, polymethyl acrylate, melamino-formaldehyde (MF) etc.The particle diameter of microballoon is less than 10 μ m, and particle size range is 50nm～5 μ m preferably, and better particle size range is 100nm～500nm.
Polyelectrolyte to be assembled: cationic polyelectrolyte is selected polymine (polyethyleneimine for use, PEI), diallyl dimethyl ammoniumchloride (poly (diallyldimethylammonium chloride), PDDA), the PAH hydrochloride (poly (allylamine hydrochloride), PAH); Anionic polyelectrolyte select for use poly-4-styrene sulfonate (poly (styrenesulfonate), PSS), polyvinyl sulfuric acid salt (poly (vinylsulfate)), polyacrylic acid (poly (acrylic acid), PAA).
M
2+The source: the cadmium ion compound is selected Cd (NO for use
3)
2, CdCl
2, Cd (ClO
4)
2, Cd (CH
3COO)
2Deng the aqueous solution; The zine ion compound is selected Zn (NO for use
3)
2, Zn (CH
3COO)
2Deng the aqueous solution.
Preparation process:
I) with positive and negative ion polyelectrolyte respectively and soluble salt, be mixed with solution A and B as NaCl or KCl etc.Making the concentration of polyelectrolyte in the solution A (and B) is 0.1～5g/L, and the concentration of salt is 0.05～0.6mol/L.
Ii) be the M of 0.001～0.1mol/L with concentration
2+Solution mixes with polyelectrolyte A (and/or B) solution respectively, vibration absorption 15min-12h, and preferably 1～6h makes ion and polyelectrolyte functional group produce complexing, obtains complexing M
2+Polyelectrolyte A
c(and B
c) solution.M
2+The concentration of solution is high more, and the size of the nanoparticle on the fluorescent microsphere that obtains at last is big more, the corresponding red shift of fluorescence peak position.
Iii) will examine the microballoon dispersed with stirring in solution A
cIn.Stir 5～60min under the room temperature, preferably 10～30min forms template microsphere absorption complexing M
2+Water solubility of polyelectrolyte suspension, complexing M
2+Polyelectrolyte amount must greater than on template microsphere absorption a unimolecular layer amount.The mass ratio of template microsphere and suspension is 0.5～5%, is preferably 1～2%.Use centrifugally then, remove supernatant liquor, with deionized water washing/centrifugal/disperse again 3～5 times.
Iv) step I is obtained the settling redispersion in solution B in ii)
cIn, and iii) ensuing set by step operation is carried out once again.
V) step I is obtained the settling redispersion in solution A in v)
cIn, and iii) ensuing set by step operation is carried out once again.So circulation repeatedly obtains the complex microsphere suspension of template microsphere pipe absocped with polyelectrolyte on surface multilayer film.
2. be assembled in the complexing M of microsphere surface
2+Multi-layer polyelectrolyte with contain X
2-Aqueous solution reaction in (X is elements such as S, Se, the Te of VI family):
Contain X
2-The aqueous solution: the sulphur source can be selected thioacetamide (TAA), Na for use
2The S aqueous solution; Selenium source can be selected NaHSe, Na for use
2SeO
3The aqueous solution; The tellurium source can be selected NaHTe, Na for use
2The Te aqueous solution.
The complex microsphere suspension that step 1 is obtained is stirred and heated to 40～100 ℃, with the X that contains of 0.001～0.1mol/L of preparing
2-The aqueous solution dropwise splash in the suspension and react X
2-With M
2+Molar ratio remain on 0.5～1.5, be preferably 1.1～1.2.Reaction times is 15min～1h.Behind the stopped reaction, with the fluorescent microsphere that generates with deionized water washing/centrifugal/disperse again 3～5 times.
Beneficial effect
Preparation method of the present invention has the following advantages: (1) has selected for use homogeneous polymer or silicon dioxide microsphere to make template, and therefore the fluorescent nano mciroball that obtains is more even; (2) the fluorescent nano mciroball size can be made template by the microballoon of selecting different size for use, and the kind and the number of plies of absorption polyelectrolyte controlled, thus the luminous intensity of regulating fluorescent microsphere; (3) nanoparticle that adopts the method for reaction in to make is subjected to the polyelectrolyte protection, can be dispersed in the microballoon top layer, is difficult for causing reunion; (4) can be at the different M of same microsphere surface assembling complexing
2+Polyelectrolyte, obtain indicating the fluorescent microsphere of multiple nanoparticle; (5) can be by regulating M
2+With the ratio of polyelectrolyte complex, the size of control microballoon top layer fluorescence nano grain, thus regulate the fluorescent emission peak position; (6) Biao Mian polyelectrolyte has stronger functionally, and better dispersed to the various aqueous solution, the active substances of can delivering a child as antibody, antigen, enzyme, protein or nucleic acid, thereby can be realized the purpose that bioluminescence detects.
Embodiment
Further illustrate content of the present invention below in conjunction with embodiment, but these embodiment do not limit protection scope of the present invention.
Embodiment 1
Step (1) is with PSS, and PDDA is mixed with solution A and B with NaCl respectively.The concentration of polyelectrolyte is 1g/L in the solution A (and B), and the concentration of salt is 0.5mol/L.
Step (2) is got the Cd that concentration is 0.01mol/L respectively
2+Solution 50mL joins in the polyelectrolyte PSS solution that 50mL concentration is 1g/L, gets the Zn of concentration 0.01mol/L
2+Solution 50mL joins in the polyelectrolyte PDDA solution that 50mL concentration is 1g/L, and vibration 4h makes ion and polyelectrolyte functional group produce complexing, obtains complexing Cd
2+Polyelectrolyte PSS-Cd
2+Solution and complexing Zn
2+Polyelectrolyte PDDA-Zn
2+Solution.
Step (3) is that melamino-formaldehyde (MF) the microballoon dispersed with stirring of 2 μ m is at 100mLPSS-Cd with the 2g median size
2+In the solution.Stir 30min under the room temperature, form MF microballoon absorption polyelectrolyte PSS-Cd
2+Water-soluble suspension.Use centrifugally then, remove supernatant liquor, with deionized water washing/centrifugal/disperse again 4 times.
Step (4) will obtain the settling redispersion at 100mL PDDA-Zn in the step (3)
2+In the solution, and (3) ensuing operation is carried out once more set by step.
Step (5) will obtain the settling redispersion at solution PSS-Cd in the step (4)
2+In, and (3) ensuing operation is carried out once more set by step.So circulation is 4 times, obtains microsphere surface first and third, five, seven layers and is absorption PSS-Cd
2+Film, second, four, six, eight layer are absorption PDDA-Zn
2+The complex microsphere suspension of film.
Step (6) places 70 ℃ of stirring in water bath with complex microsphere suspension, thioacetamide (TAA) solution of the 0.01mol/L for preparing is dropwise splashed in the suspension react S again
2-With Cd
2+And S
2-With Zn
2+Ratio remain on 1.1.Behind the reaction 0.5h, solution becomes oyster, stopped reaction.With the fluorescent microsphere that generates CdS/ZnS with deionized water washing/centrifugal/disperse again 4 times.Obtain MF base fluorescent microsphere.Wherein the mean sizes of the nanoparticle on the fluorescent microsphere is 9nm, and its fluorescence peak position is 462nm.
Embodiment 2
With median size is that the polystyrene microsphere of 200nm replaces the MF microballoon among the embodiment 1, is the 100mL PDDA-Zn among the PAH solution replacement embodiment 1 of 1g/L with 100mL concentration
2+Solution, with " thioacetamide of 0.01mol/L (TAA) solution " in " the NaHSe solution of 0.005mol/L " step of replacing (6), all the other repeat the 2nd～6 step among the embodiment 1, obtain generating the polystyrene-based fluorescent nano mciroball of CdSe at last, the mean sizes that wherein obtains the nanoparticle on the fluorescent microsphere is at 4nm, and its fluorescence peak position is at 552nm.
Embodiment 3
With median size is the SiO of 100nm
2Microballoon replaces the polystyrene microsphere among the embodiment 2, all the other repeat the 2nd～6 step among embodiment 1 with " the NaHSe solution of 0.005mol/L " among " the NaHTe solution of 0.008mol/L " replacement embodiment 2, obtain generating the silicon-dioxide-substrate fluorescence nano mciroball of CdTe at last, the mean sizes that wherein obtains the nanoparticle on the fluorescent microsphere is at 5nm, and its fluorescence peak position is at 590nm.
Claims (9)
1, a kind of preparation method of fluorescent nano mciroball, described method are to be nuclear with polymer microballoon or silicon dioxide microsphere, it is characterized in that, at first will have the polyelectrolyte of opposite charges, respectively with M
2+Ion complexation; Adopt layer-by-layer again, the polyelectrolyte of the oppositely charged that complexing is good successively alternate group is contained in microsphere surface, and its number of plies is unrestricted; Then introduce X
2-Ion, carry out reaction in and obtain uniform fluorescent nano mciroball;
Wherein, M
2+Ion is the Cd of II family
2+, Zn
2+In one or both, X
2-Ion is the S of VI family
2-, Se
2-, Te
2-In a kind of.
2, the preparation method of fluorescent nano mciroball as claimed in claim 1, it is characterized in that the described polyelectrolyte that has opposite charges is: cationic polyelectrolyte is selected a kind of in polymine, diallyl dimethyl ammoniumchloride, the PAH hydrochloride for use; Anionic polyelectrolyte is selected a kind of in poly-4-styrene sulfonate, polyvinyl sulfuric acid salt, the polyacrylic acid for use; M
2+The source is: the cadmium ion compound is selected Cd (NO for use
3)
2, CdCl
2, Cd (ClO
4)
2, Cd (CH
3COO)
2The aqueous solution in a kind of, the zine ion compound is selected Zn (NO for use
3)
2, Zn (CH
3COO)
2The aqueous solution in a kind of;
The described polyelectrolyte that has an opposite charges respectively with M
2+In the ion complexation process, positive and negative ion polyelectrolyte is mixed with solution with the aqueous solution of soluble salt respectively, in the polyelectrolyte solution for preparing, the concentration of polyelectrolyte is 0.1～5g/L, and the concentration of salts solution is 0.05～0.6mol/L;
With the polyelectrolyte solution and the concentration that prepare is the M of 0.001～0.1mol/L
2+Solution mixes, and vibration absorption 15min-12h obtains complexing M
2+Polyelectrolyte solution.
3, the preparation method of fluorescent nano mciroball as claimed in claim 1 is characterized in that, the particle diameter of microballoon is less than 10 μ m.
4, the preparation method of fluorescent nano mciroball as claimed in claim 1 is characterized in that, is assembled in the process of microsphere surface template microsphere and complexing M at the described polyelectrolyte that complexing is good
2+The mass ratio of polyelectrolyte solution be 0.5～5%.
5, the preparation method of fluorescent nano mciroball as claimed in claim 1 is characterized in that, at described introducing X
2-Ion, carry out in the process of reaction in, will be assembled in the complexing M of microsphere surface
2+Multi-layer polyelectrolyte with contain X
2-Aqueous solution reaction in, wherein, contain X
2-The aqueous solution be: the sulphur source is selected thioacetamide, Na for use
2A kind of in the aqueous solution of S; Selenium source is selected NaHSe, Na for use
2SeO
3The aqueous solution in a kind of; The tellurium source is selected NaHTe, Na for use
2A kind of in the aqueous solution of Te; Contain X
2-Concentration of aqueous solution be 0.001～0.1mol/L, X
2-With M
2+Molar ratio be 0.5～1.5, the reaction times is 15min～1h.
6, the preparation method of fluorescent nano mciroball as claimed in claim 3 is characterized in that, the particle diameter of microballoon is 50nm～5 μ m.
7, the preparation method of fluorescent nano mciroball as claimed in claim 6 is characterized in that, the particle diameter of microballoon is 100nm～500nm.
8, the preparation method of fluorescent nano mciroball as claimed in claim 4 is characterized in that, template microsphere and complexing M
2+The mass ratio of polyelectrolyte solution be 1～2%.
9, the preparation method of fluorescent nano mciroball as claimed in claim 5 is characterized in that, X
2-With M
2+Mol ratio be 1.1～1.2.
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| CN101914378A (en) * | 2010-08-06 | 2010-12-15 | 陕西师范大学 | SiO2@ZrO2@Y2O3:Eu3+ fluorescent powder with core-shell structure and preparation method thereof |
| CN102060948A (en) * | 2010-12-03 | 2011-05-18 | 黑龙江大学 | Method for preparing polymer fluorescent nano particle |
| CN102320612A (en) * | 2011-05-26 | 2012-01-18 | 东北师范大学 | Preparation method and application of fluorescence mesoporous silica nano-particle |
| CN102391875A (en) * | 2011-08-30 | 2012-03-28 | 苏州大学 | Quantum dot-polyelectrolyte fluorescent nano complex and application thereof |
| CN102423670A (en) * | 2011-08-09 | 2012-04-25 | 中国科学院合肥物质科学研究院 | Raman coding microspheres and preparation method thereof |
| CN103992449A (en) * | 2014-05-26 | 2014-08-20 | 江苏大学 | Preparation method of core-shell fluorescent microsphere |
| CN110105697A (en) * | 2019-05-09 | 2019-08-09 | 南京林业大学 | A method of preparing polyelectrolyte complex material |
| CN110846039A (en) * | 2019-10-29 | 2020-02-28 | 浙江传媒学院 | SiO (silicon dioxide)2Preparation method of @ CdTe @ Au nano composite material |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN101914378A (en) * | 2010-08-06 | 2010-12-15 | 陕西师范大学 | SiO2@ZrO2@Y2O3:Eu3+ fluorescent powder with core-shell structure and preparation method thereof |
| CN101914378B (en) * | 2010-08-06 | 2013-03-20 | 陕西师范大学 | SiO2@ZrO2@Y2O3:Eu3+ fluorescent powder with core-shell structure and preparation method thereof |
| CN102060948A (en) * | 2010-12-03 | 2011-05-18 | 黑龙江大学 | Method for preparing polymer fluorescent nano particle |
| CN102060948B (en) * | 2010-12-03 | 2012-07-04 | 黑龙江大学 | Method for preparing polymer fluorescent nano particle |
| CN102320612A (en) * | 2011-05-26 | 2012-01-18 | 东北师范大学 | Preparation method and application of fluorescence mesoporous silica nano-particle |
| CN102423670B (en) * | 2011-08-09 | 2014-01-01 | 中国科学院合肥物质科学研究院 | Raman coding microspheres and preparation method thereof |
| CN102423670A (en) * | 2011-08-09 | 2012-04-25 | 中国科学院合肥物质科学研究院 | Raman coding microspheres and preparation method thereof |
| CN102391875A (en) * | 2011-08-30 | 2012-03-28 | 苏州大学 | Quantum dot-polyelectrolyte fluorescent nano complex and application thereof |
| CN102391875B (en) * | 2011-08-30 | 2013-04-03 | 苏州大学 | Quantum dot-polyelectrolyte fluorescent nano complex and application thereof |
| CN103992449A (en) * | 2014-05-26 | 2014-08-20 | 江苏大学 | Preparation method of core-shell fluorescent microsphere |
| CN110105697A (en) * | 2019-05-09 | 2019-08-09 | 南京林业大学 | A method of preparing polyelectrolyte complex material |
| CN110846039A (en) * | 2019-10-29 | 2020-02-28 | 浙江传媒学院 | SiO (silicon dioxide)2Preparation method of @ CdTe @ Au nano composite material |
| CN110846039B (en) * | 2019-10-29 | 2020-10-30 | 浙江传媒学院 | SiO (silicon dioxide)2Preparation method of @ CdTe @ Au nano composite material |
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