CN109306539A - A kind of 3D conduction cell culturing bracket and preparation method thereof - Google Patents
A kind of 3D conduction cell culturing bracket and preparation method thereof Download PDFInfo
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- D01F6/00—Monocomponent artificial filaments or the like of synthetic polymers; Manufacture thereof
- D01F6/28—Monocomponent artificial filaments or the like of synthetic polymers; Manufacture thereof from copolymers obtained by reactions only involving carbon-to-carbon unsaturated bonds
- D01F6/38—Monocomponent artificial filaments or the like of synthetic polymers; Manufacture thereof from copolymers obtained by reactions only involving carbon-to-carbon unsaturated bonds comprising unsaturated nitriles as the major constituent
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- C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
- C12N5/0062—General methods for three-dimensional culture
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- D01—NATURAL OR MAN-MADE THREADS OR FIBRES; SPINNING
- D01F—CHEMICAL FEATURES IN THE MANUFACTURE OF ARTIFICIAL FILAMENTS, THREADS, FIBRES, BRISTLES OR RIBBONS; APPARATUS SPECIALLY ADAPTED FOR THE MANUFACTURE OF CARBON FILAMENTS
- D01F1/00—General methods for the manufacture of artificial filaments or the like
- D01F1/02—Addition of substances to the spinning solution or to the melt
- D01F1/10—Other agents for modifying properties
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- D06M10/00—Physical treatment of fibres, threads, yarns, fabrics, or fibrous goods made from such materials, e.g. ultrasonic, corona discharge, irradiation, electric currents, or magnetic fields; Physical treatment combined with treatment with chemical compounds or elements
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- D06M11/00—Treating fibres, threads, yarns, fabrics or fibrous goods made from such materials, with inorganic substances or complexes thereof; Such treatment combined with mechanical treatment, e.g. mercerising
- D06M11/73—Treating fibres, threads, yarns, fabrics or fibrous goods made from such materials, with inorganic substances or complexes thereof; Such treatment combined with mechanical treatment, e.g. mercerising with carbon or compounds thereof
- D06M11/74—Treating fibres, threads, yarns, fabrics or fibrous goods made from such materials, with inorganic substances or complexes thereof; Such treatment combined with mechanical treatment, e.g. mercerising with carbon or compounds thereof with carbon or graphite; with carbides; with graphitic acids or their salts
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- D06M15/00—Treating fibres, threads, yarns, fabrics, or fibrous goods made from such materials, with macromolecular compounds; Such treatment combined with mechanical treatment
- D06M15/19—Treating fibres, threads, yarns, fabrics, or fibrous goods made from such materials, with macromolecular compounds; Such treatment combined with mechanical treatment with synthetic macromolecular compounds
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- C12N2533/00—Supports or coatings for cell culture, characterised by material
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- D06M2101/00—Chemical constitution of the fibres, threads, yarns, fabrics or fibrous goods made from such materials, to be treated
- D06M2101/16—Synthetic fibres, other than mineral fibres
- D06M2101/18—Synthetic fibres consisting of macromolecular compounds obtained by reactions only involving carbon-to-carbon unsaturated bonds
- D06M2101/26—Polymers or copolymers of unsaturated carboxylic acids or derivatives thereof
- D06M2101/28—Acrylonitrile; Methacrylonitrile
Abstract
The invention discloses a kind of 3D conduction cell culturing brackets and preparation method thereof.The culture bracket passes through first by polyacrylonitrile and Fe3O4By electrostatic spinning technique with the reception of water phase device, polyacrylonitrile/Fe of high granular is made3O4Nano fiber scaffold wraps up PEDOT on scaffold fibers surface followed by in-situ polymerization, using the Electrostatic Absorption between graphene oxide and PEDOT, is made in 3D bracket outermost layer load graphene oxide.Culture bracket of the invention, average pore size is up to 13.8 μm between fiber, and the three-dimensional porous structure with the connection of inside and outside height, cell, which can be migrated smoothly to internal stent, forms uniform cell -3D cultivating system.Meanwhile the PEDOT with satisfactory electrical conductivity attracts each other with graphene oxide, graphene sheet layer is firmly attached to fiber outermost layer.Culture bracket of the invention has good biocompatibility, is conducive to cell adhesion, growth and proliferation.
Description
Technical field
The invention belongs to technical field of biomedical materials, it is related to a kind of 3D conduction cell culturing bracket and its preparation side
Method.
Background technique
Three-dimensional cell culture culture technique (TDCC) refer to by with three-dimensional structure different materials carrier and various differences
The cell of type co-incubation in vitro, enables cell to migrate, grow in the three-dimensional space structure of carrier, constitutes three
A kind of technology of the cell carrier compound of dimension.Compared to conventional two-dimensional plane culture, dimensional culture mode exists closer to cell
Growth conditions in natural extracellular matrix and the functionalization expression for maintaining cell, so as to simulate biology in vitro
Internal cell growth state.Therefore, dimensional culture technology can make result in drug screening and cytotoxicity detection process
It is more accurate, reliably.
Three-dimensional manometer fibrous framework possesses structure (Hogrebe N J, et similar with natural extracellular matrix because of it
al.Biomaterial microarchitecture:A potent regulator of individual cell
behavior and multicellular organization[J].Journal of Biomedical Materials
Research Part A, 2016.) and be widely used in the research of three-dimensional cell cultivation technology.Electrostatic spinning technique conduct
The optimal preparation method of nanofiber, can be used for preparing three-dimensional manometer fibrous framework.Currently based on the three of electrostatic spinning technique
The preparation strategy of dimension nano fiber scaffold mainly has: adding three-dimensional auxiliary reception device；Perforating agent is added in spinning process；Superposition
Electrospun nano-fibers film；Using polymer nature spinning process self assembly effect (Sun B, et
al.Advances in three-dimensional nanofibrous macrostructures via
electrospinning[J].Progress in Polymer Science,2014,39(5):862-890.).But above-mentioned side
The aperture of the three-dimensional manometer fibrous framework of method preparation is too small (< 5 μm), and cell can not be successfully to penetrate to migrate to internal stent and be formed
One cultivating system, therefore, cell are still to be grown in nano fiber scaffold surface, Wu Famo in a manner of approximate two bit plane cultures
Quasi- existence of the cell in natural biological body.
To versatile stem cell directed differentiation, nerve cell cynapse extends etc. has very significant electro photoluminescence culture cell
Facilitation, by electro photoluminescence in conjunction with three-dimensional cell cultivation bracket, can assign cell similar to natural extracellular matrix grow
Electro photoluminescence is introduced while environment, and the two collective effect is made to have the function that collaboration promotes (1.Tian H C, et in cell
al.Graphene oxide doped conducting polymer nanocomposite film for electrode-
tissue interface.[J].Biomaterials,2014,35(7):2120-2129.；2.Chen C,et
al.Biointerface by Cell Growth on Graphene Oxide Doped Bacterial Cellulose/
Poly(3,4-ethylenedioxythiophene)Nanofibers[J].Acs Applied Materials&
Interfaces,2016,8(16):10183.).Existing three-dimensional conductive bracket generally only has macroscopic three dimensional structure, implements electricity
Collaboration facilitation both when stimulation induction is unable to get embodiment (Chen C, et al.Three-Dimensional BC/
PEDOT Composite Nanofibers with High Performance for Electrode–Cell Interface
[J].Acs Applied Materials&Interfaces,2015,7(51):28244.).It is therefore desirable to prepare to have to fit
The three-dimensional manometer fibrous framework in suitable cell migration aperture.
Summary of the invention
It is poly- with 10 microns or more apertures, satisfactory electrical conductivity and biocompatibility that the purpose of the present invention is to provide a kind of
Acrylonitrile/Fe3O4/ polyethylene dioxythiophene/stannic oxide/graphene nano 3D conduction cell culturing bracket and preparation method thereof.
Realize that the technical solution of the object of the invention is as follows:
A kind of 3D conduction cell culturing bracket, by first by polyacrylonitrile and Fe3O4It is connect by electrostatic spinning technique with specific
Polyacrylonitrile/Fe of high granular is made in receiving apparatus3O4Nano fiber scaffold, followed by in-situ polymerization on scaffold fibers surface
It wraps up polyethylene dioxythiophene (PEDOT), using the Electrostatic Absorption between graphene oxide and PEDOT, in 3D bracket outermost layer
Graphene oxide is loaded, specific preparation process is as follows:
Step 1, polyacrylonitrile/Fe3O4Electrostatic spinning liquid preparation:
By Fe3O4Nano particle is placed in N ' dinethylformamide, and ultrasonic disperse is uniform, addition and Fe3O4Etc. quality
Triton X-100, ultrasonic disperse is uniform again, be added polyacrylonitrile fibril, ultrasonic dissolution, overnight, obtain polyacrylonitrile/
Fe3O4Electrostatic spinning liquid, wherein Fe3O4Concentration is 0.037~0.042g/mL；
Step 2, fluffy state polyacrylonitrile/Fe3O4The preparation of nanofiber:
By polyacrylonitrile/Fe3O4Electrostatic spinning liquid prepares nanofiber using wet electrostatic spinning technology, is had with bottom
The culture dish that magnet and the depth of water are 8~10mm is receiver, and nanofiber arrives fluffy state polypropylene through vacuum freeze drying
Nitrile/Fe3O4Nanofiber；
Step 3, fluffy state polyacrylonitrile/Fe3O4The preparation of/PEDOT:
By fluffy state polyacrylonitrile/Fe3O4Nanofiber is placed in the mixing diethyl ether solution of ethene dioxythiophene and ferric trichloride
In, under ice-water bath, ultrasonic confined reaction, after reaction, ethyl alcohol washs repeatedly, with the ethyl alcohol in water displacement bracket, through vacuum
It is freeze-dried to get fluffy state polyacrylonitrile/Fe is arrived3O4/ PEDOT nanofiber；
Step 4, the preparation of graphene oxide dispersion:
By graphene oxide ultrasonic disperse in water, graphene oxide dispersion is obtained；
Step 5, polyacrylonitrile/Fe3O4The preparation of/PEDOT/ graphene oxide 3D conducting bracket:
By fluffy state polyacrylonitrile/Fe3O4/ PEDOT nanofiber is placed in graphene oxide dispersion, is acutely vibrated, quiet
After setting overnight, washing removes unsupported graphene sheet layer to get polyacrylonitrile/Fe is arrived3O4/ PEDOT/ graphene oxide 3D is led
Electric bracket.
Preferably, in step 1, the polyacrylonitrile concentration is 0.125~0.15g/mL, Fe3O4The ultrasonic disperse time is
0.8~1h, Triton the X-100 ultrasonic disperse time are 0.8~1h, and the polyacrylonitrile fibril ultrasonic disperse time is 1.5~2h.
Preferably, in step 2, the magnet is ndfeb magnet, and spinning voltage is 15~16kV, and spinning flow velocity is
0.5~0.8mL/h, receiving distance is 10~12cm.
Preferably, in step 3, the concentration of the ethene dioxythiophene is 0.015~0.02g/mL, the concentration of iron chloride
For 0.02~0.03g/mL.
Preferably, in step 3, fluffy state polyacrylonitrile/Fe3O4Nanofiber is first scattered in ethene dioxythiophene solution,
Add the ferric chloride solution isometric with ethene dioxythiophene.
Preferably, in step 3, the water and ethanol replacement are replaced using gradient, and water and proportion of ethanol are followed successively by 1:9,
3:7,5:5,7:3,9:1.
Preferably, in step 4, the concentration of the graphene oxide dispersion is 0.5~1.5mg/mL.
Compared with prior art, of the invention to have the advantage that
(1) fluffy state polyacrylonitrile/Fe of the invention3O4Nanofiber density is 0.7718mg/cm3, belong to super light material,
Up to 13.8 μm, the three-dimensional porous structure with the connection of inside and outside height, cell can be migrated smoothly to bracket average pore size between fiber
Inside forms uniform cell -3D cultivating system；
(2) positive charge of the surface PEDOT institute band and surface of graphene oxide institute are negatively charged attracts each other, and makes graphene film
Layer is firmly attached to fiber outermost layer, not easily to fall off, and genotoxic potential will not be caused to cell, and the introducing of PEDOT assigns 3D branch
Frame satisfactory electrical conductivity, the building for responding model in vitro to microcurrent stimulating for native organism inner cell provide backing material,
The graphene oxide of bracket outermost layer load has good biocompatibility, is conducive to cell adhesion, growth and proliferation.
Detailed description of the invention
Fig. 1 is fluffy state polyacrylonitrile/Fe3O4Nanofiber Aqueous dispersions pictorial diagram.
Fig. 2 is fluffy state polyacrylonitrile/Fe3O4Nanofiber scanning electron microscope (SEM) photograph.
Fig. 3 is polyacrylonitrile/Fe3O4The scanning electron microscope (SEM) photograph of/PEDOT/ graphene oxide 3D conduction cell culturing bracket.
Fig. 4 is polyacrylonitrile/Fe of fluffy state3O4Nanofiber, polypropylene-base/Fe3O4/ PEDOT nanofiber and poly-
Acrylonitrile/Fe3O4The Cyclic voltamogram curve of/PEDOT/ graphene oxide 3D conduction cell culturing bracket.
Fig. 5 is the three-dimensional confocal scanning figure of 3D conducting bracket culture RGC-5.
Fig. 6 is the scanning electron microscope (SEM) photograph of 3D conducting bracket culture Hep G-2.
Fig. 7 is polyacrylonitrile/Fe prepared by comparative example 1 and 23O4Nanofiber scanning electron microscope (SEM) photograph.
Specific embodiment
The invention will be further described with attached drawing combined with specific embodiments below.
Embodiment 1
(1) 3mL n,N-Dimethylformamide is measured in screw socket bottle, weighs 0.117g Fe3O4Nano particle is placed in N, N-
In dimethylformamide, ultrasonic disperse 1h；It weighs and Fe3O4Etc. quality Triton X-100 in above-mentioned ultrasonic disperse
Fe3O4In dispersion liquid, continue ultrasonic disperse 1h；It weighs 0.45g polyacrylonitrile fibril to be added in dispersion liquid, be stood after ultrasonic 1.5h
Overnight up to spinning solution.
(2) device for spinning is built, with 8mm depth ultrapure water is equipped with, diameter is the culture dish of 22cm as reception device, is being trained
It supports ware bottom and places ndfeb magnet (10cm*10cm*1cm).Specific electrospinning parameters: 10cm 21# tack needle；Flow 0.5mL/
h；Receive distance 12cm；Voltage 15kV；Spinning time 12min.Fig. 1 is that nano fiber scaffold disperses pictorial diagram in water.
(3) nanofiber in water-bath is collected in centrifuge tube, and freezing is dry after two drop ethyl alcohol shaken wells are added thereto
Dry 60h is to get fluffy state polyacrylonitrile/Fe3O4Nanofiber.Fig. 2 is fluffy state polyacrylonitrile/Fe3O4Nanofiber scanning electricity
Mirror figure, fiber has Fe because of load as seen from Figure 23O4Nano particle and keep surface more coarse, and Fe3O4It is uniformly dispersed
In fiber, do not occur agglomeration；Fibrous framework aperture at 10 μm or more, the size aperture be conducive to cell smoothly migrate to
Internal stent forms uniform cell growth system.
(4) configuration 20mL concentration is the diethyl ether solution of 0.017g/mL ethene dioxythiophene, the 3D nanometer that will be prepared in (3)
Fibrous framework is placed in vibrating dispersion in diethyl ether solution, and configuration 20mL concentration is that second is added in the diethyl ether solution of 0.02g/mL ferric trichloride
In the diethyl ether solution of alkene dioxy thiophene, ultrasonic reaction 25min in ice-water bath.To wash Nanowire repeatedly with ethyl alcohol after the reaction was completed
Dimensional scaffold until eliminate ferric trichloride, then with water-ethanol mixed liquor gradient elution nano fiber scaffold (V completelyUltrapure water: VEthyl alcoholPoint
It Wei not 1:9；3:7；5:5；7:3；9:1), by the nano fiber scaffold freeze-drying 60h after elution to get fluffy state polypropylene
Base/Fe3O4/ PEDOT nanofiber conducting bracket.
(5) 8mL water is measured in screw socket bottle, 8mg graphene oxide ultrasonic disperse 2h is added thereto up to graphite oxide
Aqueous solution.
(6) branch prepared in (4) is placed in graphene oxide water solution, is dispersed in forced oscillation to bracket
In graphene oxide water solution, stand overnight.Be washed with water graphene oxide unadsorbed to the greatest extent, freeze-drying for 24 hours up to polyacrylonitrile/
Fe3O4/ PEDOT/ graphene oxide 3D conducting bracket.Fig. 3 is the scanning electron microscope (SEM) photograph of 3D conducting bracket, and arrow meaning is oxidation stone
Black alkene lamella.By finding out graphene sheet layer tight in figure in nano fiber outer layer, and non-oxidation graphene is filled in and receives
It in the micropore that rice fiber crossovers are constituted, therefore will not influence connectivity inside and outside 3D conducting bracket micropore, guarantee that cell can be suitable
Benefit is migrated to internal stent.
(7) respectively by polyacrylonitrile/Fe of fluffy state3O4Nanofiber, polypropylene-base/Fe3O4/ PEDOT nanofiber with
And 3D conducting bracket is scattered in PBS buffer solution, in this, as working electrode, platinum electrode is to electrode, and calomel electrode is reference
Electrode detects the bracket electrical property, scanning voltage: -0.4~0.6V using cyclic voltammetry in PBS (0.1M, PH=7.4)；
Sweep speed 100mV/s, institute's test loop VA characteristic curve are as shown in Figure 4.It is led since ferroferric oxide nano granules have
It is electrical, therefore polyacrylonitrile/Fe3O4Certain chemical property is presented in nanofiber, when by home position polymerization reaction in fiber table
Its conductivity increases after face loads PEDOT, and cyclic voltammetric electric current significantly improves；After loading not good conductor of electricity graphene oxide,
Therefore the original gap of cyclic voltammetric electric current less still shows good electric conductivity.
Embodiment 2
The present embodiment is substantially the same manner as Example 1, unique different to be Fe3O4Incorporation is 0.1125g, graphite oxide
In aqueous solution configuration, graphene oxide additional amount is 4mg, and the performance of 3D conducting bracket obtained is substantially the same manner as Example 1.
Embodiment 3
The present embodiment is substantially the same manner as Example 1, unique different to be that ethene dioxythiophene concentration is 0.02g/mL, oxygen
In the configuration of graphite aqueous solution, graphene oxide additional amount is 12mg, the performance and 1 base of embodiment of 3D conducting bracket obtained
This is identical.
Embodiment 4
Retinal ganglial cells dimensional culture
(1) 3D conducting bracket prepared by embodiment 1 is placed in 75% ethyl alcohol and impregnates sterilizing 8h, then led with PBS flushing
Ethyl alcohol is eliminated completely electric bracket 4 times.
(2) it weighs 0.1g agarose powder to be placed in 7mL PBS buffer solution, melt after high pressure sterilization in 120 DEG C, while hot will
Agarose solution is uniformly layered on 48 orifice plate bottoms, and sterilized 3D conducting bracket is added thereto after agarose cooled and solidified,
It is added after DMEM complete medium of the 500 μ L containing 15%FBS thereto and culture plate is placed in CO2It is incubated overnight in incubator.
(3) culture medium in 3D conducting bracket is removed, finely dispersed optic ganglion cell (RGC-5) cell suspension is added
Enter in 3D conducting bracket, cell concentration is 5000/hole, and into culture plate, supplement is added culture medium to 500 holes μ L/ and is placed on CO2
It is cultivated 5 days in incubator.It changes the liquid once within cell every 24 hours.
(4) the 3D conducting bracket for cultivating 5 days RGC-5 cells is taken out into contact plate, it is 2 μm of ol/L that 20 μ L concentration are added thereto
Calcein-AM solution be protected from light dyeing 15min after cell and fibre morphology are observed under three-dimensional confocal scanning microscope.Often
Observation 15min is supplemented into 3D bracket is added 20 μ L culture mediums.Fig. 4 is aobvious for the three-dimensional confocal scanning of 3D conducting bracket and cell
Micro mirror figure, from this it appears that cell tight is adhered on 3D conducting bracket, epicyte protein is by nanofiber packet in figure
It is wrapped in and wherein makes cell along the direction of nanofiber stretching, extension proliferation, to constitute extensive cell-nanofiber composite network.
The dimensional culture of liver cancer cells
(1) 3D conducting bracket prepared in embodiment 1 is placed in 75% ethyl alcohol and impregnates sterilizing 8h, then rushed with PBS
It washes conducting bracket 4 times and eliminates ethyl alcohol completely.
(2) it weighs 0.1g agarose powder to be placed in 7mL PBS buffer solution, melt after high pressure sterilization in 120 DEG C, while hot will
Agarose solution is uniformly layered on 24 orifice plate bottoms, and sterilized 3D conducting bracket is added thereto after agarose cooled and solidified,
It is added after DMEM complete medium of the 1mL containing 10%FBS thereto and culture plate is placed in CO2It is incubated overnight in incubator.
(3) culture medium in 3D conducting bracket is removed, it is conductive that 3D is added in finely dispersed liver cancer cells (HepG2) suspension
In bracket, cell concentration 2x105A/hole, into culture plate, supplement is added culture medium to the hole 1mL/ and is placed on CO2It is trained in incubator
It supports 3 days.It changes the liquid once within cell every 24 hours.
(4) culture medium removed after cell culture 3 days in culture plate washs 3D conducting bracket 3 times with PBS buffer solution, guide
The PBS solution of 1mL2.5% glutaraldehyde is added in electric bracket, is protected from light at 4 DEG C after sealing and stands 4h to fix cell.
(5) glutaraldehyde solution in conducting bracket is removed, the mixed liquor gradient elution of ethanol/water is used after washing 3 times with PBS
(VEthyl alcohol:VWater=1:9,2:8,3:7,5:5,7:3,9:1,10:0).Metal spraying observes cell shape under scanning electron microscope after freeze-drying
State.Fig. 6 is Hep G2 scanning electron microscope (SEM) photograph, and cell adhesion is on 3D conducting bracket in figure, and forms apparent cell aggregation,
This is conducive to the holding of liver cancer cells in vitro culture functionalization, so that the building for liver cancer in vitro study model provides ideal stent
Material.
Comparative example 1: polyacrylonitrile/Fe3O4The preparation of nanofiber
(1) 3mL n,N-Dimethylformamide is measured in screw socket bottle, weighs 0.135g Fe3O4Nano particle is placed in N, N-
In dimethylformamide, ultrasonic disperse 1h；It weighs and Fe3O4Etc. quality Triton X-100 in above-mentioned ultrasonic disperse
Fe3O4In dispersion liquid, continue ultrasonic disperse 1h；It weighs 0.45g polyacrylonitrile fibril to be added in dispersion liquid, be stood after ultrasonic 1.5h
Overnight up to spinning solution.
(2) device for spinning is built, with 8mm depth ultrapure water is equipped with, diameter is the culture dish of 22cm as reception device, is being trained
It supports ware bottom and places ndfeb magnet (10cm*10cm*1cm).Specific electrospinning parameters: 10cm 21# tack needle；Flow 0.5mL/
h；Receive distance 12cm；Voltage 15kV；Spinning time 12min.During the spinning process, due to Fe3O4Incorporation is excessive, makes fiber
It is attracted to receiver bottom rapidly by magnet, therefore when fiber is accumulated in water is affected by magnetic forces and is compressed, Fig. 7 a is made
Standby polyacrylonitrile/Fe3O4Nanofiber scanning electron microscope (SEM) photograph, it can be seen from the figure that aperture is and fine mostly at 5 μm between fiber
Dimension table surface roughness significantly improves, and Fe occurs3O4The phenomenon that reunion, therefore the nanofiber should not be used as three-dimensional cell cultivation
Backing material.
Comparative example 2: polyacrylonitrile/Fe3O4The preparation of nanofiber
(1) 3mL n,N-Dimethylformamide is measured in 5mL screw socket bottle, weighs 0.09g Fe3O4Nano particle is placed in N,
In dinethylformamide, ultrasonic disperse 1h；It weighs and Fe3O4Etc. quality Triton X-100 in above-mentioned ultrasonic disperse
Fe3O4In dispersion liquid, continue ultrasonic disperse 1h；It weighs 0.45g polyacrylonitrile fibril to be added in dispersion liquid, be stood after ultrasonic 1.5h
Overnight up to spinning solution.
(2) device for spinning is built, with 8mm depth ultrapure water is equipped with, diameter is the culture dish of 22cm as reception device, is being trained
It supports ware bottom and places ndfeb magnet (10cm*10cm*1cm).Specific electrospinning parameters: 10cm 21# tack needle；Flow 0.5mL/
h；Receive distance 12cm；Voltage 15kV；Spinning time 12min.During the spinning process, due to Fe3O4Incorporation is few, makes fiber institute
The magnetic force being subject to can not overcome the surface tension of water, therefore fiber can only be accumulated in the water surface, can not form discrete receive in water
Rice fibre structure.Fig. 7 b is prepared polyacrylonitrile/Fe3O4Nanofiber scanning electron microscope (SEM) photograph, fiber surface is relatively smooth in figure,
And there is Fe3O4It is distributed on fiber, but the aperture between fiber is mostly at 5 μm, and fiber wrapping phenomena occurs, therefore should
Fiber should not be used as three culture backing materials of cell.
Claims (8)
1. a kind of preparation method of 3D conduction cell culturing bracket, which is characterized in that specific step is as follows:
Step 1, polyacrylonitrile/Fe3O4Electrostatic spinning liquid preparation:
By Fe3O4Nano particle is placed in N ' dinethylformamide, and ultrasonic disperse is uniform, addition and Fe3O4Etc. quality
Triton X-100, ultrasonic disperse is uniform again, be added polyacrylonitrile fibril, ultrasonic dissolution, overnight, obtain polyacrylonitrile/
Fe3O4Electrostatic spinning liquid, wherein Fe3O4Concentration is 0.037~0.042g/mL；
Step 2, fluffy state polyacrylonitrile/Fe3O4The preparation of nanofiber:
By polyacrylonitrile/Fe3O4Electrostatic spinning liquid prepares nanofiber using wet electrostatic spinning technology, has magnet with bottom
And the culture dish that the depth of water is 8~10mm is receiver, nanofiber through vacuum freeze drying to get to fluffy state polyacrylonitrile/
Fe3O4Nanofiber；
Step 3, fluffy state polyacrylonitrile/Fe3O4The preparation of/PEDOT:
By fluffy state polyacrylonitrile/Fe3O4Nanofiber is placed in the mixing diethyl ether solution of ethene dioxythiophene and ferric trichloride,
Under ice-water bath, ultrasonic confined reaction, after reaction, ethyl alcohol washs repeatedly, with the ethyl alcohol in water displacement bracket, through vacuum refrigeration
It dries to get fluffy state polyacrylonitrile/Fe is arrived3O4/ PEDOT nanofiber；
Step 4, the preparation of graphene oxide dispersion:
By graphene oxide ultrasonic disperse in water, graphene oxide dispersion is obtained；
Step 5, polyacrylonitrile/Fe3O4The preparation of/PEDOT/ graphene oxide 3D conducting bracket:
By fluffy state polyacrylonitrile/Fe3O4/ PEDOT nanofiber is placed in graphene oxide dispersion, is acutely vibrated, was stood
After night, washing removes unsupported graphene sheet layer to get polyacrylonitrile/Fe is arrived3O4/ PEDOT/ graphene oxide 3D conduction branch
Frame.
2. preparation method according to claim 1, which is characterized in that in step 1, the polyacrylonitrile concentration is
0.125~0.15g/mL, Fe3O4The ultrasonic disperse time is 0.8~1h, and the Triton X-100 ultrasonic disperse time is 0.8~1h,
The polyacrylonitrile fibril ultrasonic disperse time is 1.5~2h.
3. preparation method according to claim 1, which is characterized in that in step 2, the magnet is ndfeb magnet,
Spinning voltage is 15~16kV, and spinning flow velocity is 0.5~0.8mL/h, and receiving distance is 10~12cm.
4. preparation method according to claim 1, which is characterized in that in step 3, the concentration of the ethene dioxythiophene
For 0.015~0.02g/mL, the concentration of iron chloride is 0.02~0.03g/mL.
5. preparation method according to claim 1, which is characterized in that in step 3, fluffy state polyacrylonitrile/Fe3O4Nanometer
Fiber is first scattered in ethene dioxythiophene solution, adds the ferric chloride solution isometric with ethene dioxythiophene.
6. preparation method according to claim 1, which is characterized in that in step 3, the water and ethanol replacement are using ladder
Degree displacement, water and proportion of ethanol are followed successively by 1:9,3:7,5:5,7:3,9:1.
7. preparation method according to claim 1, which is characterized in that in step 4, the graphene oxide dispersion
Concentration is 0.5~1.5mg/mL.
8. 3D conduction cell culturing bracket made from preparation method according to any one of claims 1 to 7.
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