CN105837730B - The method that a kind of combination laminated assembling technology and host-guest interaction build bioactivity surface - Google Patents

The method that a kind of combination laminated assembling technology and host-guest interaction build bioactivity surface Download PDF

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CN105837730B
CN105837730B CN201610183773.7A CN201610183773A CN105837730B CN 105837730 B CN105837730 B CN 105837730B CN 201610183773 A CN201610183773 A CN 201610183773A CN 105837730 B CN105837730 B CN 105837730B
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ada
multilayer film
polyelectrolyte multilayer
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CN105837730A (en
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陈红
于谦
曹利敏
韦婷
渠阳翠
胡昌明
占文俊
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Suzhou Hongfeng Hongfeng Biological Science And Technology Co Ltd
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Suzhou University
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    • C08F—MACROMOLECULAR COMPOUNDS OBTAINED BY REACTIONS ONLY INVOLVING CARBON-TO-CARBON UNSATURATED BONDS
    • C08F220/00—Copolymers of compounds having one or more unsaturated aliphatic radicals, each having only one carbon-to-carbon double bond, and only one being terminated by only one carboxyl radical or a salt, anhydride ester, amide, imide or nitrile thereof
    • C08F220/02—Monocarboxylic acids having less than ten carbon atoms; Derivatives thereof
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    • C08F220/06—Acrylic acid; Methacrylic acid; Metal salts or ammonium salts thereof
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    • A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
    • A61L27/14—Macromolecular materials
    • A61L27/22—Polypeptides or derivatives thereof, e.g. degradation products
    • A61L27/227—Other specific proteins or polypeptides not covered by A61L27/222, A61L27/225 or A61L27/24
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
    • A61L27/50—Materials characterised by their function or physical properties, e.g. injectable or lubricating compositions, shape-memory materials, surface modified materials
    • A61L27/54—Biologically active materials, e.g. therapeutic substances
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K17/00—Carrier-bound or immobilised peptides; Preparation thereof
    • C07K17/02—Peptides being immobilised on, or in, an organic carrier
    • C07K17/08—Peptides being immobilised on, or in, an organic carrier the carrier being a synthetic polymer
    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08F—MACROMOLECULAR COMPOUNDS OBTAINED BY REACTIONS ONLY INVOLVING CARBON-TO-CARBON UNSATURATED BONDS
    • C08F2/00—Processes of polymerisation
    • C08F2/38—Polymerisation using regulators, e.g. chain terminating agents, e.g. telomerisation
    • A—HUMAN NECESSITIES
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    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L2300/00—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices
    • A61L2300/20—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices containing or releasing organic materials
    • A61L2300/216—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices containing or releasing organic materials with other specific functional groups, e.g. aldehydes, ketones, phenols, quaternary phosphonium groups
    • A—HUMAN NECESSITIES
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    • A61L2300/00—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices
    • A61L2300/20—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices containing or releasing organic materials
    • A61L2300/23—Carbohydrates
    • A61L2300/232—Monosaccharides, disaccharides, polysaccharides, lipopolysaccharides
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    • A61L2300/00—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices
    • A61L2300/20—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices containing or releasing organic materials
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    • A61L2300/00—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices
    • A61L2300/40—Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices characterised by a specific therapeutic activity or mode of action
    • A61L2300/428—Vitamins, e.g. tocopherol, riboflavin

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Abstract

The present invention relates to bio-medical engineering and polymeric material field, there is provided a kind of method that combination laminated assembling technology and host-guest interaction build bioactivity surface.The present invention is by simple combined polymerization technology by guest molecule adamantane（Ada）Group is incorporated into polyacrylic acid（PAA）On obtain the copolymer of acrylic monomers and acrylic acid adamantane methanol ester monomer（P(AA‑Ada)）；Then utilize layer assembly（LbL）Technology is in a variety of amidized substrate surface deposition P (AA Ada) and polypropylene amine（PAH）Polyelectrolyte multilayer film.The beta cyclodextrin for having modified different biological molecules can be fixed to high-density using a large amount of adamantane groups contained on polyelectrolyte multilayer film（β‑CD）Derivative, so as to obtain the bioactivity surface with corresponding function.The present invention outstanding feature be：This method has universality, can be applied to a variety of different chemical property, the base material of shape；And the bioactivity surface performance built is adjustable, it is adaptable to wider bio-medical field.

Description

One kind combines laminated assembling technology and host-guest interaction builds bioactivity surface Method
Technical field
The present invention relates to bio-medical functional high polymer material field, and in particular to one kind combines laminated assembling technology and master Object interaction builds the preparation method of bioactivity surface on a variety of base materials.
Background technology
Bioactivity surface refers to the biologically functional molecules such as protein, polypeptide, nucleic acid, polysaccharide passing through physically or chemically side Method is fixed on synthetic surfaces, and then assigns the specific bioactive functions in surface（Such as biomolecule recognition performance, sterilization Performance, anticoagulation function, antifouling property etc.）, this is in many bio-medical engineerings and biological technical field extensive application.Cause How simply, efficiently, pervasively to biomaterial this, carry out surface modification significant to build bioactivity surface. The structure of usual bioactivity surface is, first in material surface one layer of matrix of formation, then biologically functional molecule to be fixed or born It is loaded in matrix.Polymer is due to its good biocompatibility, and with multiple avtive spots, feature is strong, it is easy to surface modification etc. Advantage is widely adopted as matrix.The polymer substrate preparation method generally used mainly includes self assembly, Langmuir- Blodgett films, polymer coating, surface initiation polymerization, laminated assembling technology etc..Among these, laminated assembling technology is a kind of base The polymer alternating layers with different electric charges are adsorbed to the method for modifying in substrate surface in electrostatic interaction.With its other party Method is compared, and laminated assembling technology has simple to operate, polymer film parameter is convenient to control, almost go for any shape and A variety of advantages such as the base material of physical property.Especially, the polymer film of laminated assembling technology formation has three-dimensional structure, fits very much Load immobilizing biologically active molecule is closed, by simply changing the thickness of multilayer film, fixed biological point just can be efficiently controlled The quantity of son.
In the common methods of material surface it is that biomolecule is passed through by covalent bond using chemical method by biomolecule fixation With reference in matrix.Although this method adhesion is stronger, the operation of complexity is usually required and using organic solvent etc..In recent years Come, widely paid close attention to using the affine fixation based on weak interaction.Host-guest interaction is a kind of common oversubscription Sub- chemical action, in advance by host molecule（Or guest molecule）Modified after biomolecule, just can be mutual by Subjective and Objective The fixation of biomolecule is easily realized in effect under mild conditions.Beta-schardinger dextrin（β-CD）It is a kind of common main body point Son, its hydrophobic cavity facilitates the insertion of a variety of guest molecules.β-CD itself have the hydroxyls of multiple convenient activation, can be with Realize on a molecule combine multiple biological micromolecule parts so that improve local ligand density and then so that after modify β- CD has higher bioactivity.In addition, chemically angle is set out, modification obtains having difference after we can be carried out to β-CD The β-cdderivatives of function., can be by simply mixing after two kinds or even various biomolecules using host-guest interaction The β-cdderivatives of modification, are realized difunctional or even multi-functional on the same surface of imparting.
The present invention is from the conventional polyanion/polycation of layer assembly to poly propenoic acid allylamine（PAA/PAH） With typical Subjective and Objective to beta-schardinger dextrin/adamantane（β-CD/Ada）It is used as model.First will by simple combined polymerization technology Ada groups, which are incorporated on polyanion PAA, obtains copolymer p (AA-Ada), then using laminated assembling technology in a variety of base materials Surface deposition P (AA-Ada)/PAH multilayer films, the β-CD of bioactive molecule will be modified with finally by host-guest interaction It is incorporated into and the bioactivity surface with corresponding function is obtained in polyelectrolyte multilayer film.
The content of the invention
The technical problems to be solved by the invention are：For the reaction present in current bioactivity surface preparation method The problem of journey is complicated, base material is limited, immobilizing biologically active molecular species is limited, function is single there is provided one kind it is simple to operate, have Universality, multiple biological activities molecule can be fixed, the surface modifying method of a variety of biological functions in surface is assigned.
The present invention solves its technical problem and uses following technical scheme：
A kind of structure that the present invention is provided has the preparation method of the material of correspondence biological function, it is characterised in that including Following steps：
（1）The preparation of polyelectrolyte multilayer film modified surface：
Amidized base material is immersed in the copolymer (P of acrylic monomers and 1- acrylic acid adamantane methanol ester monomers (AA-Ada) a period of time in solution), a period of time in polypropylene amine (PAH) solution is then immersed in again, i.e., in amination Substrate surface obtain one layer of P (AA-Ada)/PAH bilayer, immersion is repeated several times, if being had dried layer P (AA- Ada the polyelectrolyte multilayer film modified surface of)/PAH bilayers；
（2）Polyelectrolyte multilayer film modified surface immobilizing biologically active molecule：
Gained is placed in polyelectrolyte multilayer film modified surface and spread out containing the β-CD for having modified bioactive molecule Reacted in biological solution, that is, obtain being fixed with the polyelectrolyte multilayer film modified surface of bioactive molecule.
Another structure that the present invention is provided has the preparation method of the material of correspondence biological function, it is characterised in that bag Include following steps：
（1）The preparation of polyanion P (AA-Ada) containing adamantane group：
By certain mol ratio by acrylic monomers, 1- acrylic acid adamantane methanol ester monomer, chain-transferring agent and initiator It is added in reaction solution and carries out polymerisation, the method purified after reaction using sedimentation separation is produced containing adamantane group Polyanion P (AA-Ada)；
（2）The preparation of polyelectrolyte multilayer film modified surface：
Amidized base material is immersed in the copolymer (P of acrylic monomers and 1- acrylic acid adamantane methanol ester monomers (AA-Ada) a period of time in solution), a period of time in polypropylene amine (PAH) solution is then immersed in again, i.e., in amination Substrate surface obtain one layer of P (AA-Ada)/PAH bilayer, immersion is repeated several times, if being had dried layer P (AA- Ada the polyelectrolyte multilayer film modified surface of)/PAH bilayers；
（3）Polyelectrolyte multilayer film modified surface immobilizing biologically active molecule：
Gained is placed in polyelectrolyte multilayer film modified surface and spread out containing the β-CD for having modified bioactive molecule Reacted in biological solution, that is, obtain being fixed with the polyelectrolyte multilayer film modified surface of bioactive molecule.
Further, described bioactive molecule is biotin, mannose or arginine-glutamic acid-aspartic acid-figured silk fabrics ammonia Sour polypeptide.
Further, the preparation process for being modified with the β-cdderivatives of the biotin molecule is：By propargyl biotin and 7 β-the CD of individual azido group modification（β-CD-(N3)7）Click-reaction is carried out, the β-CD of biotin modification are made.
Further, the preparation process for being modified with the β-cdderivatives of the mannose molecules is：By propargyl mannose and 7 β-the CD of individual azido group modification（β-CD-(N3)7）Click-reaction is carried out, the β-CD of mannose-modified are made.
Further, the β-cdderivatives of the arginine-glutamic acid-aspartic acid-valine peptide molecule are modified with Preparation process is：By propargyl-arginine-glutamic acid-aspartic acid-valine polypeptide and the β-CD of 7 azido group modifications （β-CD-(N3)7）Click-reaction is carried out, the peptide modified β-CD of arginine-glutamic acid-aspartic acid-valine are made.
Further, the amidized base material be monocrystalline silicon or glass, the amidized monocrystalline silicon piece or sheet glass Preparation process is：Monocrystalline silicon piece or sheet glass are placed in 3- aminopropyls-triethoxysilane（APTES）Toluene solution in it is anti- The monocrystalline silicon or glass baseplate of surface amination should be obtained for a period of time.
Further, the amidized base material is gold-plated monocrystalline silicon piece, the preparation of the amidized gold-plated monocrystalline silicon piece Process is：Gold-plated monocrystalline silicon piece is placed in the aqueous solution of mercaptoethylamine hydrochloride and reacts a period of time, surface amination is obtained Gold-plated single crystal silicon substrate.
Further, modified bioactive molecule β-cdderivatives solution for the aqueous solution, phosphate buffer solution or Other can dissolve the solution of corresponding bioactive molecule.
Further, described to be polymerized to Invertible ideal, the chain-transferring agent is 2-（Phenyl first Acyl sulfenyl）Propionic acid；Described initiator is azodiisobutyronitrile, ABVN, dibenzoyl peroxide, the carbon of peroxidating two Sour ethylhexyl, isopropyl benzene hydroperoxide, potassium peroxydisulfate-sulphite system or hydrogen peroxide-perferrite system.
Further, the solution containing chain-transferring agent is DMF solution, the solution containing initiator For N,N-dimethylformamide solution.
Further, the β-CD of 7 azido group modifications（β-CD-(N3)7）Preparation method be：First by No. six positions of β-CD Hydroxyl carries out bromine substitution（β-CD-(Br)7）Or iodine substitution（β-CD-(I)7）, then carry out Azide with sodium azide.
A kind of structure that the present invention is provided has the preparation method of the material of correspondence biological function, and it includes having in detailed below Body step：
（1）The preparation of polyanion P (AA-Ada) containing adamantane group：
In reaction unit, acrylic monomers, 1- acrylic acid adamantane methanol ester monomer, chain are turned by certain mol ratio Move agent and initiator is added in reaction solution, under nitrogen protection stirring reaction, reaction temperature is 65 DEG C, the reaction time is 24 small When；Reaction is settled after terminating with ether, and the polyanion P (AA-Ada) containing adamantane group can be obtained after separating-purifying； The P (AA-Ada) is the english abbreviation of the copolymer of 1- acrylic acid adamantane methanol ester monomer and acrylic monomers；
The chain-transferring agent is 2-（Phenyl formyl sulfenyl）Propionic acid；Described initiator is azodiisobutyronitrile etc.； Described reaction solution is DMF solution.
（2）The preparation of polyelectrolyte multilayer film modified surface：
Prepare polyelectrolyte solution, including polyanion P (AA-Ada) solution and polycation PAH solution；By amination Base material be immersed in P (AA-Ada) solution, through over cleaning remove combine weaker polymer after be immersed in again in PAH solution, P (AA-Ada)/PAH bilayer can be obtained on surface after over cleaning, this process repeated several times is finally reached The number of plies needed, obtains polyelectrolyte multilayer film modified surface；
Described polyelectrolyte solution is acetic acid/sodium acetate buffer that pH value is 5.0；Taken from polyelectrolyte solution The cleaning solution on the surface gone out is acetic acid/sodium acetate buffer that pH value is 5.0；The amidized base material be monocrystalline silicon, Glass or gold-plated monocrystalline silicon.
The preparation process of the amidized base material comprises the following steps：
（a）Clean monocrystalline silicon piece or sheet glass are first used into " piranha " solution（Sulfuric acid:Hydrogen peroxide=7:3, volume ratio） Pre-processed, be subsequently placed in 3- aminopropyls-triethoxysilane（APTES）Toluene solution in react and stay overnight, obtain table The amidized monocrystalline silicon in face or glass baseplate；
（b）The gold-plated monocrystalline silicon piece ammoniacal liquor that will be cleaned by UV ozone instrument, the mixing of hydrogen peroxide and deionized water is molten Liquid（Ammoniacal liquor:Hydrogen peroxide:Deionized water=1:1:5, volume ratio）Pre-processed, be subsequently placed in the water of mercaptoethylamine hydrochloride Reaction is stayed overnight in solution, obtains the gold-plated single crystal silicon substrate of surface amination；
（3）Polyelectrolyte multilayer film modified surface immobilizing biologically active molecule：
The polyelectrolyte multilayer film modified surface of gained is placed in containing the molten of the β-cdderivatives for having modified biomolecule Reacted in liquid, reaction temperature is room temperature, the reaction time is 12 ~ 24 hours, and reaction cleans surface after terminating with deionized water, The polyelectrolyte multilayer film modified surface of bioactive molecule can be fixed；
Described bioactive molecule is biotin, mannose and REDV polypeptides；Described has modified bioactive molecule β-cdderivatives solution for the aqueous solution, phosphate buffer solution or other can dissolve the molten of corresponding bioactive molecule Liquid.
The preparation process of the β-cdderivatives for being modified with bioactive molecule comprises the following steps：
（a）β-the CD of 7 azido group modifications（β-CD-(N3)7）Preparation；
（b）β-the CD of biotin modification preparation：
In reaction unit, β-CD- (N are first added3)7And biotin-PEG4- alkyne, is then added under nitrogen protection Catalyst/ligand solution, stirring reaction, reaction temperature is 50 DEG C, and the reaction time is 36 hours, is reacted mixed solution after terminating Dialysed with MWCO 1000 bag filter, the β-CD of biotin modification are can obtain after freeze-drying.
（c）β-the CD of mannose-modified preparation：
By 1- under nitrogen protection（2 '-propargyl）- α-D-MANNOSE and β-CD- (N3)7Add in catalyst system solution, Click-reaction is carried out, reaction temperature is 50 DEG C, the reaction time is 24 hours, and reaction removes solvent after terminating, and uses absolute methanol Precipitation is collected by centrifugation, the β-CD of mannose-modified are can obtain after purification.
（d）β-CD peptide modified REDV preparation：
By propargyl-REDV and β-CD- (N under nitrogen protection3)7Add in catalyst system solution, carry out click-reaction, Reaction temperature is 50 DEG C, and the reaction time is 24 hours, and post-reaction treatment is to be dialysed using MWCO 500 bag filter, and freezing is dry It is dry to obtain the peptide modified β-CD of REDV.
It is preferred that, step（a）Described in preparation method be following one or two kinds of：
The hydroxyl of No. six positions of β-CD first is carried out into bromine to replace（β-CD-(Br)7）, then carry out Azide with sodium azide；
The hydroxyl of No. six positions of β-CD first is carried out into iodine to replace（β-CD-(I)7）, then carry out Azide with sodium azide.
It is preferred that, step（b）Described in catalyst/ligand solution be dimethyl sulphoxide solution, the catalyst/ligand System is 2,2'- bipyridyls/copper bromide etc..
It is preferred that, step（c）Described in catalyst solution be dimethyl sulphoxide solution, the catalyst/ligand system For 2,2'- bipyridyls/copper bromide etc..
It is preferred that, step（d）Described in catalyst system solution be dimethyl sulfoxide (DMSO)/water mixed solution, the catalysis System is copper sulphate/sodium ascorbate etc..
By above-mentioned steps, the biology for finally giving the high density fixed biologically functional molecule with correspondence biological function is living Property surface.
The present invention has the advantages that following main compared with prior art：
The method that the present invention is provided is that combination laminated assembling technology and host-guest interaction prepare bioactivity surface Method.Compared with prior art, the present invention has following prominent characteristics：
1. it is simple to operate, easy：Layer assembly and Subjective and Objective reaction condition are gentle, can be under room temperature, aqueous environment Complete.
2. applicability is wider：The method that laminated assembling technology builds polyelectrolyte multilayer film can apply to different chemistry Property, base material of different shapes can be realized in any substrate surface containing amino, it is adaptable to such as biological detection, tissue work Many bio-medical fields such as journey, protein separation.
3. surface property is adjustable：On the one hand can be simply by changing layer assembly number of plies control load biomolecule Capacity, the β-CD that the modification of different biologically functional molecules on the other hand can be designed as needed assign surface different biological work( Energy.For example, after fixing the β-CD of biotin modification on the polyelectrolyte multilayer film surface obtained using this method, can be selective With reference to Avidin（See Fig. 1）；, can be with specific absorption canavaline after fixing the β-CD of mannose-modified on surface（Con A）（See Fig. 2）；Fixed on surface after the peptide modified β-CD of REDV, sticking for surface endothelial cell can be promoted（See Fig. 3）.
Brief description of the drawings
Fig. 1 is that the β-CD front and rear surfaces of biotin modification are fixed to fluorescent decoration in monocrystalline silicon piece polyelectrolyte multilayer film surface Avidin absorption situation；
Fig. 2 is that the β-CD front and rear surfaces of mannose-modified are fixed to fluorescence in gold-plated monocrystalline silicon piece polyelectrolyte multilayer film surface The absorption situation of the canavaline of modification.
Fig. 3 is that table before and after the peptide modified β-CD of REDV is fixed on polyelectrolyte multilayer film surface on gold-plated monocrystalline silicon piece base material Stick situation in face of endothelial cell.
Embodiment
The method that the combination laminated assembling technology and host-guest interaction that the present invention is provided build bioactivity surface, be First carry out amination pretreatment to substrate surface, then by polyelectrolyte P (AA-Ada) and PAH by laminated assembling technology on surface Deposition obtains the polyelectrolyte multilayer film containing Ada groups.The host-guest interaction between Ada and β-CD is finally utilized, in β-the CD for being modified with different bioactive molecules are fixed on polyelectrolyte multilayer film surface, assign surface corresponding biological function.
Below by embodiment, the present invention is further elaborated, but does not limit the present invention.
Embodiment 1
Being built with reference to laminated assembling technology and host-guest interaction in monocrystalline silicon sheet surface, there is biomolecule to recognize work( The surface of energy
Monocrystalline silicon piece " piranha " solution that will be cleaned with acetone soln ultrasound（Sulfuric acid:Hydrogen peroxide=7:3, volume Than）Pre-processed, be subsequently placed in 3- aminopropyls-triethoxysilane（APTES）Toluene solution in react 18 hours, i.e., Obtain the monocrystalline silicon piece of surface amination.Polyanion P (AA-Ada) solution and polycation PAH solution, solvent are prepared respectively The concentration for being 5.0 for pH value is acetic acid/sodium acetate buffer of 0.05 mol/L.Amidized monocrystalline silicon piece is immersed in In P (AA-Ada) solution, removed through peracetic acid/sodium acetate buffer cleaning after combining weaker polymer and be immersed in PAH again In solution, P (AA-Ada)/PAH bilayer is obtained on surface after over cleaning, this process repeated several times finally reaches To the number of plies of needs, you can obtain the monocrystalline silicon piece of polyelectrolyte multilayer film modification.
The hydroxyl of No. six positions of β-CD first is carried out into bromine to replace（β-CD-(Br)7）, then carry out Azide system with sodium azide β-the CD of standby 7 azido groups modification（β-CD-(N3)7）；Finally by β-CD- (N3)7Carry out clicking on instead with propargyl biotin Should, obtain the β-CD of biotin modification（β-CD-(biotin)7）.
The monocrystalline silicon piece that above-mentioned polyelectrolyte multilayer film is modified is placed in 1 mM every liter of β-CD- (biotin)7It is water-soluble In liquid, and reaction is stayed overnight at room temperature.Reaction cleans surface after terminating with water, you can be fixed biotin has biology The surface of molecular recognition function.
Embodiment 2
Being built with reference to laminated assembling technology and host-guest interaction in gold-plated monocrystalline silicon sheet surface has specific adsorption The surface of canavaline function
By the gold-plated monocrystalline silicon piece ammoniacal liquor clean with UV ozone instrument is passed through, the mixed solution of hydrogen peroxide and deionized water （Ammoniacal liquor:Hydrogen peroxide:Deionized water=1:1:5, volume ratio）Pre-processed, be subsequently placed in the water-soluble of mercaptoethylamine hydrochloride Reaction is stayed overnight in liquid, that is, obtains the gold-plated monocrystalline silicon piece of surface amination.Polyanion P (AA-Ada) solution is prepared respectively and poly- Cation PAH solution, solvent is acetic acid/sodium acetate buffer that the concentration that pH value is 5.0 is 0.05 mol/L.By amino The gold-plated monocrystalline silicon piece changed is immersed in P (AA-Ada) solution, is combined through peracetic acid/sodium acetate buffer cleaning removal weaker Polymer after be immersed in again in PAH solution, P (AA-Ada)/PAH bilayer is obtained on surface after over cleaning, This process repeated several times finally reaches the number of plies of needs, you can obtain the gold-plated monocrystalline silicon piece of polyelectrolyte multilayer film modification.
The hydroxyl of No. six positions of β-CD first is carried out into bromine to replace（β-CD-(Br)7）, then carry out Azide system with sodium azide β-the CD of standby 7 azido groups modification（β-CD-(N3)7）；Finally by β-CD- (N3)7And 1-（2 '-propargyl）- α-D-MANNOSE Click-reaction is carried out, the β-CD of mannose-modified are obtained（β-CD-(mannose)7）.
The gold-plated monocrystalline silicon piece that above-mentioned polyelectrolyte multilayer film is modified is placed in 1 mM every liter of β-CD- (mannose)7In the aqueous solution, and reaction is stayed overnight at room temperature.Reaction cleans surface after terminating with water, you can be fixed sweet That reveals sugar has specific adsorption canavaline（Con A）The surface of function.
Embodiment 3
Building to have in gold-plated monocrystalline silicon sheet surface with host-guest interaction with reference to laminated assembling technology promotes endothelium thin The surface of born of the same parents' adhesive function
By the gold-plated monocrystalline silicon piece ammoniacal liquor clean with UV ozone instrument is passed through, the mixed solution of hydrogen peroxide and deionized water （Ammoniacal liquor:Hydrogen peroxide:Deionized water=1:1:5, volume ratio）Pre-processed, be subsequently placed in the water-soluble of mercaptoethylamine hydrochloride Reaction is stayed overnight in liquid, that is, obtains the gold-plated monocrystalline silicon piece of surface amination.Polyanion P (AA-Ada) solution is prepared respectively and poly- Cation PAH solution, solvent is acetic acid/sodium acetate buffer that the concentration that pH value is 5.0 is 0.05 mol/L.By amino The gold-plated monocrystalline silicon piece changed is immersed in P (AA-Ada) solution, is combined through peracetic acid/sodium acetate buffer cleaning removal weaker Polymer after be immersed in again in PAH solution, P (AA-Ada)/PAH bilayer is obtained on surface after over cleaning, This process repeated several times finally reaches the number of plies of needs, you can obtain the gold-plated monocrystalline silicon piece of polyelectrolyte multilayer film modification.
The hydroxyl of No. six positions of β-CD first is carried out into bromine to replace（β-CD-(Br)7）, then carry out Azide system with sodium azide β-the CD of standby 7 azido groups modification（β-CD-(N3)7）；Finally by β-CD- (N3)7Clicked on propargyl-REDV polypeptides Reaction, obtains the peptide modified β-CD of REDV（β-CD-(REDV)7）.
The gold-plated monocrystalline silicon piece that above-mentioned polyelectrolyte multilayer film is modified is placed in 1 mM every liter of β-CD- (REDV)7Water In solution, and reaction is stayed overnight at room temperature.Reaction cleans surface after terminating with water, you can be fixed having for REDV polypeptides Promote the surface of adhesion function of monocytes.
This method has potential application prospect in many bio-medicals and technical field of biology.Wherein need to particularly point out Be that party's science of law is not only limited to us and worked PAA/PAH, the β-CD/Ada of use, and can be generalized to other polymers With other Subjective and Objectives pair.For needing difunctional or even multifunction surface field, simply it can be introduced altogether using mixing Mode introduces two or more, and there is the cyclodextrin molecular of different ligands to be achieved.
The desirable embodiment according to the present invention is enlightenment above, by above-mentioned description, and related personnel completely can be with Without departing from the scope of the technological thought of the present invention', various changes and amendments are carried out.The technical scope of this invention It is not limited to the content on specification, it is necessary to determine the technical scope according to the scope of the claims.

Claims (10)

1. a kind of method that combination laminated assembling technology and host-guest interaction build bioactivity surface, it is characterised in that Comprise the following steps：
（1）The preparation of polyelectrolyte multilayer film modified surface：
Amidized base material is immersed in the copolymer p (AA-Ada) of acrylic monomers and 1- acrylic acid adamantane methanol ester monomers Solution in a period of time, a period of time is then immersed in polypropylene amine PAH solution again, i.e., is obtained in amidized substrate surface To one layer of P (AA-Ada)/PAH bilayer, immersion is repeated several times, if being had double points of dried layer P (AA-Ada)/PAH The polyelectrolyte multilayer film modified surface of sublayer；
（2）Polyelectrolyte multilayer film modified surface immobilizing biologically active molecule：
Gained is placed in polyelectrolyte multilayer film modified surface and spread out containing the beta-schardinger dextrin for having modified bioactive molecule Reacted in biological solution, that is, obtain being fixed with the polyelectrolyte multilayer film modified surface of bioactive molecule.
2. a kind of method that combination laminated assembling technology and host-guest interaction build bioactivity surface, it is characterised in that Comprise the following steps：
（1）The preparation of polyanion P (AA-Ada) containing adamantane group：
Acrylic monomers, 1- acrylic acid adamantane methanol ester monomer, chain-transferring agent and initiator are added by certain mol ratio Polymerisation is carried out into reaction solution, the method purified after reaction using sedimentation separation produces poly- the moon containing adamantane group Ion P (AA-Ada)；
（2）The preparation of polyelectrolyte multilayer film modified surface：
Amidized base material is immersed in the copolymer p (AA-Ada) of acrylic monomers and 1- acrylic acid adamantane methanol ester monomers Solution in a period of time, a period of time is then immersed in polypropylene amine PAH solution again, i.e., is obtained in amidized substrate surface To one layer of P (AA-Ada)/PAH bilayer, immersion is repeated several times, if being had double points of dried layer P (AA-Ada)/PAH The polyelectrolyte multilayer film modified surface of sublayer；
（3）Polyelectrolyte multilayer film modified surface immobilizing biologically active molecule：
Gained is placed in polyelectrolyte multilayer film modified surface and spread out containing the beta-schardinger dextrin for having modified bioactive molecule Reacted in biological solution, that is, obtain being fixed with the polyelectrolyte multilayer film modified surface of bioactive molecule.
3. method according to claim 1 or 2, it is characterised in that：Described bioactive molecule is biotin, mannose Or arginine-glutamic acid-aspartic acid-valine polypeptide.
4. method according to claim 3, it is characterised in that：It is modified with the system of the β-cdderivatives of the biotin molecule It is for process：β-CD by propargyl biotin and the modification of 7 azido groups are：β-CD-(N3)7, click-reaction is carried out, is made β-the CD of biotin modification.
5. method according to claim 3, it is characterised in that：It is modified with the system of the β-cdderivatives of the mannose molecules It is for process：β-CD by propargyl mannose and the modification of 7 azido groups are：β-CD-(N3)7, click-reaction is carried out, is made β-the CD of mannose-modified.
6. method according to claim 3, it is characterised in that：It is modified with the arginine-glutamic acid-aspartic acid-figured silk fabrics The preparation process of the β-cdderivatives of propylhomoserin peptide molecule is：Propargyl-arginine-glutamic acid-aspartic acid-valine is more Peptide and the β-CD of 7 azido group modifications are：β-CD-(N3)7, click-reaction is carried out, arginine-glutamic acid-asparagus fern ammonia is made Peptide modified β-the CD of acid-valine.
7. method according to claim 1 or 2, it is characterised in that：The amidized base material is monocrystalline silicon or glass, institute The preparation process for stating amidized monocrystalline silicon piece or sheet glass is：Monocrystalline silicon piece or sheet glass are placed in the second of 3- aminopropyls-three TMOS（APTES）Toluene solution in react a period of time, obtain the monocrystalline silicon or glass baseplate of surface amination.
8. method according to claim 1 or 2, it is characterised in that：The amidized base material is gold-plated monocrystalline silicon piece, institute The preparation process for stating amidized gold-plated monocrystalline silicon piece is：Gold-plated monocrystalline silicon piece is placed in the aqueous solution of mercaptoethylamine hydrochloride Reaction a period of time, obtain the gold-plated single crystal silicon substrate of surface amination.
9. method according to claim 1 or 2, it is characterised in that：The β-cdderivatives of bioactive molecule are modified Solution be the aqueous solution, phosphate buffer solution or other can dissolve the solution of corresponding bioactive molecule.
10. method according to claim 2, it is characterised in that：Described is polymerized to Invertible ideal, The chain-transferring agent is 2-（Phenyl formyl sulfenyl）Propionic acid；Described initiator is azodiisobutyronitrile, azo two different heptan Nitrile, dibenzoyl peroxide, dicetyl peroxydicarbonate ethylhexyl, isopropyl benzene hydroperoxide, potassium peroxydisulfate-sulphite body System or hydrogen peroxide-perferrite system.
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