CN102813963A - Method for fixing functional molecules on biomedical material surface with dopamine serving as bridging - Google Patents
Method for fixing functional molecules on biomedical material surface with dopamine serving as bridging Download PDFInfo
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- CN102813963A CN102813963A CN2012103316666A CN201210331666A CN102813963A CN 102813963 A CN102813963 A CN 102813963A CN 2012103316666 A CN2012103316666 A CN 2012103316666A CN 201210331666 A CN201210331666 A CN 201210331666A CN 102813963 A CN102813963 A CN 102813963A
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- VYFYYTLLBUKUHU-UHFFFAOYSA-N dopamine Chemical compound NCCC1=CC=C(O)C(O)=C1 VYFYYTLLBUKUHU-UHFFFAOYSA-N 0.000 title claims abstract description 108
- 229960003638 dopamine Drugs 0.000 title claims abstract description 54
- 239000003519 biomedical and dental material Substances 0.000 title claims abstract description 48
- 238000000034 method Methods 0.000 title claims abstract description 32
- 238000006243 chemical reaction Methods 0.000 claims abstract description 34
- HLXZNVUGXRDIFK-UHFFFAOYSA-N nickel titanium Chemical compound [Ti].[Ti].[Ti].[Ti].[Ti].[Ti].[Ti].[Ti].[Ti].[Ti].[Ti].[Ni].[Ni].[Ni].[Ni].[Ni].[Ni].[Ni].[Ni].[Ni].[Ni].[Ni].[Ni].[Ni].[Ni] HLXZNVUGXRDIFK-UHFFFAOYSA-N 0.000 claims description 31
- 229910001000 nickel titanium Inorganic materials 0.000 claims description 31
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 claims description 28
- HTTJABKRGRZYRN-UHFFFAOYSA-N Heparin Chemical compound OC1C(NC(=O)C)C(O)OC(COS(O)(=O)=O)C1OC1C(OS(O)(=O)=O)C(O)C(OC2C(C(OS(O)(=O)=O)C(OC3C(C(O)C(O)C(O3)C(O)=O)OS(O)(=O)=O)C(CO)O2)NS(O)(=O)=O)C(C(O)=O)O1 HTTJABKRGRZYRN-UHFFFAOYSA-N 0.000 claims description 28
- 229920000669 heparin Polymers 0.000 claims description 28
- 229960002897 heparin Drugs 0.000 claims description 28
- 239000012153 distilled water Substances 0.000 claims description 22
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 17
- 229910052757 nitrogen Inorganic materials 0.000 claims description 14
- QKNYBSVHEMOAJP-UHFFFAOYSA-N 2-amino-2-(hydroxymethyl)propane-1,3-diol;hydron;chloride Chemical compound Cl.OCC(N)(CO)CO QKNYBSVHEMOAJP-UHFFFAOYSA-N 0.000 claims description 7
- 238000001035 drying Methods 0.000 claims description 7
- 238000002791 soaking Methods 0.000 claims description 7
- KIUKXJAPPMFGSW-DNGZLQJQSA-N (2S,3S,4S,5R,6R)-6-[(2S,3R,4R,5S,6R)-3-Acetamido-2-[(2S,3S,4R,5R,6R)-6-[(2R,3R,4R,5S,6R)-3-acetamido-2,5-dihydroxy-6-(hydroxymethyl)oxan-4-yl]oxy-2-carboxy-4,5-dihydroxyoxan-3-yl]oxy-5-hydroxy-6-(hydroxymethyl)oxan-4-yl]oxy-3,4,5-trihydroxyoxane-2-carboxylic acid Chemical compound CC(=O)N[C@H]1[C@H](O)O[C@H](CO)[C@@H](O)[C@@H]1O[C@H]1[C@H](O)[C@@H](O)[C@H](O[C@H]2[C@@H]([C@@H](O[C@H]3[C@@H]([C@@H](O)[C@H](O)[C@H](O3)C(O)=O)O)[C@H](O)[C@@H](CO)O2)NC(C)=O)[C@@H](C(O)=O)O1 KIUKXJAPPMFGSW-DNGZLQJQSA-N 0.000 claims description 4
- RKDVKSZUMVYZHH-UHFFFAOYSA-N 1,4-dioxane-2,5-dione Chemical compound O=C1COC(=O)CO1 RKDVKSZUMVYZHH-UHFFFAOYSA-N 0.000 claims description 4
- 239000012901 Milli-Q water Substances 0.000 claims description 4
- 229920002674 hyaluronan Polymers 0.000 claims description 4
- 229960003160 hyaluronic acid Drugs 0.000 claims description 4
- SQDAZGGFXASXDW-UHFFFAOYSA-N 5-bromo-2-(trifluoromethoxy)pyridine Chemical compound FC(F)(F)OC1=CC=C(Br)C=N1 SQDAZGGFXASXDW-UHFFFAOYSA-N 0.000 claims description 2
- 229920001661 Chitosan Polymers 0.000 claims description 2
- 229920001287 Chondroitin sulfate Polymers 0.000 claims description 2
- LSDPWZHWYPCBBB-UHFFFAOYSA-N Methanethiol Chemical compound SC LSDPWZHWYPCBBB-UHFFFAOYSA-N 0.000 claims description 2
- 235000001014 amino acid Nutrition 0.000 claims description 2
- 229940024606 amino acid Drugs 0.000 claims description 2
- 150000001413 amino acids Chemical class 0.000 claims description 2
- 229940045110 chitosan Drugs 0.000 claims description 2
- 229940059329 chondroitin sulfate Drugs 0.000 claims description 2
- 229920001577 copolymer Polymers 0.000 claims description 2
- JJTUDXZGHPGLLC-UHFFFAOYSA-N lactide Chemical compound CC1OC(=O)C(C)OC1=O JJTUDXZGHPGLLC-UHFFFAOYSA-N 0.000 claims description 2
- 229920000747 poly(lactic acid) Polymers 0.000 claims description 2
- 239000000463 material Substances 0.000 abstract description 14
- 230000004071 biological effect Effects 0.000 abstract description 6
- 230000004048 modification Effects 0.000 abstract description 4
- 238000012986 modification Methods 0.000 abstract description 4
- 238000006116 polymerization reaction Methods 0.000 abstract description 4
- 230000008569 process Effects 0.000 abstract description 3
- 229920001690 polydopamine Polymers 0.000 abstract 3
- 230000035484 reaction time Effects 0.000 abstract 1
- 238000006557 surface reaction Methods 0.000 abstract 1
- 210000004027 cell Anatomy 0.000 description 7
- 230000000694 effects Effects 0.000 description 5
- 230000010100 anticoagulation Effects 0.000 description 4
- 210000001519 tissue Anatomy 0.000 description 4
- 239000000470 constituent Substances 0.000 description 3
- 239000000126 substance Substances 0.000 description 3
- 108090000190 Thrombin Proteins 0.000 description 2
- 210000004204 blood vessel Anatomy 0.000 description 2
- 238000007654 immersion Methods 0.000 description 2
- 235000018102 proteins Nutrition 0.000 description 2
- 108090000623 proteins and genes Proteins 0.000 description 2
- 102000004169 proteins and genes Human genes 0.000 description 2
- 230000004044 response Effects 0.000 description 2
- 229960004072 thrombin Drugs 0.000 description 2
- 108090000790 Enzymes Proteins 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 238000010521 absorption reaction Methods 0.000 description 1
- 230000002776 aggregation Effects 0.000 description 1
- 238000004220 aggregation Methods 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 230000033115 angiogenesis Effects 0.000 description 1
- 239000003146 anticoagulant agent Substances 0.000 description 1
- 229940127219 anticoagulant drug Drugs 0.000 description 1
- 230000008901 benefit Effects 0.000 description 1
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- 210000000988 bone and bone Anatomy 0.000 description 1
- 230000002308 calcification Effects 0.000 description 1
- 210000000845 cartilage Anatomy 0.000 description 1
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- 230000002708 enhancing effect Effects 0.000 description 1
- 238000007306 functionalization reaction Methods 0.000 description 1
- 210000002216 heart Anatomy 0.000 description 1
- 210000003709 heart valve Anatomy 0.000 description 1
- 230000036039 immunity Effects 0.000 description 1
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- 229910010272 inorganic material Inorganic materials 0.000 description 1
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Abstract
The invention discloses a method for fixing functional molecules on a biomedical material surface with dopamine serving as a bridging. The method uses the dopamine to serve as a bridging medium, firstly uses strong auto polymerization characters of the dopamine under certain pH environment to be assembled on the biomedical material surface to form a poly-dopamine layer, and then a chemical reaction is used for stem grafting of the functional molecules on the poly-dopamine layer to build the biomedical material surface with biological activity. The method does not damage the biological activity of the functional molecules and can change assembling thickness of the poly-dopamine layer by controlling reaction time so as to change grafting percent of the functional molecules on the surface. The method is simple and practical in process, good in repeatability and capable of widely applying to surface functionalization modification of biomedical materials such as intervention materials and tissue engineering materials, and has good clinical application prospects.
Description
Technical field
The present invention relates to the biological activity modification of used in tissue engineering base material, relating in particular to a kind of is the method for the bio-medical material surface fixed function molecule of bridging with the dopamine.
Background technology
Organizational project is one and relates to multidisciplinary, multi-field crossing research problems such as medical science, chemistry, biology, materialogy.In the research of organizational project, functional molecular effectively is fixed to the bio-medical material surface, making up the bioactivation timbering material is important step wherein, it directly has influence on the activity and the functional expression of purpose cell and destination protein enzyme.Functional molecular is meant the macromolecular material with some specific function.Why they have particular functionality, are owing in its strand, combined the particular functionality group, or molecule and other materials with specific function carried out compoundly, and perhaps the two haves both at the same time.Functional molecular through special, high-affinity with cell membrane on receptors bind, finally stimulate cellular proliferation and produce the polypeptides matter of other biological effect.Functional molecular can transmit, conversion or storage material, energy and information, thereby influences growth, propagation and the differentiation of surrounding tissue cell.Functional molecular shows as multifunctionality to the effect of target cell.With heparin and hyaluronic acid is example.Heparin is a kind of anticoagulant commonly used, can strengthen the affinity of antithrombase and thrombin, quickens the inactivation of thrombin; Suppress hematoblastic adhesion and aggregation; Increase the permeability of blood vessel wall, and adjustable angiogenesis, be the choice drug that need reach anticoagulation rapidly therefore.Hyaluronic acid can make cell keep separated from one another, can make moisture get into intercellular substance, and form protein gel with protein bound; Cell is sticked together, and maintenance cell moisture is played in the cellular metabolism effect of bringing into normal play; The protection cell does not receive the infringement of pathogen, accelerates to recover skin histology, improves the wound healing regeneration capacity; Reduce cicatrix, effects such as enhancing immunity.Through this type functional molecular is assembled into substrate surface, can give base material different functions characteristics, the most at last the base material functionalization.
The compound of functional molecular and bio-medical material is the important means that realizes the bio-medical material biological activityization, and the bio-medical material that is compounded with functional molecular can have the different functions characteristics.Traditional complex technique mainly contains physisorphtion and chemical fixation.And simple physics absorption can not reach the purpose of held stationary functional molecular, fixing its biological activity of functional molecular forfeiture that then might make of chemical method.
Summary of the invention
The purpose of this invention is to provide a kind of simple and easy to do be the method for the bio-medical material surface fixed function molecule of bridging with the dopamine.
Of the present invention is the method for the bio-medical material surface fixed function molecule of bridging with the dopamine, may further comprise the steps:
(1) bio-medical material that will remove oxide layer and surface blot is soaked in the buffer that contains 0.1-3mg/ml dopamine, and this buffer is the Tris-HCl buffer of 10mM, and its pH value is 8.5, soaks reaction 1-36 hours, reaction temperature 4-60
oC moved in the distilled water rinsing 0.1-100 minutes then, and reuse nitrogen dries up;
(2) bio-medical material that is fixed with dopamine after will drying up is soaked in the buffer that concentration is 0.1-3mg/ml functional molecular, and this buffer is the PBS buffer, and its pH value is 7.4, reacts 1-36 hours, and soaking reaction temperature is 4-37
oC moved in the distilled water rinsing 0.1-100 minutes then, dried up with nitrogen at last, can obtain to have fixed the bio-medical material of functional molecular.
Among the present invention, said bio-medical material is a Nitinol, and cochrome gathers Acetic acid, hydroxy-, bimol. cyclic ester, polylactide, the copolymer of lactide and Acetic acid, hydroxy-, bimol. cyclic ester or pla-pcl.Said functional molecular can be heparin, hyaluronic acid, chondroitin sulfate, chitosan, aminoacid or mercaptan.Said distilled water can be a distilled water, tri-distilled water or Milli-Q water.
Among the present invention, the preferred concentration of dopamine in buffer is 1-2mg/ml.In the step of the present invention (1), bio-medical material preferred immersion response time in dopamine solution is 12-24 hours, and reaction temperature is 25-50
oC.Step (1) bio-medical material preferred rinsing time in distilled water is 10-30 minutes.
Among the present invention, the preferred concentration of functional molecular is 1-2mg/ml.In the step of the present invention (2), the bio-medical material preferred immersion response time in functional molecular that is fixed with dopamine is 12-24 hours, and reaction temperature is 25-30
oC.The bio-medical material preferred rinsing time in distilled water that is fixed with functional molecular in the step (2) is 10-30 minutes.
The present invention is the bridging medium with the dopamine; Utilize its auto polymerization characteristic stronger under the certain pH environment to be assembled into the formation of bio-medical material surface and gather the dopamine layer; Utilize chemical reaction that functional molecular is grafted to then and gather on the dopamine layer, make up the bio-medical material of biologically active.
Advantage of the present invention
This method has made full use of the auto polymerization characteristic of dopamine under certain pH conditions, through gathering dopamine and having the functional molecular generation chemical reaction of specific function, thereby reach functional molecular is fixed to the surperficial purpose of bio-medical material.This bio-medical material scope is wide, can be metal material, inorganic material, high-molecular organic material etc.This method adopts first polymerization; But stay the reactive activity group on the bio-medical material surface; The molecule of grafting fixed function does not more destroy the function of functional molecular again when solving assembling material stabilization problem, be particularly suitable for the strict bioactivation modification of functional molecular functional requirement.This method be applicable to can with the functional molecular that gathers dopamine generation chemical reaction, have the popularity of application.The tissue engineering bracket of biological activityization can be prepared through the present invention, multiple tissue engineering bracket modifications such as heart, blood vessel, skin, cartilage, bone, nerve, tendon, cardiac valve can be widely used in specific function.
Description of drawings
Fig. 1 is the variation of Nitinol surface contact angle before and after the assembling heparin
Fig. 2 a is the variation of having assembled the whole constituent content in Nitinol surface, heparin front and back
Fig. 2 b is the variation of having assembled Nitinol surface, heparin front and back Ti 2p constituent content
Fig. 2 c is the variation of having assembled Nitinol surface, heparin front and back S 2p constituent content
Fig. 3 a is the surperficial SEM figure of the Nitinol of unassembled dopamine and heparin
Fig. 3 b is the surperficial SEM figure that has assembled the Nitinol of the unassembled heparin of dopamine
Fig. 3 c is the surperficial SEM figure that has assembled the Nitinol of dopamine and heparin
Fig. 4 is the variation of the anticoagulation function of the Nitinol before and after the assembling heparin
Fig. 5 is the variation of the topological pattern of the Nitinol before and after assembling dopamine and the heparin
The specific embodiment
Below in conjunction with embodiment the present invention is elaborated.
Instance 1:
A kind of is the method for the bio-medical material surface fixed function molecule of bridging with the dopamine, may further comprise the steps:
(1) Nitinol that will remove oxide layer and surface blot is soaked in the buffer that contains the 2mg/ml dopamine, and this buffer is the Tris-HCl of 10mM, and its pH value is 8.5, soaks reaction 24 hours, reaction temperature 50
oC moved in the distilled water rinsing 20 minutes then, and reuse nitrogen dries up;
(2) Nitinol that is fixed with dopamine after will drying up is soaked in the buffer that concentration is the 2mg/ml heparin, and this buffer is the PBS buffer, and its pH value is 7.4, reacts 12 hours, and soaking reaction temperature is 37
oC.Move in the distilled water rinsing then 10 minutes, and dried up with nitrogen at last, can obtain to have fixed the Nitinol of heparin.Gather behind dopamine and the heparin respectively the generation (Fig. 1) that chemical reaction is followed the tracks of in the variation of measuring its contact angle with DSA10-MK2 type surface tension instrument thereby assembled.Visible by Fig. 1, along with the generation of assembling process, variation has also taken place in the contact angle on Nitinol surface simultaneously, has proved that these two kinds of materials successively have been assembled into the surface of Nitinol.
Thereby assembled behind the heparin generation (Fig. 2 a, Fig. 2 b, Fig. 2 c) that the changes of contents of measuring surface-element with the XPS analysis appearance is followed the tracks of chemical reaction.Visible by Fig. 2 a, Fig. 2 b, Fig. 2 c, along with the generation of assembling process, variation has also taken place in the element and the content on Nitinol surface simultaneously, has proved that these two kinds of materials successively have been assembled into the surface of Nitinol.
Assemble application scanning ultramicroscope (SEM) behind the heparin and observed the variation (Fig. 3 a, Fig. 3 b, Fig. 3 c) of its surface topography.The result shows, assembled that the Nitinol surface presents a large amount of polymer behind the heparin, proved that two kinds of materials successively have been assembled into the surface of Nitinol.
Use atomic force scanning probe microscopy (AFM) and observe the variation (Fig. 4) of its surface topology pattern before and after assembling.The result shows, assembled that obvious variation has taken place for Nitinol surface topology pattern and roughness behind the heparin, proved that two kinds of materials successively have been assembled into the surface of Nitinol.
| Sample | Ni/Ti | Ni/Ti-DA | Ni/Ti-DA-Hep |
| Roughness | 0.37 | 2.8 | 3.0 |
Adopt the multiple calcification time (PRT) of anticoagulate plasma to analyze the variation (Fig. 5) of the anticoagulation function on Nitinol surface, assembling front and back.Visible among the figure, the anticoagulation function on Nitinol surface is significantly improved behind the assembling heparin, also proved heparin successful group installed to the surface of Nitinol, and still kept good biological activity.
Instance 2:
A kind of is the method for the bio-medical material surface fixed function molecule of bridging with the dopamine, may further comprise the steps:
(1) cochrome that will remove oxide layer and surface blot is soaked in the buffer that contains the 1mg/ml dopamine, and this buffer is the Tris-HCl of 10mM, and its pH value is 8.5, soaks reaction 24 hours, reaction temperature 37
oC moved in the distilled water rinsing 30 minutes then, and reuse nitrogen dries up;
(2) cochrome that is fixed with dopamine after will drying up is soaked in the buffer that concentration is the 2mg/ml heparin, and this buffer is the PBS buffer, and its pH value is 7.4, reacts 12 hours, and soaking reaction temperature is 37
oC.Move in the distilled water rinsing then 10 minutes, and dried up with nitrogen at last, can obtain to have fixed the cochrome of heparin.
Instance 3:
A kind of is the method for the bio-medical material surface fixed function molecule of bridging with the dopamine, may further comprise the steps:
(1) Nitinol that will remove oxide layer and surface blot is soaked in the buffer that contains the 3mg/ml dopamine, and this buffer is the Tris-HCl of 10mM, and its pH value is 8.5, soaks reaction 12 hours, reaction temperature 37
oC moved in the tri-distilled water rinsing 60 minutes then, and reuse nitrogen dries up;
(2) Nitinol that is fixed with dopamine after will drying up is soaked in the buffer that concentration is the 3mg/ml heparin, and this buffer is the PBS buffer, and its pH value is 7.4, reacts 12 hours, and soaking reaction temperature is 37
oC.Move in the distilled water rinsing then 10 minutes, and dried up with nitrogen at last, can obtain to have fixed the Nitinol of heparin.
Instance 4:
A kind of is the method for the bio-medical material surface fixed function molecule of bridging with the dopamine, may further comprise the steps:
(1) the PE film that will remove oxide layer and surface blot is soaked in the buffer that contains the 0.1mg/ml dopamine, and this buffer is the Tris-HCl of 10mM, and its pH value is 8.5, soaks reaction 24 hours, reaction temperature 50
oC moved in the tri-distilled water rinsing 30 minutes then, and reuse nitrogen dries up;
(2) the PE film that is fixed with dopamine after will drying up is soaked in the buffer that concentration is the 2mg/ml heparin, and this buffer is the PBS buffer, and its pH value is 7.4, reacts 12 hours, and soaking reaction temperature is 37
oC.Move in the tri-distilled water rinsing then 20 minutes, and dried up with nitrogen at last, can obtain to have fixed the PE film of heparin.
Instance 5:
A kind of is the method for the bio-medical material surface fixed function molecule of bridging with the dopamine, may further comprise the steps:
(1) Nitinol that will remove oxide layer and surface blot is soaked in the buffer that contains the 1mg/ml dopamine, and this buffer is the Tris-HCl of 10mM, and its pH value is 8.5, soaks reaction 24 hours, reaction temperature 50
oC moved in the Milli-Q water rinsing 30 minutes then, and reuse nitrogen dries up;
(2) to be soaked in concentration be in the hyaluronic buffer of 1mg/ml to the Nitinol that is fixed with dopamine after will drying up, and this buffer is the PBS buffer, and its pH value is 7.4, reacts 12 hours, and soaking reaction temperature is 37
oC.Move in the Milli-Q water rinsing then 10 minutes, and dried up with nitrogen at last, can obtain to have fixed hyaluronic Nitinol.
Claims (10)
1. one kind is the method for the bio-medical material surface fixed function molecule of bridging with the dopamine, it is characterized in that may further comprise the steps:
(1) bio-medical material that will remove oxide layer and surface blot is soaked in the buffer that contains 0.1-3mg/ml dopamine, and this buffer is the Tris-HCl buffer of 10mM, and its pH value is 8.5, soaks reaction 1-36 hours, reaction temperature 4-60
oC moved in the distilled water rinsing 0.1-100 minutes then, and reuse nitrogen dries up;
(2) bio-medical material that is fixed with dopamine after will drying up is soaked in the buffer that concentration is 0.1-3mg/ml functional molecular, and this buffer is the PBS buffer, and its pH value is 7.4, reacts 1-36 hours, and soaking reaction temperature is 4-37
oC moved in the distilled water rinsing 0.1-100 minutes then, dried up with nitrogen at last, can obtain to have fixed the bio-medical material of functional molecular.
2. according to claim 1 is the method for the bio-medical material surface fixed function molecule of bridging with the dopamine, it is characterized in that said bio-medical material is Nitinol, cochrome, gathers the copolymer or the pla-pcl of Acetic acid, hydroxy-, bimol. cyclic ester, polylactide, lactide and Acetic acid, hydroxy-, bimol. cyclic ester.
According to claims 1 described a kind of be the method for the bio-medical material surface fixed function molecule of bridging with the dopamine, it is characterized in that the concentration of dopamine in buffer is 1-2mg/ml.
4. according to claim 1 is the method for the bio-medical material surface fixed function molecule of bridging with the dopamine, it is characterized in that step (1) bio-medical material soaks reaction 12-24 hours in dopamine solution, and reaction temperature is 25-50
oC.
5. according to claim 1 is the method for the bio-medical material surface fixed function molecule of bridging with the dopamine, it is characterized in that the rinsing 10-30 minutes in distilled water of step (1) bio-medical material.
6. according to claim 1 is the method for the bio-medical material surface fixed function molecule of bridging with the dopamine, and the concentration that it is characterized in that functional molecular is 1-2mg/ml.
7. according to claim 1 is the method for the bio-medical material surface fixed function molecule of bridging with the dopamine; It is characterized in that the bio-medical material that step (2) is fixed with dopamine soaks reaction 12-24 hours in functional molecular, reaction temperature is 25-30
oC.
8. according to claim 1 is the method for the bio-medical material surface fixed function molecule of bridging with the dopamine, it is characterized in that the rinsing 10-30 minutes in distilled water of bio-medical material that step (2) is fixed with functional molecular.
9. according to claim 1 is the method for the bio-medical material surface fixed function molecule of bridging with the dopamine, it is characterized in that said functional molecular is heparin, hyaluronic acid, chondroitin sulfate, chitosan, aminoacid or mercaptan.
According to claims 1 described a kind of be the method for the bio-medical material surface fixed function molecule of bridging with the dopamine, it is characterized in that said distilled water is a distilled water, tri-distilled water or Milli-Q water.
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