CN101156967A - Preparation and usage of fibroin albumen antipriming pipe - Google Patents
Preparation and usage of fibroin albumen antipriming pipe Download PDFInfo
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- CN101156967A CN101156967A CNA200710177104XA CN200710177104A CN101156967A CN 101156967 A CN101156967 A CN 101156967A CN A200710177104X A CNA200710177104X A CN A200710177104XA CN 200710177104 A CN200710177104 A CN 200710177104A CN 101156967 A CN101156967 A CN 101156967A
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Abstract
The preparation and the purpose of a silk fibroin protein porous duct relates to the biological material field, in particular to the tissue engineering scaffold field. The invention is to arrange silk fibroin aqueous solution with 30 percent to 50 percent of high concentration in a vessel, the solution is formed into silk fibroin fiber with the diameter of 200 nm to 3000 nm under the function of electrostatic force with 500 V to 60 kV voltage, the fiber is received by a revolving metal rod or tube, the receiving distance is in 5 to 100 cm, and after 5-480 minutes, the silk fibroin porous duct with the diameter of 1 to 10 mm, the wall thickness of 0.01 to 1 mm and the length not lower than 30 cm can be obtained. Then the silk fibroin porous duct is processed with 90 to 10 percent of methyl alcohol aqueous solution to lead the silk fibroin porous duct to be denaturized, after the vacuum drying in the normal temperature, the non water-soluble silk fibroin protein porous duct is obtained. The diameter of the silk fibroin duct of the invention is controllable, the invention has the biological compatibility and the porosity which are beneficial to the cells to grow, and can be used as a tissue engineering scaffold, the preparation process is easy and feasible, the cost is low, and the commercialization production can be realized.
Description
Technical field
The present invention relates to technical field of biological material for preparation has the new method of the fibroin albumen antipriming pipe of good biocompatibility, is specifically related to the tissue engineering bracket field.
Background technology
In recent years, the onset of illness of vascular occlusive diseases such as arteriosclerosis is cumulative many, and the patient that need carry out vascular transplant is also more and more.Because the rejection of allosome blood vessel and from the finiteness of body blood vessel, the used blood vessel multi-source of transplant operation is in artificial blood vessel.At present, diameter has particularly been realized commercialization greater than 10 millimeters artificial blood vessel greater than 6 millimeters, then become an international difficult problem less than 6 millimeters small-caliber vasculars, its main cause is the formation of thrombosis and thickening of new intima, causes blood vessel blockage, and makes graft failure.In the artificial blood vessel of medium-small diameter, past many uses ePTFE (expanded polytetrafluoroethylsealing) and Dacron (terylene), but its vascular compliance is relatively poor, and clinical data shows that ePTFE artificial blood vessel patency rate only is 30%.
U.S. natural science fund engineering council had proposed the organizational project notion in the biological engineering seminar in 1987, for the preparation of small-caliber vascular provides a new thinking, promptly " principle of application project and zoobiotism and method are understood organizational structure-function relationship of the mammal of normal and pathology; and develop biological tissue's succedaneum, and be used to repair, keep and improve the function of tissue ".For blood vessel and since synthetic material in vivo life-time service can bring negative effect to human body, prepare a kind of original small-caliber vascular particularly necessity that seems of repairing and substitute.The ultimate principle and the method for organizational project be, it is good to be adsorbed in a kind of biocompatibility after the high concentration histiocyte amplification with In vitro culture, and can progressively on the extracellular matrix of degraded and absorbed (ECM) material, be formed cell-biomaterial composites by human body.This material can be the three dimensions that cell provides existence, helps cell and obtains enough nutrient substance, carries out metabolism, makes the three-dimensional rack growth of cell by prefabricated form; Then with this species complex implanting to human body desired area.The cell of plantation continues the hypertrophy breeding in biological support degraded and absorbed process, formed new respective organization with original specific function and form and organ, thereby can reach the purpose of repair tissue profile and reconstruction.Ideal tissue engineering bracket material or cell carrier should have following condition: 1. excellent biological compatibility; 2. Shi Yi biological degradability; 3. effective surface activity; 4. certain plasticity; 5. has three-dimensional porous structure.Organize often to be the three dimensions configuration in vivo, appropriate nutrition condition and steric requirements are provided for the growth of cell, help the physiological function of organizing performance specific.Therefore, implant design should mainly be considered the steric configuration of carrier material, as the size in hole, the connectedness between the hole, the surface area of material internal etc.The open-cell material surface can more help seeking connections with of cell pseudopodium than common material surface, increases the physical force between cellular layer and the base material.Perforate on the base material helps the transportation and the exchange of moisture, inorganic salt and other nutrient substance and products of cellular metabolism, thereby more helps the normal growth and the physiological metabolism of cell.In order to carry out the In vitro culture of multi-layer cellular, the cell carrier material that how to prepare three-dimensional porous structure has become the focus of material science research.At present, the primary structure form in the organizational project of being applied in has two kinds of fibrous framework and porous foams.
The preparation method of tissue engineering bracket mainly contains the fibrage method, salting out method, gas foaming method, melt molding method, methods such as phase separation method, Freeze Drying Technique and electrostatic spinning.For the intravascular tissue engineering support, not only need material to possess excellent biological compatibility, also need in the practical application support is prepared into and be three-dimensional tubing.At present, the method for preparing the perforated tubular support mostly is weave, low-temperature extrusion forming, the cylindric back spraying of perforated membrane coiled polymer solution, lyophilization molding in the mould.Electrostatic spinning has its unique advantage as a kind of porous material for preparing, its process is: solution is divided into nanometer to micron-sized fiber under the effect of electric field force, solvent vapors away in the process that arrives receiving system, and resulting fiber overlaps mutually on the receiving system surface and forms porous material.Recently, the method with electrostatic spinning prepares the existing report of gene recombination spider silk fibroin tubular bracket.
Fibroin albumen is as natural biological albumen, and with its excellent biocompatibility, low inflammatory reaction and good mechanical performance are subjected to biomaterial circle extensive studies.At biomedical sector, fibroin is as the operation suture thread history of existing centuries, at artificial skin, prosthetic ligament, artificial bone, artificial blood vessel, medicament slow release material and enzyme immobilization material aspect have obtained a series of basic research achievement, have shown huge application potential.Because the fibroin albumen side chain has the cell growth and sticks factor, and can make nanoscale to micron-sized azelon,, therefore also obtain extensive studies in field of tissue engineering technology with the in-vivo tissue structural similarity by regeneration.
Fibroin albumen not only has excellent biocompatibility, and cheap and easy to get, makes that fibroin albumen is subjected to paying close attention to more widely, is more conducive to commercially produce.Fibroin albumen comprises electrostatic spinning by various method, lyophilization, and saltings out method etc. are prepared into porous support, but the preparation of fibroin albumen antipriming pipe is not still reported.Because the special physicochemical property of fibroin albumen is difficult to prepare uniform tubing by mould.And after deliberation more comprehensive of the electrostatic spinning of fibroin albumen, and studies have shown that the fibroin albumen non-woven fabrics by the method for electrostatic spinning preparation is suitable for the cell growth and breeding.
This paper is prepared into antipriming pipe by direct method with electrostatic spinning with the fibroin aqueous solution.This method synthesis the spinning of fibroin aqueous solution advantage and avoided use by cross-linking agent in the film tubulation process, make preparation process simple.The fibroin pipe caliber of preparing is controlled, has excellent biological compatibility and porous concurrently and is beneficial to the cell growth, is expected to be used for scaffold for vascular tissue engineering.
Summary of the invention
The objective of the invention is in order to prepare the fibroin albumen antipriming pipe support, to utilize the fibroin albumen excellent biological compatibility, and the high-specific surface area that electricity spinning fibre had, thereby the exploitation fibroin albumen antipriming pipe comprises intravascular tissue engineering support, tracheal tissue's engineering rack, urethral tissue engineering rack and other tubular brackets in the application in tissue engineering bracket field.
The present invention is achieved in that
The same patent of silk fibroin water solution " preparation of insoluble fibroin albumen thin film and fibroin albumen pipe " (patent No. is 03131142.3) of preparation high concentration, promptly White Steam Filature Yarns is through coming unstuck, dissolving, dialysis, sucking filtration, concentrating and obtain the silk fibroin water solution that concentration is 30%-50% (w/w).
The present invention places container with the silk fibroin water solution of 30%-50% (w/w) high concentration, solution is under the electrostatic force of 500V-60kV at voltage, forming diameter is the fibroin fiber of 200nm-3000nm, metal bar or pipe with rotation receive silk fiber, receiving range is 5-100cm, just can obtain caliber behind the 5-480min on metal bar is 1-10mm, wall thickness is 0.01-1mm, length is not less than the fibroin antipriming pipe of 30cm, use 90%-100% (v/v) methanol aqueous solution to handle then and make its degeneration, behind the room temperature vacuum drying, obtain non-water-soluble fibroin albumen antipriming pipe.The diameter of employed metal bar or pipe and length are adjustable as required.Prepared fibroin albumen antipriming pipe is used for tissue engineering bracket, comprises intravascular tissue engineering support, tracheal tissue's engineering rack, urethral tissue engineering rack and other tubular brackets.
The fibroin albumen antipriming pipe of preparing is got sample its microscopic appearance of sem observation wherein, uses Fourier transform infrared spectroscopy, and X-ray diffraction is measured the structure of fibroin albumen in the material, and tubing is carried out mechanics property analysis.Test result shows that fibroin albumen is mainly the random coil structure in the material, the dry state hot strength is not less than 0.37MPa, elongation at break 23.3%, carry out with methanol that silk II structure increases after the post processing, material is water insoluble, and hot strength is 1.17MPa under the hygrometric state, elongation at break 82.2%, elastic modelling quantity is 3.6MPa, has demonstrated good toughness.
Advantage of the present invention is:
1. directly prepare fibroin albumen antipriming pipe, avoided the use of organic binder bond and the problem of fibroin Mould Machining difficulty.
2. adopt the method for electrostatic spinning, technology is simple, and consumption is few, easily realizes commodity production.
3. silk fibroin water solution spinning, safety and environmental protection is avoided the use of organic solvent, good biocompatibility, and cost is low.
4. the fibroin albumen antipriming pipe of preparing has high porosity, and the hole has good connectedness, is beneficial to the cell growth, the nutrient substance transmission.
So the application on biomedicine is significant and practical value to fibroin albumen antipriming pipe in the present invention.
Description of drawings
Fig. 1-be the macrograph of fibroin albumen antipriming pipe of the present invention.
The specific embodiment
Embodiment 1:
With the Na of silkworm silk at 0.5% (w/w)
2CO
3Boil 60min in the solution under 100 ℃ to slough the sericin on silkworm silk top layer, the fibroin albumen after coming unstuck repeatedly washs after drying with distilled water, obtains pure fibroin albumen.A certain amount of fibroin albumen is dissolved in CaCl
2/ H
2O/CH
3CH
2In OH (mol ratio is 1: 8: 2) ternary solution or other dense saline solution, filtering solution promptly obtained the silk fibroin water solution that concentration is 3% (w/w) to remove calcium chloride and the ethanol in the solution in three days in dialysis in distilled water.
Embodiment 2:
Prepare pure silk fibroin water solution shown in embodiment 1.
1. spinning liquid preparation: 500mL pure silk fibroin aqueous solution is at 70 ℃ of following heated and stirred 18h, and mixing speed is 50rpm, obtains concentration and be 30% silk fibroin water solution.
2. electrostatic spinning process: the employing diameter is 4mm, length is that the metal bar rotation of 300mm receives, rotary speed is 5rpm, receiving range is 20cm, syringe needle place voltage is 20kV, and spinning speed is 0.15mL/min, behind the 2h, receive filament diameter on the metal bar about 700nm, thickness is about the puffy fibroin fiber of 0.4mm.
3. antipriming pipe forms: the puffy fibroin fiber places 90% methanol solution to handle 1h, and it is 0.2mm that room temperature vacuum drying 24h obtains thickness, and bore is the fibroin albumen antipriming pipe of 4mm.
Embodiment 3:
Prepare pure silk fibroin water solution shown in embodiment 1.
1. spinning liquid preparation: 500mL pure silk fibroin aqueous solution is at 70 ℃ of following heated and stirred 22h, and mixing speed is 50rpm, obtains concentration and be 50% silk fibroin water solution.
2. electrostatic spinning process: the employing diameter is 4mm, length is that the metal bar rotation of 300mm receives, rotary speed is 5rpm, receiving range is 20cm, syringe needle place voltage is 20kV, and spinning speed is 0.15mL/min, behind the 2h, receiving filament diameter on the metal bar is 2500nm, and thickness is about the puffy fibroin fiber of 0.4mm.
3. antipriming pipe forms: the puffy fibroin fiber places 90% methanol solution to handle 1h, and it is 0.2mm that room temperature vacuum drying 24h obtains thickness, and bore is the fibroin albumen antipriming pipe of 4mm.
Embodiment 4:
Prepare pure silk fibroin water solution shown in embodiment 1.
1. spinning liquid preparation: pure silk fibroin aqueous solution is at 70 ℃ of following heated and stirred 20h, and mixing speed is 50rpm, obtains concentration and be 40% silk fibroin water solution.
2. electrostatic spinning process: the employing diameter is 1mm, length is that the metal bar rotation of 300mm receives, rotary speed is 5rpm, receiving range is 20cm, syringe needle place voltage is 20kV, and spinning speed is 0.15mL/min, behind the 2h, receive filament diameter on the metal bar about 2000nm, thickness is about the puffy fibroin fiber of 0.4mm.
3. antipriming pipe forms: the puffy fibroin fiber places 90% methanol solution to handle 1h, and room temperature vacuum drying 24h obtains thickness and is about 0.2mm, and internal diameter is the fibroin albumen antipriming pipe of 1mm.
Embodiment 5:
Prepare pure silk fibroin water solution shown in embodiment 1.
1. spinning liquid preparation: pure silk fibroin aqueous solution is at 70 ℃ of following heated and stirred 20h, and mixing speed is 50rpm, obtains concentration and be 40% silk fibroin water solution.
2. electrostatic spinning process: the employing diameter is 10mm, length is that the metal bar rotation of 300mm receives, rotary speed is 5rpm, receiving range is 20cm, syringe needle place voltage is 20kV, spinning speed is 0.15mL/min, behind the 2h, receives the puffy fibroin fiber that thickness is about 0.4mm on the metal bar.
3. antipriming pipe forms: the puffy fibroin fiber places 90% methanol solution to handle 1h, and room temperature vacuum drying 24h obtains thickness and is about 0.2mm, and internal diameter is the fibroin albumen antipriming pipe of 10mm.
Embodiment 6:
Prepare pure silk fibroin water solution shown in embodiment 1.
1. spinning liquid preparation: pure silk fibroin aqueous solution is at 70 ℃ of following heated and stirred 20h, and mixing speed is 50rpm, obtains concentration and be 40% silk fibroin water solution.
2. electrostatic spinning process: the employing diameter is 4mm, and length is that the metal bar rotation of 20mm receives, and rotary speed is 5rpm, receiving range is 20cm, and syringe needle place voltage is 20kV, and spinning speed is 0.15mL/min, behind the 2h, receive the puffy fibroin fiber that thickness is about 0.4mm on the metal bar.
3. antipriming pipe forms: the puffy fibroin fiber places 90% methanol solution to handle 1h, and obtaining internal diameter behind the room temperature vacuum drying is 4mm, and thickness is about 0.2mm, and length is the fibroin albumen antipriming pipe of 20mm.(see figure 1)
Embodiment 7:
Prepare pure silk fibroin water solution shown in embodiment 1.
1. spinning liquid preparation: pure silk fibroin aqueous solution is at 70 ℃ of following heated and stirred 20h, and mixing speed is 50rpm, obtains concentration and be 40% silk fibroin water solution.
2. electrostatic spinning process: diameter is 4mm, and length is that the metal bar rotation of 300mm receives, and rotary speed is 5rpm, receiving range is 20cm, and syringe needle place voltage is 20kV, and spinning speed is 0.15mL/min, behind the 2h, receive the puffy fibroin fiber that thickness is about 0.4mm on the metal bar.
3. antipriming pipe forms: the puffy fibroin fiber places 90% methanol solution to handle 1h, obtains thickness behind the room temperature vacuum drying and is about 0.2mm, and internal diameter is the fibroin albumen antipriming pipe of 4mm.
Embodiment 8:
Prepare pure silk fibroin water solution shown in embodiment 1.
1. spinning liquid preparation: pure silk fibroin aqueous solution is at 70 ℃ of following heated and stirred 20h, and mixing speed is 50rpm, obtains concentration and be 40% silk fibroin water solution.
2. electrostatic spinning process: diameter is 4mm, and length is that the metal bar rotation of 300mm receives, and rotary speed is 5rpm, receiving range is 20cm, and syringe needle place voltage is 20kV, and spinning speed is 0.15mL/min, behind the 5min, receive the puffy fibroin fiber that thickness is about 0.01mm on the metal bar.
3. antipriming pipe forms: the puffy fibroin fiber places methanol solution to handle 1h, and obtaining internal diameter behind the room temperature vacuum drying is 4mm, and thickness is about 0.005mm, and length is the fibroin albumen antipriming pipe of 300mm.
Embodiment 9:
Prepare pure silk fibroin water solution shown in embodiment 1.
1. spinning liquid preparation: pure silk fibroin aqueous solution is at 70 ℃ of following heated and stirred 20h, and mixing speed is 50rpm, obtains concentration and be 40% silk fibroin water solution.
2. electrostatic spinning process: diameter is 4mm, and length is that the metal bar rotation of 300mm receives, and rotary speed is 5rpm, receiving range is 20cm, and syringe needle place voltage is 20kV, and spinning speed is 0.15mL/min, behind the 480min, receive the puffy fibroin fiber that thickness is about 1mm on the metal bar.
3. antipriming pipe forms: the puffy fibroin fiber places methanol solution to handle 1h, and obtaining internal diameter behind the room temperature vacuum drying is 4mm, and thickness is about 0.5mm, and length is the fibroin albumen antipriming pipe of 300mm.
Claims (4)
1. the preparation of fibroin albumen antipriming pipe and purposes, it is characterized in that: the silk fibroin water solution of 30%-50% (w/w) high concentration is placed container, solution is under the electrostatic force of 500V-60kV at voltage, forming diameter is the fibroin fiber of 200nm-3000nm, metal bar or pipe with rotation receive silk fiber, receiving range is 5-100cm, just can obtain caliber behind the 5-480min on metal bar is 1-10mm, wall thickness is 0.01-1mm, length is not less than the fibroin antipriming pipe of 30cm, use 90%-100% (v/v) methanol aqueous solution to handle then and make its degeneration, behind the room temperature vacuum drying, obtain non-water-soluble fibroin albumen antipriming pipe.
2. the preparation of fibroin albumen antipriming pipe and purposes, it is characterized in that: prepared fibroin albumen antipriming pipe is used for tissue engineering bracket.
3. the preparation of fibroin albumen antipriming pipe as claimed in claim 1 and purposes, it is characterized in that: the diameter of employed metal bar or pipe and length are adjustable as required.
4. the preparation of fibroin albumen antipriming pipe as claimed in claim 2 and purposes, it is characterized in that: tissue engineering bracket comprises the intravascular tissue engineering support, tracheal tissue's engineering rack, urethral tissue engineering rack and other tubular bracket.
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| CNB200710177104XA CN100548391C (en) | 2007-11-09 | 2007-11-09 | The preparation method of fibroin albumen antipriming pipe |
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| CN100548391C CN100548391C (en) | 2009-10-14 |
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Cited By (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101879330A (en) * | 2010-06-02 | 2010-11-10 | 苏州大学 | Small-caliber silk fibroin tubular material and preparation method thereof |
| CN101579246B (en) * | 2009-05-31 | 2010-12-08 | 苏州大学 | Artificial silk fibroin nano-fiber nerve repair conduit and preparation method thereof |
| CN102488926A (en) * | 2011-12-16 | 2012-06-13 | 东华大学 | Tissue engineering scaffold for urethra reconstruction and preparation method thereof |
| CN102652841A (en) * | 2012-04-17 | 2012-09-05 | 北京航空航天大学 | Method for preparing sulfated silk fibroin vascular tissue engineering scaffold by using electrostatic spinning |
| CN103223192A (en) * | 2013-04-03 | 2013-07-31 | 浙江大学 | Preparation method of composite material of silk fibroin and cyclodextrin |
| CN103505760A (en) * | 2012-06-29 | 2014-01-15 | 复旦大学附属华山医院 | Airway epithelium-porous silk fibroin protein complex, and preparation method and application thereof |
| CN104018245A (en) * | 2014-06-13 | 2014-09-03 | 东华大学 | Preparation method of tubular silk fibroin/keratin composite nanofiber material |
| WO2018156613A1 (en) * | 2017-02-21 | 2018-08-30 | Trustees Of Tufts College | Silk fibroin tracheal stent |
-
2007
- 2007-11-09 CN CNB200710177104XA patent/CN100548391C/en not_active Expired - Fee Related
Cited By (12)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101579246B (en) * | 2009-05-31 | 2010-12-08 | 苏州大学 | Artificial silk fibroin nano-fiber nerve repair conduit and preparation method thereof |
| CN101879330A (en) * | 2010-06-02 | 2010-11-10 | 苏州大学 | Small-caliber silk fibroin tubular material and preparation method thereof |
| CN101879330B (en) * | 2010-06-02 | 2013-04-10 | 苏州大学 | Small-caliber silk fibroin tubular material and preparation method thereof |
| CN102488926A (en) * | 2011-12-16 | 2012-06-13 | 东华大学 | Tissue engineering scaffold for urethra reconstruction and preparation method thereof |
| CN102652841A (en) * | 2012-04-17 | 2012-09-05 | 北京航空航天大学 | Method for preparing sulfated silk fibroin vascular tissue engineering scaffold by using electrostatic spinning |
| CN103505760A (en) * | 2012-06-29 | 2014-01-15 | 复旦大学附属华山医院 | Airway epithelium-porous silk fibroin protein complex, and preparation method and application thereof |
| CN103505760B (en) * | 2012-06-29 | 2015-05-20 | 复旦大学附属华山医院 | Airway epithelium-porous silk fibroin protein complex, and preparation method and application thereof |
| CN103223192A (en) * | 2013-04-03 | 2013-07-31 | 浙江大学 | Preparation method of composite material of silk fibroin and cyclodextrin |
| CN103223192B (en) * | 2013-04-03 | 2016-04-20 | 浙江大学 | The preparation method of a kind of fibroin albumen and cyclodextrin composite |
| CN104018245A (en) * | 2014-06-13 | 2014-09-03 | 东华大学 | Preparation method of tubular silk fibroin/keratin composite nanofiber material |
| CN104018245B (en) * | 2014-06-13 | 2016-11-02 | 东华大学 | A kind of preparation method of fibroin albumen/keratin composite nano fiber tubular material |
| WO2018156613A1 (en) * | 2017-02-21 | 2018-08-30 | Trustees Of Tufts College | Silk fibroin tracheal stent |
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