CN106237387A - A kind of manufacture method of 3 D-printing bio-ink material - Google Patents

A kind of manufacture method of 3 D-printing bio-ink material Download PDF

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Publication number
CN106237387A
CN106237387A CN201610729225.XA CN201610729225A CN106237387A CN 106237387 A CN106237387 A CN 106237387A CN 201610729225 A CN201610729225 A CN 201610729225A CN 106237387 A CN106237387 A CN 106237387A
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China
Prior art keywords
growth factor
bio
ink
manufacture method
cell
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Pending
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CN201610729225.XA
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Chinese (zh)
Inventor
周佳
厉民
王峥
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Zhejiang Provincial Peoples Hospital
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Zhejiang Provincial Peoples Hospital
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Priority to CN201610729225.XA priority Critical patent/CN106237387A/en
Publication of CN106237387A publication Critical patent/CN106237387A/en
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    • 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
    • A61L27/36—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix
    • A61L27/38—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix containing added animal cells
    • 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
    • A61L27/14—Macromolecular materials
    • A61L27/20—Polysaccharides
    • 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
    • A61L27/14—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
    • A61L27/14—Macromolecular materials
    • A61L27/22—Polypeptides or derivatives thereof, e.g. degradation products
    • A61L27/24—Collagen
    • 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
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B33—ADDITIVE MANUFACTURING TECHNOLOGY
    • B33Y—ADDITIVE MANUFACTURING, i.e. MANUFACTURING OF THREE-DIMENSIONAL [3-D] OBJECTS BY ADDITIVE DEPOSITION, ADDITIVE AGGLOMERATION OR ADDITIVE LAYERING, e.g. BY 3-D PRINTING, STEREOLITHOGRAPHY OR SELECTIVE LASER SINTERING
    • B33Y70/00—Materials specially adapted for additive manufacturing
    • 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
    • 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
    • A61L2300/252—Polypeptides, proteins, e.g. glycoproteins, lipoproteins, cytokines
    • 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
    • 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
    • A61L2300/412—Tissue-regenerating or healing or proliferative agents
    • A61L2300/414—Growth factors

Abstract

The present invention relates to the manufacture method of a kind of 3 D-printing bio-ink material, comprise the steps: under 24 DEG C of environment, cell growth factor, hydrophilic macromolecule material and deionized water are sequentially added container；Mass percent is deionized water 45 75%, hydrophilic macromolecule material 20 50%, growth factor-21 2%；Magnetic stirrer 10 20min, obtains gel aqueous fluid；Add the biological cell through counting and make suspension.The present invention has following feature and beneficial effect: the gel aqueous fluid material therefor of bio-ink is natural polymer, good biocompatibility, and catabolite safety non-toxic does not exist the potential danger of anaphylaxis or toxic reaction；Bio-ink is similar with the mechanical property of body tissue, there is not foreign body sensation；Bio-ink, in print procedure, can be controlled print density by the regulation of print parameters and printed form, it is achieved personalized printing.

Description

A kind of manufacture method of 3 D-printing bio-ink material
Technical field
The present invention relates to the manufacture method of a kind of 3 D-printing bio-ink material, particularly relate to a kind of containing biological cell With the manufacture method of the 3 D-printing bio-ink material of somatomedin, belong to technical field of biological materials.
Background technology
Three-dimensional printing technology has had the developing history of more than 30 year from the proposition of concept, and it is to use to print material Material, carrys out the technology of constructed object by the way of successively stacking accumulation, also known as increasing material manufacturing technology, and actually rapid shaping neck A kind of emerging technology in territory, is a kind of based on mathematical model file, carrys out the skill of constructed object by the way of successively printing Art.Owing to 3 D-printing is without proembryo and mould, the object of any shape, letter just directly can be generated according to computer graphics data Change the fabrication schedule of product, shorten the lead time of product, improve efficiency and reduce cost.
Along with the progress of technology, the application of 3 D-printing is the most extensive, and for biomedical sector, 3 D-printing Technology can obtain the higher medical model of fidelity, more to meet the armarium of ergonomics, application scenarios more extensively biological Medical apparatus and instruments, the higher artificial organ of personalization level or organ.Artificial organ or organ often structure is complicated, by multiple not Same cell composition, and interlaced composition function unit, thus play various function.Therefore, at manufacturing artificial tissue or device During official, need to consider biological cell is placed in the position that bionical degree is optimal, reach immediately to make the requirement that immediately can use.
Hydrogel water content based on biomacromolecule is high, and mechanical property is similar to soft tissue, has good biofacies Capacitive, conveying nutrient and the high efficiency of excretion metabolism thing and the great ability of parcel cell, these features make it by widely For building tissue engineering bracket, drug controlled release system etc..But as bio-ink, the most only solvent is inadequate, for life Thing cell can play biological function in local in the short time, in addition it is also necessary to has various nutrient substance and cytokine to provide material for it Support and ensure.
I.e. beat i.e. use in order to realize 3 D-printing artificial organ organ, build functional organization's organ list that many cells are compound Unit, simplifies and builds flow process, improves and builds efficiency, needs the simple and convenient bio-ink that can meet again above-mentioned requirements of design configurations.
Summary of the invention
It is an object of the invention to the deficiency for existing biometric print material, it is provided that a kind of by biological cell, cell growth The factor, hydrophilic macromolecule material and the bio-ink of deionized water composition.The artificial organ obtained after 3 D-printing or device Official possesses the functional unit that can play biological effect.
For realizing above-mentioned technical purpose, present invention employs techniques below scheme: this 3 D-printing bio-ink material Manufacture method, comprise the steps:
1) under 2-4 DEG C of environment, cell growth factor, hydrophilic macromolecule material and deionized water are sequentially added appearance Device；Mass percent is deionized water 45-75%, hydrophilic macromolecule material 20-50%, growth factor-21-2%；
2) magnetic stirrer 10-20min, obtains gel aqueous fluid；
3) add the biological cell through counting and make suspension.
As preferably: step 1) in, hydrophilic macromolecule material is: agarose, alginic acid, hyaluronic acid, collagen, methyl One or more in cellulose, fibroin, keratin.
As preferably: step 1) in, cell growth factor is: nerve growth factor, epidermal growth factor, bone growth because of Son, hemopoietic growth factor, endothelial cell growth factor (ECGF), glial growth factor, fibroblast growth factor, insulin-like growth A kind of or the most several in the factor, polypeptide growth factor.
As preferably: step 3) in, described biological cell is: zooblast, plant cell, fungus or antibacterial, individually freezes Exist in liquid nitrogen container, recovery of thawing when needing, wash 2-4 time with PBS after recovery, centrifugal segregation PBS, resuspended with hydrogel and count Number.
The present invention has following feature and a beneficial effect: 1) the gel aqueous fluid material therefor of bio-ink is natural polymer Son, good biocompatibility, catabolite safety non-toxic, there is not the potential danger of anaphylaxis or toxic reaction；2) biological Ink is similar with the mechanical property of body tissue, there is not foreign body sensation；3) bio-ink is in print procedure, can be joined by printing The regulation of number, controls print density and prints form, it is achieved personalization prints；4) after bio-ink has printed, can be in short-term In possess biological function, improve printing effect；5) the bio-ink configuration process of the present invention is simple, reduces operator Training difficulty.
Accompanying drawing explanation
Fig. 1 is the unit are aixs cylinder number comparison diagram of the regenerating nerve cross section of embodiment 1；
Fig. 2 is the unit are aixs cylinder number comparison diagram of the regenerating nerve cross section of embodiment 2；
Fig. 3 is the unit are aixs cylinder number comparison diagram of the regenerating nerve cross section of embodiment 3；
Fig. 4 is the unit are aixs cylinder number comparison diagram of the regenerating nerve cross section of embodiment 4.
Detailed description of the invention
Below in conjunction with embodiment, the present invention is described further.The explanation of following embodiment is only intended to help to understand this Invention.It should be pointed out that, for those skilled in the art, under the premise without departing from the principles of the invention, also The present invention can be carried out some improvement and modification, these improve and modify the protection domain also falling into the claims in the present invention In.
Embodiment 1
1) configuration of gel aqueous fluid: according to mass percent, takes hyaluronic acid 45%, epidermal growth factor 1%, ionized water 54%, under 2 DEG C of environment, above-mentioned substance is sequentially added container, magnetic stirrer 20min.
2) biological cell is thawed recovery, wash 2 times with PBS, centrifugal segregation PBS, resuspended with hydrogel and count.
3) in gel aqueous fluid, add the biological cell through counting and make suspension.
Embodiment 2
1) configuration of gel aqueous fluid: according to mass percent, takes hyaluronic acid 25%, collagen 20%, epidermal growth factor 1%, ionized water 54%, under 2 DEG C of environment, above-mentioned substance is sequentially added container, magnetic stirrer 20min.
2) biological cell is thawed recovery, wash 2 times with PBS, centrifugal segregation PBS, resuspended with hydrogel and count.
3) in gel aqueous fluid, add the biological cell through counting and make suspension.
Embodiment 3
1) configuration of gel aqueous fluid: according to mass percent, takes hyaluronic acid 25%, collagen 20%, epidermal growth factor 1%, ionized water 54%, under 2 DEG C of environment, above-mentioned substance is sequentially added container, magnetic stirrer 20min.
2) biological cell is thawed recovery, wash 3 times with PBS, centrifugal segregation PBS, resuspended with hydrogel and count.
3) in gel aqueous fluid, add the biological cell through counting and make suspension.
Embodiment 4
1) configuration of gel aqueous fluid: according to mass percent, takes methylcellulose 15%, collagen 10%, fibroin 10%, Epidermal growth factor 0.5%, insulin like growth factor 0.5%, ionized water 64%, under 2 DEG C of environment, by above-mentioned substance sequentially Add container, magnetic stirrer 15min.
2) biological cell is thawed recovery, wash 3 times with PBS, centrifugal segregation PBS, resuspended with hydrogel and count.
3) in gel aqueous fluid, add the biological cell through counting and make suspension.
The 3 D-printing bio-ink performance test of embodiment 1-4 is as follows:
After the bio-ink that embodiment 1 is configured prints on nerve trachea, it is used for repairing rat defect, and commonly leads Pipe and autologous nerve repair rat defect after 6 weeks the most as shown in Figure 1.Unit are axle to regenerating nerve cross section Prominent number counts and compares.Fig. 1 respectively organizes regenerating nerve cross section aixs cylinder count value and all there is notable difference (P < 0.05).Implement After the bio-ink that example 1 is configured prints on nerve trachea, it is used for repairing neurologic defect, obtained regenerating nerve cross section axis Though prominent count value is not as good as autologous nerve group (P < 0.05), but is better than and simple conduit set (P < 0.05).
After the bio-ink that embodiment 2 is configured prints on nerve trachea, it is used for repairing rat defect, and commonly leads Pipe and autologous nerve repair rat defect after 6 weeks the most as shown in Figure 2.Unit are axle to regenerating nerve cross section Prominent number counts and compares.Fig. 2 respectively organizes regenerating nerve cross section aixs cylinder count value and all there is notable difference (P < 0.05).Implement After the bio-ink that example 2 is configured prints on nerve trachea, it is used for repairing neurologic defect, obtained regenerating nerve cross section axis Though prominent count value is not as good as autologous nerve group (P < 0.05), but is better than and simple conduit set (P < 0.05).
After the bio-ink that embodiment 3 is configured prints on nerve trachea, it is used for repairing rat defect, and commonly leads Pipe and autologous nerve repair rat defect after 6 weeks the most as shown in Figure 3.Unit are axle to regenerating nerve cross section Prominent number counts and compares.Fig. 3 respectively organizes regenerating nerve cross section aixs cylinder count value and all there is notable difference (P < 0.05).Implement After the bio-ink that example 3 is configured prints on nerve trachea, it is used for repairing neurologic defect, obtained regenerating nerve cross section axis Though prominent count value is not as good as autologous nerve group (P < 0.05), but is better than and simple conduit set (P < 0.05).
After the bio-ink that embodiment 4 is configured prints on nerve trachea, it is used for repairing rat defect, and commonly leads Pipe and autologous nerve repair rat defect after 6 weeks the most as shown in Figure 4.Unit are axle to regenerating nerve cross section Prominent number counts and compares.Fig. 4 respectively organizes regenerating nerve cross section aixs cylinder count value and all there is notable difference (P < 0.05).Implement After the bio-ink that example 4 is configured prints on nerve trachea, it is used for repairing neurologic defect, obtained regenerating nerve cross section axis Though prominent count value is not as good as autologous nerve group (P < 0.05), but is better than and simple conduit set (P < 0.05).

Claims (4)

1. the manufacture method of a 3 D-printing bio-ink material, it is characterised in that comprise the steps:
1) under 2-4 DEG C of environment, cell growth factor, hydrophilic macromolecule material and deionized water are sequentially added container；Matter Amount percentage ratio is deionized water 45-75%, hydrophilic macromolecule material 20-50%, growth factor-21-2%；
2) magnetic stirrer 10-20min, obtains gel aqueous fluid；
3) add the biological cell through counting and make suspension.
The manufacture method of 3 D-printing bio-ink material the most according to claim 1, it is characterised in that: step 1) in, Hydrophilic macromolecule material is: in agarose, alginic acid, hyaluronic acid, collagen, methylcellulose, fibroin, keratin Plant or several.
The manufacture method of 3 D-printing bio-ink material the most according to claim 1, it is characterised in that: step 1) in, Cell growth factor is: nerve growth factor, epidermal growth factor, SGF, hemopoietic growth factor, endothelial growth factor One in son, glial growth factor, fibroblast growth factor, insulin like growth factor, polypeptide growth factor Or it is the most several.
The manufacture method of 3 D-printing bio-ink material the most according to claim 1, it is characterised in that: step 3) in, Described biological cell is: zooblast, plant cell, fungus or antibacterial, the most frozen in liquid nitrogen container, thaws multiple when needing Soviet Union, washs 2-4 time with PBS after recovery, centrifugal segregation PBS, resuspended with hydrogel and count.
CN201610729225.XA 2016-08-24 2016-08-24 A kind of manufacture method of 3 D-printing bio-ink material Pending CN106237387A (en)

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Cited By (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN107744602A (en) * 2017-09-30 2018-03-02 广东泰宝医疗器械技术研究院有限公司 A kind of preparation method of bio-ink material available for 3D printing
CN107998449A (en) * 2017-12-15 2018-05-08 杭州捷诺飞生物科技股份有限公司 A kind of 3D printing high intensity bio-ink material
CN109957539A (en) * 2017-12-14 2019-07-02 深圳先进技术研究院 A kind of artificial islet tissue and its preparation and application
CN112931849A (en) * 2021-02-09 2021-06-11 南京农业大学 Food leavening agent based on 3D printing and preparation method and application thereof
CN113677505A (en) * 2019-01-24 2021-11-19 中国科学院遗传与发育生物学研究所 Three-dimensional biological printing system and method

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN103977453A (en) * 2014-05-28 2014-08-13 段升华 3D (three-dimensional) bioprinting hydrogel material and application thereof
WO2015002707A1 (en) * 2013-05-28 2015-01-08 The Johns Hopkins University Bone regeneration using stromal vascular fraction. platelet-derived growth factor-rich hydrogel, three dimensional printed poly-epsilon-caprolactone scaffolds

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2015002707A1 (en) * 2013-05-28 2015-01-08 The Johns Hopkins University Bone regeneration using stromal vascular fraction. platelet-derived growth factor-rich hydrogel, three dimensional printed poly-epsilon-caprolactone scaffolds
CN103977453A (en) * 2014-05-28 2014-08-13 段升华 3D (three-dimensional) bioprinting hydrogel material and application thereof

Cited By (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN107744602A (en) * 2017-09-30 2018-03-02 广东泰宝医疗器械技术研究院有限公司 A kind of preparation method of bio-ink material available for 3D printing
CN109957539A (en) * 2017-12-14 2019-07-02 深圳先进技术研究院 A kind of artificial islet tissue and its preparation and application
CN107998449A (en) * 2017-12-15 2018-05-08 杭州捷诺飞生物科技股份有限公司 A kind of 3D printing high intensity bio-ink material
CN113677505A (en) * 2019-01-24 2021-11-19 中国科学院遗传与发育生物学研究所 Three-dimensional biological printing system and method
CN113677505B (en) * 2019-01-24 2023-08-18 中国科学院遗传与发育生物学研究所 Three-dimensional bioprinting system and method
CN112931849A (en) * 2021-02-09 2021-06-11 南京农业大学 Food leavening agent based on 3D printing and preparation method and application thereof
CN112931849B (en) * 2021-02-09 2023-07-07 南京农业大学 Food leavening agent based on 3D printing and preparation method and application thereof

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