CN105708858A - Preparation method of growth-factor-platelet-rich fibrin and releasate - Google Patents
Preparation method of growth-factor-platelet-rich fibrin and releasate Download PDFInfo
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- CN105708858A CN105708858A CN201410730708.2A CN201410730708A CN105708858A CN 105708858 A CN105708858 A CN 105708858A CN 201410730708 A CN201410730708 A CN 201410730708A CN 105708858 A CN105708858 A CN 105708858A
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Abstract
The invention discloses a preparation method of a growth-factor-platelet-rich fibrin and releasate; the preparation method includes the following steps: providing mammalian autologous blood, and placing the blood in a container; shaking the blood; centrifuging the blood and then allowing to be static, making the blood separated into three layers of substances in the container, wherein the upper layer and the lower layer are liquid substances and the middle layer is condensed into a bulk substance; taking out the bulk substance, and allowing the bulk substance to stand until a liquid substance is released; and separating to take the liquid substances, namely the growth-factor-platelet-rich fibrin and releasate. According to the preparation method of the growth-factor-platelet-rich fibrin and releasate, through the steps of shaking the blood and standing, the prepared growth-factor-platelet-rich fibrin can has higher-concentration growth factors compared with a studied or known platelet-rich fibrin.
Description
Technical field
The present invention is about a kind of platelet fibrin rich in somatomedin and release liquid (Growth-Factor-Platelet-RichFibrinandreleasate thereof, GFPRF) preparation method, especially a kind of compared to existing rich in hematoblastic fibrin (Platelet-RichFibrin, PRF), there is the platelet fibrin rich in somatomedin of the somatomedin (GrowthFactor) of higher concentration and the preparation method of release liquid thereof.
Background technology
In the past, when doctor carries out the process of wound or surgical wound, going, other than with antibiotic, inflammation of avoiding infection, the healing of wound then can only rely on the reparation power of patient self.But, along with the appearance of autologous platelet concentrate, accelerate healing rate no longer simply dream.Autologous platelet concentrate has multiple somatomedin, cytohormone and reactive protein etc., it is possible to puts wound into or adds in implant, with the healing of accelerated in wounds.
Progress along with research, autologous platelet concentrate is from simple Platelet Concentrate, to need to by the thrombin of cattle and calcium chloride cause platelet activation rich in hematoblastic blood plasma (Platelet-RichPlasma, PRP), developing deeply be do not need other additives rich in hematoblastic fibrin.Platelet contains many biological activity proteins, including several somatomedin, has the effect promoting that stem cell generates with the movement of repair cell, hypertrophy, differentiation, new vascular generation and collagen protein, therefore can accelerating wound healing.When hematoblastic concentration is higher, somatomedin the more, not only can more accelerate healing rate, for the treatment of complicated wound, is also very helpful.Additionally, can also be used in the reparation of other biological body soft tissue and sclerous tissues rich in hematoblastic fibrin, for instance for the cosmetic surgery of autologous fat transplantation, or dentistry promotes in the middle of the technology of osteanagenesis.
At present rich in hematoblastic fibrinous preparation method, it is mostly, by the venous blood of mammal autologous offer about 10 ~ 60cc, to obtain then through centrifugal treating, and there is following advantage: be entirely autologous product, so there is no the exclusive problem of immunizing antigen antibody；During centrifugal blood, the fibrin matrix that self-assembling formation is fine and close, relatively there is tolerance；Platelet, leukocyte and cytohormone are stumbled betwixt by highly dense fibrin matrix, and platelet can disengage somatomedin and reactive protein for a long time at leisure, and can support and the relevant signaling release with cytohormone of healing；The leukocyte stumbled at fine and close fibrin matrix, has the effect to infection and immunomodulating aspect.
Summary of the invention
In view of the above problems, present invention generally provides a kind of platelet fibrin rich in somatomedin and the preparation method of release liquid thereof, especially one is compared to existing rich in hematoblastic fibrin, has the platelet fibrin rich in somatomedin of the somatomedin of higher concentration and the preparation method of release liquid thereof.
In order to reach object above, the present invention provides a kind of platelet fibrin rich in somatomedin and the preparation method of release liquid thereof, comprises the steps of the blood providing mammal autologous, is placed in container；Blood is rocked；Blood is carried out one centrifugal after static, make blood be separated into three layers material in a reservoir, it is liquid with lower floor at the middle and upper levels, and intermediate layer is then condensed into a cake mass, is the platelet fibrin rich in somatomedin；Take out cake mass, and cake mass is stood, treat that it discharges liquid；And, separate and take out liquid, be the fibrinoplatelet release liquid rich in somatomedin.
In one embodiment of the invention, wherein container be shaped as elongated, and there is two ends and an interlude, and blood rocked, refer to the two ends of the container fulcrum relative to interlude, repeat upper and lower straight line and back and forth or draw the motion of circle.
In one embodiment of the invention, the time wherein rocking blood is 45 to 90 seconds, it is advantageous to be 60 seconds.
In one embodiment of the invention, back and forth or the number of times of the motion drawing circle is 30 to 60 times wherein to repeat upper and lower straight line.
In one embodiment of the invention, the wherein centrifugal time is 9 to 12 minutes, it is advantageous to be 10 minutes.
In one embodiment of the invention, wherein centrifugal rotating speed is 2800 to 3200rpm, it is advantageous to for 3000rpm.
In one embodiment of the invention, wherein take out cake mass, be utilize aseptic tweezers to be pressed from both sides out by cake mass.
In one embodiment of the invention, wherein container is serum separator tube (SerumSeparationTube), it is advantageous to for camouflage color first-bornization blood taking tube, for instance be made by BD company, model is the blood taking tube of 367988.It addition, container before placing blood, can first preset coagulant and gel (clotactivatorandgel).
In one embodiment of the invention, wherein being stood by cake mass, refer to and be placed in test tube by cake mass, and be placed in warm water by test tube, the temperature of warm water is preferably 37 degree Celsius.
In one embodiment of the invention, it it is wherein 2 to 5 hours by the time that this cake mass is statically placed in, it is advantageous to be 3 hours.
In one embodiment of the invention, wherein comprise following somatomedin rich in the fibrin platelet of somatomedin and release liquid thereof, it is and comprises platelet derived growth factor (Platelet-derivedgrowthfactor, PDGF), somatomedin transition (transforminggrowthfactor), VEGF (Vascularendothelialgrowthfactor, VEGF), epidermal growth factor (Epidermalgrowthfactor, EGF), insulin like growth factor (Insulin-likegrowthfactor1, IGF-1), fiber mother cell growth factor (Fibroblastgrowthfactor, and nerve growth factor (Nervegrowthfactor FGF), NGF).
Effect of the present invention is in that, the platelet fibrin rich in somatomedin that disclosed herein and the preparation method of release liquid thereof, through the step rocking blood and standing, make the platelet fibrin rich in somatomedin that it is prepared, compared to practise or former times know rich in hematoblastic fibrin, it is possible to there is the somatomedin of higher concentration.
Accompanying drawing explanation
Figure 1A ~ 1B: in the fibrin platelet rich in somatomedin that disclosed herein and the preparation method of release liquid thereof, the explanation embodiment of slight wobble blood.
Fig. 2: in the fibrin platelet rich in somatomedin that disclosed herein and the preparation method of release liquid thereof, carry out standing the relation schematic diagram of somatomedin concentration in liquid and time of repose during step.
Fig. 3: the fibrin platelet rich in somatomedin that disclosed herein and the flow chart of steps of the preparation method of release liquid thereof.
Description of reference numerals:
210 centrifuge tubes；
220 fulcrums；
230,240 direction of motion；
250 blood；
260 coagulant and gel；
S510 ~ S550 step.
Detailed description of the invention
Below in conjunction with the drawings and specific embodiments, the invention will be further described, so that those skilled in the art can be better understood from the present invention and can be practiced, but illustrated embodiment is not as a limitation of the invention.
The present invention provides a kind of platelet fibrin rich in somatomedin and the preparation method of release liquid thereof.Due in the preparation method that disclosed herein, rich in somatomedin platelet fibrin and release liquid prepare raw material, it is gather from the autologous blood of mammal, therefore with this platelet fibrin rich in somatomedin and release liquid thereof or its goods, when this mammal is carried out the various course for the treatment of, the exclusive problem of possible immunizing antigen antibody will not be caused rich in the platelet fibrin of somatomedin and release liquid thereof because of this.
The preparation method that disclosed herein, is first to provide the blood that mammal is autologous, is placed among container.Such as in the way of blood drawing, from mammiferous vein, directly gather the blood of about 10cc in container.Wherein, container can be blood taking tube, or the centrifuge tube of glass or plastics, and the latter is more conducive to follow-up centrifugation step and is made directly operation.
Then, by blood slight wobble, it is preferred to stop before presenting homogeneous condensation state to blood.The purpose of this step, is in order to the platelet composition in activating blood so that it is disengage somatomedin.Again owing to platelet composition is activated, therefore blood has the phenomenon of condensation.
Illustrating, container is elongated blood taking tube or centrifuge tube, it is advantageous to for camouflage color first-bornization blood taking tube, for instance be made by BD company, model is the blood taking tube of 367988.It addition, container before placing blood, can first preset coagulant and gel, wherein coagulant can accelerate blood coagulation, and gel then can assist blood after follow-up centrifuging process, separates the part of serum and blood cell grumeleuse.Container has two ends and interlude.When being placed in when blood among blood taking tube or centrifuge tube, by the two ends of the container fulcrum relative to interlude, repeat upper and lower straight line and back and forth or draw the motion of circle, i.e. the purpose of slight wobble blood in attainable cost step.As shown in Figures 1 A and 1 B 1, centrifuge tube 210 is equipped with blood 250, one fulcrum 220 of definition centrifuge tube 210, and the two ends of centrifuge tube 210 will according to shown in the direction of motion 230 or 240, in the way of fulcrum 220 is motionless, repeat upper and lower straight line respectively and back and forth or draw round motion, make blood 250 can flow up and down within centrifuge tube 210.Or, it is possible to by centrifuge tube 210 horizontal positioned, then whole centrifuge tube 210 of moving in the way of the figure of eight, it is also possible to reach the purpose making blood 250 can flow up and down within centrifuge tube 210.Provided above go out the mode of slight wobble centrifuge tube, be used as illustrating the use of the present invention to be not limited to the scope of the present invention.One skilled in the art, after understanding the spirit of the present invention, should various embodiments implement according to this.
Further illustrating, the time of slight wobble blood 250 can be 45 to 90 seconds, it is advantageous to is 60 seconds；And if defining is once that then repeating upper and lower straight line back and forth or draws round motion up and down back and forth, or the number of times with figure of eight moving, it is possible to be 30 to 60 times.
Then, the blood presenting coagulated state is centrifuged after static.Wherein, the centrifugal time is 9 to 12 minutes, it is advantageous to be 10 minutes；And centrifugal rotating speed is 2800 to 3200rpm(revolutionperminute, rotating speed per minute), it is advantageous to for 3000rpm.
The blood stood afterwards by centrifugation, will be rendered obvious by out the distribution of three layers material in a reservoir.Its at the middle and upper levels and lower floor be all liquid, upper strata is the platelet poor plasma (Platelet-PoorPlasma, PPP) of yellow, lower floor then presents redness, Main Ingredients and Appearance is erythrocyte, and middle level is then condensed into a linen cake mass, and containing abundant platelet composition.That is, in general venous blood, hematoblastic content concn is every microlitre (microliter) 15 ten thousand to 35 ten thousand；And above-mentioned canescence cake mass, then can further hematoblastic content concn be promoted to 1.5 times to 20 times of venous blood originally.
Then, linen cake mass is taken out, for instance utilize aseptic tweezers to be pressed from both sides out by cake mass.And cake mass is stood, treat that it discharges liquid.Being preferably and be placed in a test tube by this cake mass, and be placed in a warm water by this test tube, wherein the temperature of warm water is preferably 37 degree Celsius.The time stood can be 2 to 5 hours, it is advantageous to is 3 hours.As in figure 2 it is shown, when the time stood is more of a specified duration, somatomedin concentration in liquid is also just big, but after the time of repose more than 3 hours, concentration is namely gradually full closes.Therefore, considering time cost and extract the choice between concentration, time of repose is to be preferred with 2.5 ~ 3.5 hours.
Further illustrate, in the liquid that cake mass discharges, the composition of various somatomedin will be comprised, including platelet derived growth factor (platelet-derivedgrowthfactor, PDGF), make the transition somatomedin (transforminggrowthfactor, TGF), VEGF (vascularendothelialgrowthfactor, VEGF), epidermal growth factor (epithelialgrowthfactor, EGF), insulin like growth factor (insulin-likegrowthfactor, IGF), fiber mother cell growth factor (Fibroblastgrowthfactor, and nerve growth factor (Nervegrowthfactor FGF), NGF).Above-mentioned somatomedin, at signaling, hyperplasia, cell differentiation, collagen protein synthesis and angiogenesis etc., all can play the effect of auxiliary.
Finally, the liquid that the material that platelet condenses with fiber discharges is separated taking-up, for instance utilizing aseptic injection tube to extract liquid, this liquid is rich in multiple somatomedin.
Further illustrate, the platelet fibrin rich in somatomedin prepared by the present invention and release liquid thereof or with it for the medicine manufactured by raw material, preferably administering mode is topical, including local or mucosal administration, or it is administered to wound location, including operating wound, it is such as by using a kind of fluid, spraying, gel, Emulsion or ointment etc..Administering mode can also be any other easily mode include Formulations for systemic administration, such as with the form of injection, by subcutaneous or intra-articular injection, it is injected into skin layer, skin corium, the muscle of fat pad of various soft tissue, spongy bone or bone marrow, or it is sprayed onto tissue surface, mix with body fluid etc..
Various known drug-supplying systems (drugdeliverysystem), including syringe, syringe needle, conduit, bag etc., all can use.Concrete drug-supplying system has medical proof fabric or implant, and its Chinese medicine adhesive plaster can be used in the application of surface topical.Preferably drug-supplying system is the preparation of slow releasing, for instance the gel of hydrogel, semisolid or solid-state, to provide long-acting treatment.
The platelet fibrin rich in somatomedin prepared by the present invention and release liquid thereof or with it for the medicine manufactured by raw material, can be individually dosed, or combine or be combined in arbitrary another kind of reagent or any other therapeutic agent of use in indication, such as it is used for treating osteoarthritis, osteoarthritis, osteonecrosis, bone or ischemic patient.The platelet fibrin rich in somatomedin prepared by the present invention and release liquid thereof or with it for the medicine manufactured by raw material, it is possible to before the using of other reagent, while or use afterwards.
Additionally, alleged by this specification pharmaceutically acceptable " carrier ", refer to the platelet fibrin rich in somatomedin prepared by any and all suitable, present invention and the solvent compatible in physical characteristic of release liquid, dispersant, coating (coating), antibacterial and antifungal, etc. blend absorption delay agent or its analog.Other example includes sterilized water, saline, phosphate buffered saline (PBS), glucose, glycerol, ethanol etc., and their combination in any.The platelet fibrin rich in somatomedin prepared by the present invention and release liquid thereof, it is possible to based on selected administering mode, and with one or more suitable being combined of above-mentioned carrier.Such carrier and administering mode are belonging to general knowledge at pharmaceutical field.Carrier also can comprise gel, hydrogel, gellant, controlled-release material or other materials as known in the art.
Fig. 3 is the flow chart of steps of the preparation method of the platelet fibrin rich in somatomedin that disclosed herein and release liquid thereof, is arranged aforesaid preparation method to form, the step including following:
As described in step S510, it is provided that the blood that mammal is autologous, it is placed in container.As it was previously stated, gather the blood that mammal is autologous, the exclusive problem of immunizing antigen antibody will not be caused.
As described in step S520, by blood slight wobble, it is preferred to stop before being homogeneous condensation state to blood.The embodiment of slight wobble, it is possible to reference to aforesaid related description.
As described in step S530, static after blood is centrifuged, make blood be separated into three layers material in a reservoir, it is liquid with lower floor at the middle and upper levels, and intermediate layer is then condensed into a linen cake mass, is this platelet fibrin rich in somatomedin.The embodiment being centrifuged, it is possible to reference to aforesaid related description.
As described in step S540, take out linen cake mass, and cake mass is stood, treat that it discharges liquid.Take out canescence cake mass and the embodiment of the mode of standing, it is possible to reference to aforesaid related description.
As described in step S550, separate and take out liquid, be the fibrinoplatelet release liquid rich in somatomedin.Separate the embodiment taken out, it is possible to reference to aforesaid related description.
Embodiment described above is only the preferred embodiment lifted for absolutely proving the present invention, and protection scope of the present invention is not limited to this.Equivalent replacement that those skilled in the art make on basis of the present invention or conversion, all within protection scope of the present invention.Protection scope of the present invention is as the criterion with claims.
Claims (17)
1. the preparation method of the platelet fibrin rich in somatomedin and release liquid thereof, it is characterised in that comprise the steps of
The blood that one mammal is autologous is provided, is placed in a container；
This blood is rocked；
This blood is carried out one centrifugal after static, make this blood be separated into three layers material in this embodiment, it is liquid with lower floor at the middle and upper levels, and intermediate layer is then condensed into a cake mass and is this platelet fibrin rich in somatomedin；
Take out this cake mass, and this cake mass is stood, treat that it discharges a liquid；And
Separate and take out this liquid, be this fibrinoplatelet release liquid rich in somatomedin.
2. the preparation method of the platelet fibrin rich in somatomedin as claimed in claim 1 and release liquid thereof, it is characterized in that, being shaped as of this container is elongated, and there is two ends and an interlude, and this blood is rocked, refer to the two ends of this container fulcrum relative to this interlude, repeat about one straight line and back and forth or draw the motion of circle.
3. the preparation method of the platelet fibrin rich in somatomedin as claimed in claim 2 and release liquid thereof, it is characterised in that the time rocking this blood is 45 to 90 seconds.
4. the preparation method of the platelet fibrin rich in somatomedin as claimed in claim 3 and release liquid thereof, it is characterised in that the time rocking this blood is 60 seconds.
5. the preparation method of the platelet fibrin rich in somatomedin as according to any one of Claims 1-4 and release liquid thereof, it is characterised in that back and forth or the number of times of the motion drawing circle is 30 to 60 times to repeat this upper and lower straight line.
6. the preparation method of the platelet fibrin rich in somatomedin as claimed in claim 1 and release liquid thereof, it is characterised in that this centrifugal time is 9 to 12 minutes.
7. the preparation method of the platelet fibrin rich in somatomedin as claimed in claim 6 and release liquid thereof, it is characterised in that this centrifugal time is 10 minutes.
8. the preparation method of the platelet fibrin rich in somatomedin as according to any one of claim 6 or 7 and release liquid thereof, it is characterised in that this centrifugal rotating speed is 2800 to 3200rpm.
9. the preparation method of the platelet fibrin rich in somatomedin as claimed in claim 8 and release liquid thereof, it is characterised in that this centrifugal rotating speed is 3000rpm.
10. the preparation method of the platelet fibrin rich in somatomedin as claimed in claim 1 and release liquid thereof, it is characterised in that take out this cake mass, is utilize aseptic tweezers to be pressed from both sides out by this cake mass.
11. the preparation method of the platelet fibrin rich in somatomedin as claimed in claim 1 and release liquid thereof, it is characterised in that this container is a serum separator tube.
12. the preparation method of the platelet fibrin rich in somatomedin as described in claim 1 or 11 and release liquid thereof, it is characterised in that preset a coagulant and a gel in this container.
13. the preparation method of the platelet fibrin rich in somatomedin as claimed in claim 1 and release liquid thereof, it is characterised in that this cake mass is stood, refers to and this cake mass is placed in a test tube, and this test tube is placed in a warm water.
14. the preparation method of the platelet fibrin rich in somatomedin as claimed in claim 13 and release liquid thereof, it is characterised in that the temperature of this warm water is 37 degree Celsius.
15. the preparation method of the platelet fibrin rich in somatomedin as described in claim 1,13 or 14 and release liquid thereof, it is characterised in that the time being undertaken this cake mass standing is 2.5 to 5 hours.
16. the preparation method of the platelet fibrin rich in somatomedin as claimed in claim 15 and release liquid thereof, it is characterised in that the time being undertaken this cake mass standing is 3 hours.
17. the preparation method of the platelet fibrin rich in somatomedin as claimed in claim 1 and release liquid thereof, it is characterized in that, multiple somatomedin should be comprised rich in the platelet fibrin of somatomedin and release liquid thereof, and this somatomedin comprised platelet derived growth factor, make the transition somatomedin, VEGF, epidermal growth factor, insulin like growth factor, fiber mother cell growth factor and nerve growth factor.
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| CN108324992A (en) * | 2017-01-19 | 2018-07-27 | 国玺干细胞应用技术股份有限公司 | Composition for cartilage repair and method for repairing articular cartilage |
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| CN108324992A (en) * | 2017-01-19 | 2018-07-27 | 国玺干细胞应用技术股份有限公司 | Composition for cartilage repair and method for repairing articular cartilage |
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Inventor after: Zhuang Mingxi Inventor after: Guo Zongfu Inventor after: Zhuang Jixuan Inventor after: Li Qian Inventor after: Lin Pocheng Inventor after: Chen Junhong Inventor after: Zhuang Linxiang Inventor before: Zhuang Mingxi Inventor before: Guo Zongfu Inventor before: Zhuang Jixuan Inventor before: Li Qian Inventor before: Ma Yuxin Inventor before: Yan Zhengan |
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Application publication date: 20160629 |