CN105219707B - A kind of method of recovery fat mesenchymal stem cell - Google Patents
A kind of method of recovery fat mesenchymal stem cell Download PDFInfo
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- CN105219707B CN105219707B CN201510800480.4A CN201510800480A CN105219707B CN 105219707 B CN105219707 B CN 105219707B CN 201510800480 A CN201510800480 A CN 201510800480A CN 105219707 B CN105219707 B CN 105219707B
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Abstract
The present invention relates to field of biotechnology, disclose a kind of method of recovery fat mesenchymal stem cell.After this method will be passaged to the fat mesenchymal stem cell complete medium culture in P3-P5 generation, the DMEM/F12 culture medium culture of serum-free is used again, respectively culture for 24 hours, 48h, 72h when collect culture medium supernatant, will each supernatant centrifugation after take supernatant to mix again, obtain conditioned medium；The fat mesenchymal stem cell frozen is thawed, and is added in conditioned medium centrifugation and removes supernatant, conditioned medium is then added again and is resuspended and cultivates to cell fusion degree reaches requirement.The present invention was to cultivate culture medium of the culture medium supernatant of fat mesenchymal stem cell as recovery when, the mescenchymal stem cell complete medium generallyd use in substitution existing method, the growth of fat mesenchymal stem cell after promoting recovery by the active material that various kinds of cell wherein included is secreted in breeding, and then improve its vigor.
Description
Technical field
The present invention relates to field of biotechnology, a kind of method for particularly relating to recovery fat mesenchymal stem cell.
Background technique
Mescenchymal stem cell (mesenchymalstemcells, MSC) is the important member of stem cell line, from hair
The mesoderm and ectoderm for educating early stage belong to multipotential stem cell, and MSC initially has found in marrow, because it is latent with Multidirectional Differentiation
Can, hematopoiesis support and it promote stem cell implantation, immunoregulation and the concern that people are increasingly subject to the features such as self-replacation.As between
Mesenchymal stem cells under specific inductive condition, can be divided into fat, bone, cartilage, muscle, tendon, ligament, mind in vivo or in vitro
Through Various Tissues cells such as, liver, cardiac muscle, endotheliums, still there is multi-lineage potential after continuous passage culture and freezen protective, can make
It is ideal seed cell for injuries of tissues and organs reparation caused by aging and lesion.
Fat mesenchymal stem cell is derived from the mescenchymal stem cell of adult adipose, dry thin with umbilical cord, medulla mesenchyma
Born of the same parents are the same, with stronger self-renewing and multi-lineage potential.Mescenchymal stem cell needs after being separately cultured by low
Warm freezen protective carries out injuries of tissues and organs reparation using recovery when in use.
Document " being separately cultured and freeze the variation of front and back biology of human adipose mesenchymal stem cells " (Chinese neuroimmunology
With neurology magazine in July, 2,013, the 4th phase of volume 20) describe the universal side of current recovery fat mesenchymal stem cell
Method:
It is removed from liquid nitrogen after cell is immediately placed in 38 DEG C of -40 DEG C of water-baths thawing rapidly and is transferred in advance after 6 months
5min is centrifuged with 300g in centrifuge tube equipped with culture solution, cell is resuspended with ADSCs complete medium and is transferred in culture bottle often
Rule culture, ADSCs complete medium are the low sugar DMEM containing 10% volume fraction fetal calf serum.
Although the method for above-mentioned recovery is fairly simple, quick, the cell viability after this method recovery is relatively low, is unfavorable for
The clinical applications such as subsequent progress injuries of tissues and organs reparation.
Summary of the invention
In view of this, the purpose of the present invention is to provide a kind of methods of recovery fat mesenchymal stem cell, so that described
Method can be improved the vigor of fat mesenchymal stem cell after recovery, and still maintains and can be divided into osteoblast, at rouge
The ability of cell.
To achieve the goals above, the invention provides the following technical scheme:
A kind of method of recovery fat mesenchymal stem cell, comprising:
The fat mesenchymal stem cell for being passaged to P3-P5 generation is first used mescenchymal stem cell complete medium culture by step 1
Afterwards, then with the DMEM/F12 culture medium culture of serum-free, respectively culture for 24 hours, 48h, 72h when collect culture medium supernatant, will be each
It takes supernatant to mix again after the centrifugation of period supernatant, obtains conditioned medium；
Step 2, by the fat mesenchymal stem cell frozen thaw, and be added to step 1 collection conditioned medium in from
The heart removes supernatant, and conditioned medium is then added again and is resuspended and cultivates to cell fusion degree and reach requirement.
The not high problem of cell viability after the fat mesenchymal stem cell frozen for existing method recovery, the present invention is multiple
It does not use commercial medium usual in conventional method (mescenchymal stem cell complete medium) to recover when Soviet Union, but uses
The conditioned medium of fat mesenchymal stem cell was cultivated to recover, can significantly improve after recovery that cell viability is not high shows
As.Present invention discover that the vigor of recovery cell or tissue can be effectively improved using conditioned medium, and the condition of separate sources
Culture solution has different promotion or inhibiting effect to the vigor of different cells, tissue.
Wherein, preferably, step 1 are as follows:
The fat mesenchymal stem cell in P3-P5 generation will be passaged to according to 5000-10000/cm2Cell density mesenchyma
The culture of stem cell complete medium for 24 hours, after being cleaned with Hanks, then with the DMEM/F12 culture medium culture of serum-free, is being trained respectively
Support for 24 hours, 48h, 72h when collect culture medium supernatant, 1000-2000rpm is centrifuged 5-10min, and it is mixed to collect all supernatants for removal precipitating
It closes, obtains conditioned medium after membrane filtration.
For parameter, incubation time and the ring of cell-seeding-density, centrifugally operated in conditioned medium of the present invention
Border etc. can be adjusted according to the actual situation, and the present invention is provided than more preferably parameter, it is not limited to these parameters
And combinations thereof, for the similar parameters occurred in other methods step of the present invention be also not limited to provided by preferred parameter and its
Combination.
Preferably, the mescenchymal stem cell complete medium is the DMEM/F12 culture medium containing 10%FBS.
The fat mesenchymal stem cell in P3-P5 generation can be come according to the separation method and propagating method of this field routine
It obtains, the present invention provides following preferred preparation methods:
By the adult adipose of extraction 0.1%-0.5%I Collagenase Type enzymolysis, digestion, centrifuged deposit is dry thin with mesenchyma
Born of the same parents' complete medium is resuspended and cultivates, and changes liquid within every 2-3 days, reaches 80%-90% to cell fusion degree, obtains P0 generation；
With pancreatin digestion P0 for cell, precipitating and secondary culture is resuspended with mescenchymal stem cell complete medium after centrifugation,
It is passaged to P3-P5 generation in the manner described.
Still more preferably, by the adult adipose of extraction, after being cleaned with PBS, adipose tissue is shredded, and 0.1%- is added
0.5%I Collagenase Type, at 37 DEG C, digestion enzymatic hydrolysis 30-50 minutes under the conditions of 100-200rpm turn centrifugation 5-10 with 1000-2000
Minute, supernatant is removed, mescenchymal stem cell complete medium is added, cell precipitation is resuspended, be subsequently placed at 37 DEG C, 5%CO2Cell
Culture culture, changes liquid in every 2-3 days, reaches 80%-90% to cell fusion degree, obtains P0 generation；
With 0.25% pancreatin digestion P0 for cell, precipitating, 5000- is resuspended with mescenchymal stem cell complete medium after centrifugation
10000/cm2Continue to cultivate in cell density inoculated and cultured ware, is passaged to P3-P5 generation in the manner described.
In step 2 recovery link, by conditioned medium of the present invention substitute it is existing through frequently with business culture
Base is recovered according to general way.
Preferably, step 2 are as follows:
The fat mesenchymal stem cell frozen is placed in 35-42 DEG C of water-bath, concussion incubation 30-60 seconds, then
It is added in the conditioned medium of 5-10 times of volume, with 1000-2000rpm centrifugation 3-5 minutes, removes supernatant, use conditioned medium
Precipitating is resuspended, piping and druming mixes, and according to 5000-10000/cm2Cell density be placed on 37 DEG C, 5%CO with conditioned medium2
After incubator stationary culture 24 hours, replacement conditioned medium continues culture to cell fusion degree and reaches 80%-90%.
The fat mesenchymal stem cell frozen with the method for the invention recovery, compared to using existing method (" people's fat
Mescenchymal stem cell be separately cultured and freeze front and back biology variation " in record method for resuscitation) recovery after cell, live
Power significantly improves, and still keeps the good biological property of mescenchymal stem cell, has and is divided into osteoblast and thin at rouge
The ability of born of the same parents.Based on this, the present invention provides application of the conditioned medium in recovery fat mesenchymal stem cell.This hair
The bright conditioned medium can be placed in -20 DEG C of incubators when not in use and save -12 months 6 months or so or be placed on -80
It DEG C saves 1 year or more.It needs to use in use, being restored to 37 DEG C of room temperature.
From the above technical scheme, when the present invention was to cultivate the culture medium supernatant of fat mesenchymal stem cell as recovery
Culture medium, the mescenchymal stem cell complete medium that generallys use in existing method is substituted, by wherein included a variety of thin
The active material that born of the same parents secrete in breeding promotes the growth of the fat mesenchymal stem cell after recovery, and then improves its work
Power.
Detailed description of the invention
Fig. 1 show vigor column diagram after method for resuscitation of the present invention and existing method for resuscitation cell recovery；
Fig. 2 show method for resuscitation of the present invention and existing method for resuscitation cell recovery and cultivates the total number of cells column after 72h
Shape figure；
Fig. 3 show blank group cell surface antigen flow cytomery figure；
Fig. 4 show experimental group cell surface antigen flow cytomery figure；
Fig. 5 show the microscopy figure of cell osteogenic induction after recovery；
Fig. 6 show the microscopy figure of cell adipogenic induction after recovery.
Specific embodiment
The embodiment of the invention discloses a kind of methods of recovery fat mesenchymal stem cell.Those skilled in the art can borrow
Reflect present disclosure, is suitably modified realization of process parameters.In particular, it should be pointed out that all similar substitutions and modifications are to this field
It is it will be apparent that they are considered as being included in the present invention for technical staff.Method of the invention passes through preferable reality
Example is applied to be described, related personnel obviously can not depart from the content of present invention, in spirit and scope to product as described herein
And method is modified or appropriate changes and combinations, carrys out implementation and application the technology of the present invention.
It is dry to a kind of recovery fat mesenchymal provided by the invention below with reference to embodiment for a further understanding of the present invention
The method of cell is described in detail.
Embodiment 1: the culture of fat mesenchymal stem cell and the collection of conditioned medium
By the adult adipose of extraction, after being cleaned with PBS, adipose tissue is shredded, and 0.25I Collagenase Type is added, at 37 DEG C,
Digestion enzymatic hydrolysis 50 minutes under the conditions of 200rpm, turn centrifugation 5-10 minutes with 1000-2000, remove supernatant, complete medium is added
(DMEM/F12+10%FBS) cell precipitation is resuspended, is subsequently placed at 37 DEG C, 5%CO2Cell culture culture changes liquid in every 2-3 days,
Reach 80%-90% to cell fusion degree, with 0.25% trypsin digestion cell, is resuspended and is precipitated with complete medium after centrifugation, (5-
10)×103Continue to cultivate in/cm2 cell density inoculated and cultured ware.P3-P5 is taken to carry out conditioned medium collection for cell.
Conditioned medium is collected: for the fat mesenchymal stem cell of conditioned medium, first according to (5-10) × 103/cm2
After being cleaned with Hanks, the DMEM/ of serum-free is added after complete medium carries out culture 24 hours in cell density inoculated and cultured ware
F12 culture medium, after 24 hours, 48 hours, 72 hours, respectively collect cell culture medium supernatant, by supernatant be sub-packed in 50mL from
In heart pipe, 1000-2000rpm is centrifuged 5-10min.Removal precipitating, collects supernatant.CMC model is drawn with disposable syringe
Base is collected in sterile collection bottle with 0.22 μm of membrane filtration.- 20 DEG C of incubators are placed on to save -12 months 6 months or so
Or it is placed on -80 DEG C of preservations 1 year or more.It needs in use, being restored to room temperature use.
Embodiment 2: the recovery of fat mesenchymal stem cell
The fat mesenchymal stem cell frozen is removed from liquid nitrogen, is placed in 35-42 DEG C of water-bath, concussion incubates
60 seconds, after cell thaws, frozen stock solution is added to in the conditioned medium of 5 times of volumes cell suspension immediately with 1000rpm centrifugation 5
Minute, supernatant is removed, is resuspended and is precipitated with conditioned medium；Gently piping and druming mixes, and according to 7 × 103/cm2Cell density inoculated and cultured
In ware, with conditioned medium culture.37 DEG C are placed on, 5%CO2After incubator stands 24 hours, replacement conditioned medium continues
Culture, daily to observe cell amplification situation, cell carries out continuing to cultivate to cell fusion degree reaching 80%-90%.
Embodiment 3: recovery cell viability comparison
Experimental group: conditioned medium is prepared according to embodiment 1, by the fat mesenchymal stem cell frozen from liquid nitrogen
It takes out, is placed in 35-42 DEG C of water-bath, concussion incubates 60 seconds, and after cell thaws, frozen stock solution is added to 5 times of bodies immediately
In long-pending conditioned medium, a small amount of cell suspension is taken, trypan blue is added, cell viability calculating instrument is placed on and carries out cell viability inspection
It surveys, remaining cell suspension removes supernatant with 1000rpm centrifugation 5 minutes, is resuspended and is precipitated with conditioned medium；Gently piping and druming mixes, and
According to 7 × 103/cm2Cell density is inoculated in six orifice plates, with conditioned medium culture.37 DEG C are placed on, 5%CO2Incubator training
It supports 72 hours, digests centrifuge cell with pancreatin, cell is resuspended with conditioned medium, takes a small amount of cell suspension, trypan blue is added, puts
It sets and carries out cell viability detection in cell viability calculating instrument.
Control group: it is (dry for the fat mesenchymal of identical source with experimental group to be removed from liquid nitrogen fat mesenchymal stem cell
Cell), it puts into 38 DEG C of -40 DEG C of water-baths melt immediately.Be transferred to be pre-loaded with the culture solution of the DMEM/F12 containing 10%FBS from
In heart pipe, cell progress cell viability detection is taken out after mixing cell, 5min is centrifuged with 300g, removes supernatant, it is complete with hADSCs
Cell is resuspended in culture medium, according to 7 × 103/cm2Cell density is inoculated with six orifice plates, and is placed in 37 DEG C, 5%CO2It is trained in incubator
It supports 72 hours.Centrifuge cell is digested with pancreatin, cell is resuspended with the DMEM/F12 culture medium containing 10%FBS, takes a small amount of cell outstanding
Trypan blue is added in liquid, is placed on cell counter and carries out cell count.
The two comparing result is shown in Fig. 1 and Fig. 2, by Fig. 1, it is apparent that between fat after recovering via the method for the present invention
The vigor of mesenchymal stem cells is 92% or so, and the cell viability after existing method recovery is 85% or so, and the two is compared to having
Significant difference.Meanwhile as seen from Figure 2, in cell recovery and after the culture of 72h, by the method for the present invention culture
Total number of cells show that the cell viability after method for resuscitation of the present invention is recovered is higher, are proliferated energy considerably beyond existing method
Power is strong.
Embodiment 4: the fat mesenchymal stem cell surface marker after flow cytomery recovery
It recovers according to embodiment 1,2 and cultivates cell, cell fusion degree reaches 80%-90% after recovering, with 0.25% pancreas
Enzymic digestion centrifuge cell, and every pipe is added 2 × 10 after counting5Cell number, dye solution are washed 1 time, and 1000rpm is centrifuged 5min；
Supernatant is abandoned, is blown and beaten with dye solution and mixes cell；Each 2 μ L of CD73, CD90, CD45 and HLA-DRA antibody is added, and sets one
Pipe is blank control；At 4 DEG C, it is protected from light 15-20min；Dye solution is washed once, and 1000rpm is centrifuged 5min；Directly mark
The cell of note abandons supernatant, is protected from light the sample-loading buffer that 500 μ L are added, and mixes, with 200 mesh screen filtration cell samples, streaming is thin
Born of the same parents' instrument detects cell surface antigen, as a result sees Fig. 3 and Fig. 4.
As can be seen from figs. 3 and 4 mescenchymal stem cell feminine gender surface marker CD45 (leucocyte is positive), HLA-DR (MHC-II
Class molecule) it is feminine gender to be presented, while the positive is presented in mescenchymal stem cell surface marker CD73, CD90.After illustrating recovery
Fat mesenchymal stem cell still keeps the good biological feature of mescenchymal stem cell.
Embodiment 5: the mescenchymal stem cell induced osteogenesis differentiation identification for recovering adipose-derived
It recovers according to embodiment 1,2 and cultivates cell, cell fusion degree reaches 80%-90% after recovering, with 0.25% pancreas
Enzymic digestion centrifuge cell, according to 1 × 103Cell/cm2Be inoculated in six orifice plates, be added complete medium, for 24 hours after, be added skeletonization
After induced medium is cultivated for 24 hours, liquid was changed every 2-3 days, Alizarin red staining is carried out after 21 days, as a result sees Fig. 5.
Mescenchymal stem cell after recovery is illustrated to pass through after osteogenic induction 3 weeks by the visible calcium scoring of Alizarin red staining
The fat mesenchymal stem cell for crossing conditioned medium recovery still maintains potential to Osteoblast Differentiation.
Embodiment 6: the mescenchymal stem cell induction for recovering adipose-derived is broken up at rouge to be identified
It recovers according to embodiment 1,2 and cultivates cell, cell confluency degree reaches 80%-90% after recovering, with 0.25% pancreas
Enzymic digestion centrifuge cell, after collecting cell, and according to 1 × 103Cell/cm2It is inoculated in six orifice plates, complete medium is added,
After for 24 hours, adipogenic induction culture medium is added and is cultivated.Adipogenic induction culture medium includes basal medium DMEM, 10% tire ox blood
Clearly, 0.5mM IBMX, 1 μM of dexamethasone, 100 μM of Indomethacins, 5 μ g/mL insulin, 2mm/L glutamine etc..It changes every three days
Liquid.Oil red O stain is carried out after four weeks, is identified fat drips formational situation, is as a result seen Fig. 6.
Fat mesenchymal stem cell after recovery, by oil red O stain red color visible oil droplet, is said after adipogenic induction 4 weeks
The bright fat mesenchymal stem cell by conditioned medium recovery is still maintained to the potential broken up at rouge.
The above description of the embodiment is only used to help understand the method for the present invention and its core ideas.It should be pointed out that pair
For those skilled in the art, without departing from the principle of the present invention, the present invention can also be carried out
Some improvements and modifications, these improvements and modifications also fall within the scope of protection of the claims of the present invention.
Claims (4)
1. a kind of method of recovery fat mesenchymal stem cell characterized by comprising
Step 1 will be passaged to the fat mesenchymal stem cell in P3-P5 generation according to 5000-10000/cm2Cell density use contain 10%
The DMEM/F12 culture medium culture of FBS for 24 hours, after being cleaned with Hanks, then with the DMEM/F12 culture medium culture of serum-free, respectively
Culture for 24 hours, 48h, 72h when collect culture medium supernatant, 1000-2000rpm be centrifuged 5-10min, removal precipitating, collect it is all on
It is clear to mix, conditioned medium is obtained after membrane filtration；
The fat mesenchymal stem cell frozen is placed in 35-42 DEG C of water-bath by step 2, concussion incubation 30-60 seconds, so
It is added in the conditioned medium of 5-10 times of volume afterwards, with 1000-2000rpm centrifugation 3-5 minutes, removes supernatant, use CMC model
The outstanding precipitating of base weight, piping and druming mix, and according to 5000-10000/cm2Cell density be placed on 37 DEG C, 5% with conditioned medium
CO2 After incubator stationary culture 24 hours, replacement conditioned medium continues culture to cell fusion degree and reaches 80%-90%.
2. method according to claim 1, which is characterized in that the P3-P5 fat subsitutes mescenchymal stem cell passes through with lower section
Method obtains:
By the adult adipose of extraction 0.1%-0.5% collagenase type I enzymolysis, digestion, centrifuged deposit mescenchymal stem cell is complete
Full culture medium is resuspended and cultivates, and changes liquid within every 2-3 days, reaches 80%-90% to cell fusion degree, obtains P0 generation；
With pancreatin digestion P0 for cell, precipitating and secondary culture, passage is resuspended with mescenchymal stem cell complete medium after centrifugation
To P3-P5 generation.
3. method according to claim 2, which is characterized in that the P3-P5 fat subsitutes mescenchymal stem cell passes through with lower section
Method obtains:
By the adult adipose of extraction, after being cleaned with PBS, adipose tissue is shredded, and 0.1%-0.5% collagenase type I is added, 37
DEG C, digestion enzymatic hydrolysis 30-50 minutes under the conditions of 100-200rpm turn centrifugation 5-10 minutes with 1000-2000, remove supernatant, are added
Cell precipitation is resuspended in mescenchymal stem cell complete medium, is subsequently placed at 37 DEG C, 5% CO2Cell culture culture, every 2-3 days
Liquid is changed, reaches 80%-90% to cell fusion degree, obtains P0 generation；
With 0.25% pancreatin digestion P0 for cell, precipitating, 5000- is resuspended with mescenchymal stem cell complete medium after centrifugation
10000/cm2 Continue to cultivate in cell density inoculated and cultured ware, is passaged to P3-P5 generation in the manner described.
4. application of the conditioned medium described in claim 1 in recovery fat mesenchymal stem cell.
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| CN105749347A (en) * | 2016-02-24 | 2016-07-13 | 广州赛莱拉干细胞科技股份有限公司 | Cosmetic filler and preparation method thereof |
| CN108373995B (en) * | 2018-02-14 | 2020-11-17 | 浙江生创精准医疗科技有限公司 | Stem cell conditioned medium, preparation method and application thereof |
| CN109161526A (en) * | 2018-10-12 | 2019-01-08 | 希瑞干细胞科技有限公司 | A kind of chorion mescenchymal stem cell recovery medium |
| CN110791477A (en) * | 2019-11-21 | 2020-02-14 | 陕西九州细胞基因工程有限公司 | Culture method of mesenchymal stem cells after cryopreservation and recovery of adipocytes |
| CN111304164A (en) * | 2019-11-30 | 2020-06-19 | 杭州拜欧津生物科技有限公司 | Preparation method and application of adipose-derived stem cell exosomes |
| CN111849880B (en) * | 2020-06-23 | 2022-04-29 | 和携科技有限公司 | Recovery method of human adipose mesenchymal stem cells after ultralow-temperature cryopreservation |
| CN112155008A (en) * | 2020-09-30 | 2021-01-01 | 北京银丰鼎诚生物工程技术有限公司 | Cryopreservation and recovery method of adipose tissues and adipose tissue cryopreservation recovery kit |
| CN115305232B (en) * | 2022-02-22 | 2024-02-20 | 王泰华 | Adipose-derived mesenchymal stem cell resuscitating culture solution and resuscitating method |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102676452A (en) * | 2012-04-26 | 2012-09-19 | 天津美德太平洋科技有限公司 | Culture medium containing human umbilical cord mesenchymal stem cell exudates and preparation method and applications thereof |
| CN103784474A (en) * | 2014-01-26 | 2014-05-14 | 广州赛莱拉生物科技有限公司 | Human fat mesenchymal stem cell extract and lyophilized powder and application thereof |
| CN104762260A (en) * | 2015-04-23 | 2015-07-08 | 广州赛莱拉干细胞科技股份有限公司 | Preparation method and application of adipose-derived mesenchymal stem cells and preparation thereof |
| KR20150100260A (en) * | 2014-02-25 | 2015-09-02 | 주식회사 미래셀바이오 | Non-antibiotics serum-free mesenchymal stem cells medium, method for preparing the same, and cosmetic composition comprising the same |
-
2015
- 2015-11-17 CN CN201510800480.4A patent/CN105219707B/en active Active
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102676452A (en) * | 2012-04-26 | 2012-09-19 | 天津美德太平洋科技有限公司 | Culture medium containing human umbilical cord mesenchymal stem cell exudates and preparation method and applications thereof |
| CN103784474A (en) * | 2014-01-26 | 2014-05-14 | 广州赛莱拉生物科技有限公司 | Human fat mesenchymal stem cell extract and lyophilized powder and application thereof |
| KR20150100260A (en) * | 2014-02-25 | 2015-09-02 | 주식회사 미래셀바이오 | Non-antibiotics serum-free mesenchymal stem cells medium, method for preparing the same, and cosmetic composition comprising the same |
| CN104762260A (en) * | 2015-04-23 | 2015-07-08 | 广州赛莱拉干细胞科技股份有限公司 | Preparation method and application of adipose-derived mesenchymal stem cells and preparation thereof |
Non-Patent Citations (3)
| Title |
|---|
| 人脂肪间充质干细胞的分离培养及冻存前后生物学变化;谢甬淋等;《中国神经免疫学和神经病学杂志》;20130731;第20卷(第4期);第263-266页 * |
| 低温冻存对人脂肪间充质干细胞生物学特性的影响;李兴天等;《江苏大学学报（医学版）》;20140531;第24卷(第3期);第190-194页 * |
| 雌激素影响冻存肾脂肪囊来源脂肪间充质干细胞的成脂分化;张慧等;《中国组织工程研究》;20130702;第17卷(第27期);第4998-5004页 * |
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