CN106148269A - A kind of mammalian cell culture and additive thereof - Google Patents
A kind of mammalian cell culture and additive thereof Download PDFInfo
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- CN106148269A CN106148269A CN201510166524.2A CN201510166524A CN106148269A CN 106148269 A CN106148269 A CN 106148269A CN 201510166524 A CN201510166524 A CN 201510166524A CN 106148269 A CN106148269 A CN 106148269A
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Abstract
The present invention relates to the culture medium that used during In vitro culture mammalian cell and for constituting the additive of this culture medium, described culture medium can make mammalian somatic cell Effective multiplication when cultivating the somatic cell of mammal in the case of not using serum.By blending pilose antler active polysaccharide in the medium, even if the somatic cell Effective multiplication of mammal can also be made in the case of culture medium is entirely free of serum.
Description
Technical field
The invention belongs to cell culture medium field, the culture medium that used during more particularly to In vitro culture mammalian cell and for constituting the additive of this culture medium.
Background technology
Mammalian cell is cell therapy and the basis of regenerative medicine area research.The cell that its described cell contains the mammiferous cell with many differentiation functions including people and these cells break up out.Advance and cultivate these cells so that it is fast breeding, become this area research and productive key.
After known mammalian cell is in vitro, it is generally incubated in the addition of the culture medium of serum of about 10% cultivation propagation.But as serum, typically from the autoblood (such as acquiring somatic blood samples of patients to be cultivated) of the cell cultivated, or other derived sera such as Ox blood serum.But, when using from autologous serum, it is necessary to use the blood of patient relatively largely, therefore increased the weight of the burden of patient.And when using the serum from other kinds, there is the problem that may be mixed into the infectious pathogens such as unknown virus or Protein virus.Owing to the composition of any serum is understood the most completely, source according to the serum used or the difference of commercially available prod batch, its contained composition is likely to difference, therefore when a large amount of use serum are cultivated, it is difficult to the cell quality making cultivation gained is consistent.Accordingly it is desirable to somatic cell can be made under not using serum profiles can to breed by fast and stable equally.
The serum-free basal medium known the most altogether at present has: the MEM (MEM) of Eagle culture medium etc., Dulbecco improvement Eagle culture medium (DMEM), MEM α (MEM-α), between leaf system Cell Basal Medium (MSCBM), Ham ' s F-12 and F-10 culture medium, DMEM/F12 culture medium, Williams culture medium E, RPMI-1640 culture medium, MCDB culture medium, 199 culture medium, Fisher culture medium, Iscove improvement Dulbecco culture medium (IMDM), McCoy improved culture medium etc., wherein RPMI-1640 culture medium due to the suitability strong, it is widely used in mammaliancellculture.But, owing to RPMI-1640 culture medium often can add serum when cultivating zooblast, and then easily cause cell contamination or pathogen infection.Simultaneously as RPMI-1640 culture medium does not have the specificity of cultivated cell category, so for for some special cells, causing the relative drop of ability of cell proliferation on the contrary.
Summary of the invention
The required problem solved of invention
The problem of the present invention is to provide mammalian cell culture and additive thereof, and described culture medium, when cultivating mammalian cell, can make the somatic cell proliferation of mammal in the case of suppressing as much as possible to add serum in the medium effectively.For the method solving problem
Present inventor has performed in-depth study, found that: by blending pilose antler active polysaccharide component in the medium, the somatic cell Effective multiplication of mammal can be made.Thus complete the present invention.That is, the present invention provides mammalian cell culture medium, and this culture medium contains pilose antler active polysaccharide component.The present invention also provides for the additive of mammalian cell culture medium, and this additive contains pilose antler active polysaccharide.
The effect of invention
The culture medium of the present invention and culture medium additive can make mammalian cell proliferation under conditions of not using serum effectively.The problem that therefore, it can solve to produce due to use serum.That is, to the burden of patient, the problem such as be mixed into of infectious pathogen.By the cultural method of the present invention, the usage amount of serum can be suppressed to cultivate mammalian cell, it is provided that the cultivation somatic cell of stabilizing quality.Therefore, obtained cultivation cell is that infectious pathogen is mixed into problem and is suppressed to the cultivation somatic cell of minimal stabilizing quality.
The best mode carried out an invention
In this specification, " mammalian cell " refers to the cell in mammalian cell in addition to proliferative cell, carry out specialization without becoming the noble cells of cell in addition including for certain purpose, and have and be divided into the cell with several difference in functionality cell abilities.The former cell is the functional cell of adult, the cell of each organ in formation organism such as including Skin Cell, neurocyte, myocyte, blood system cell, fibroblast, hepatocyte, chondrocyte, adipose cell.The latter is referred to as stem cell, is can to break up to be converted to the cell of the noble cells that at least one is above in above-mentioned differentiated cell, comprises embryonic stem cell and adult stem cell." adult stem cell " is to have in the stem cell of the ability being divided into the cell with several difference in functionality and precursor the cell in addition to embryonic stem cell, including induction versatile stem cell, hematopoietic stem cell, leaf system stem cell, neural stem cell, skin progenitor cell, liver stem cells, pancreatic stem cells etc..
As the cell cultivated in the culture medium of the present invention, as long as the somatic cell of mammal, there is no any restriction.Preferential example can the adult stem cell such as example T lymphocyte, bone marrow stem cell, but be not so limited.
Containing pilose antler active polysaccharide as essential component in the culture medium of the present invention.Pilose antler active polysaccharide is compared with other polysaccharide, in addition to the biological function with other active polysaccharide, also has unique biological activity, and sulphuric acid Cornu Cervi Pantotrichum chrondroitin is one of its main component, and it is possibly together with somatomedin simultaneously.They can promote osteogenesis, wound healing and treatment tumor.Additionally, experimental studies have found that by us, pilose antler active polysaccharide can promote cell proliferation, and effect is notable.
The Method and way of pilose antler active polysaccharide component is the most relatively simple.Mainly through Cornu Cervi Pantotrichum water logging, cell breakage, colloid mill grindings, thawing, add after enzyme hydrolysis, repeatedly sedimentation etc. operate, the pilose antler polysaccharides of purification can be obtained.
Accompanying drawing explanation
Fig. 1 is the change (concentration of pilose antler active polysaccharide is 0.01mM) of the population doublings quantity representing mononuclearcell in human blood
Fig. 2 is the change (concentration of pilose antler active polysaccharide is 0.1mM) of the population doublings quantity representing mononuclearcell in human blood
Fig. 3 is the change (concentration of pilose antler active polysaccharide is 5mM) of the population doublings quantity representing mononuclearcell in human blood
Fig. 4 is the change (concentration of pilose antler active polysaccharide is 10mM) of the population doublings quantity representing mononuclearcell in human blood
Fig. 5 is the change (concentration of pilose antler active polysaccharide is 100mM) of the population doublings quantity representing mononuclearcell in human blood
Fig. 6 is the change (concentration of pilose antler active polysaccharide is 0.01mM) of the population doublings quantity representing human fat tissue derived stem cells
Fig. 7 is the change (concentration of pilose antler active polysaccharide is 0.1mM) of the population doublings quantity representing human fat tissue derived stem cells
Fig. 8 is the change (concentration of pilose antler active polysaccharide is 5mM) of the population doublings quantity representing human fat tissue derived stem cells
Fig. 9 is the change (concentration of pilose antler active polysaccharide is 10mM) of the population doublings quantity representing human fat tissue derived stem cells
Figure 10 is the change (concentration of pilose antler active polysaccharide is 100mM) of the population doublings quantity representing human fat tissue derived stem cells
Figure 11 is the change (concentration of pilose antler active polysaccharide is 0.01mM) of the population doublings quantity representing people's bone marrow stem cell
Figure 12 is the change (concentration of pilose antler active polysaccharide is 0.1mM) of the population doublings quantity representing people's bone marrow stem cell
Figure 13 is the change (concentration of pilose antler active polysaccharide is 5mM) of the population doublings quantity representing people's bone marrow stem cell
Figure 14 is the change (concentration of pilose antler active polysaccharide is 10mM) of the population doublings quantity representing people's bone marrow stem cell
Figure 15 is the change (concentration of pilose antler active polysaccharide is 100mM) of the population doublings quantity representing people's bone marrow stem cell
Detailed description of the invention
For making the present invention easier to understand, below in conjunction with instantiation, the present invention is expanded on further.Should be understood that these examples are merely to illustrate the present invention rather than limit the scope of the present invention.
It should be noted that in following Examples, as do not carried out specified otherwise, the component of following term and culture medium is as described below:
Basal medium: for the cultivation product containing cell growth desired nutritional materials such as saccharide, aminoacid, inorganic salt, vitamin, lipids of artificial preparation.Basal medium of the present invention is DMEM in high glucose basal medium.
Serum substitute: replace the culture additive of serum for class I goodsization, composition typically determines.Serum substitute used by the present invention is N2 (a kind of business-like blood serum substituting additive, composition is insulin, transferrins, progesterone, putrescine, sodium selenite).
Pilose antler active polysaccharide: for the additive of basal culture medium, the mainly polysaccharose substance of extraction purification from Cornu Cervi Pantotrichum stem cell.Composition is complex.Mainly contain lentinan, Cordyceps polysaccharide, sulphuric acid Cornu Cervi Pantotrichum chrondroitin and some somatomedin etc..Wherein, conventional experimentation shows, sulphuric acid Cornu Cervi Pantotrichum chrondroitin is the main composition of pilose antler active polysaccharide, has and promotes bone mass cells growth, wound healing and effect for the treatment of tumor.It did not occur for the constituent report of cell culture medium.
The concentration (containing time multiple, for its total concentration) of the pilose antler polysaccharides composition in culture medium is typically about 0.01-100mM, further preferred 0.1-10mM.Wherein, the sulphuric acid Cornu Cervi Pantotrichum chrondroitin preferred 0.1-100mM of concentration, further preferred 0.25-20mM in the medium.Lentinan concentration in the medium is preferably 0.25-10mM, further preferred 0.5-5mM.Cordyceps polysaccharide concentration in the medium is preferably 0.25-10mM, further preferred 0.5-5mM.
The culture medium of the present invention further preferably contains antioxidant.The preferred embodiment of antioxidant is Cys.Known antioxidants has programmed cell death inhibitory action, therefore for cultivating the holding of cell, propagation effectively.Antioxidant can be used alone, it is also possible to is used in combination of two or more.Antioxidant concentration (containing time multiple, for its total concentration) in culture medium preferably 0.01mM-10mM, further preferred 0.1mM-1mM.
The culture medium of the present invention preferably further contains somatomedin.By containing somatomedin, cell proliferation can be promoted further.The preferred embodiment of somatomedin is transforming growth factor (TGF), granulocyte colony-stimulating factor (G-CSF) and epidermal growth factor (EGF).These somatomedin bases are all known in this field.Somatomedin can be used alone and can also be used in combination of two or more.Somatomedin concentration (containing time multiple, for its total concentration) in culture medium preferably 0.1-100ng/mL, further preferred 1-10ng/mL.
The culture medium of the present invention can contain surfactant further.It is believed that by the surfactant containing low concentration, there is the dysgenic effect reduced cell membrane.If known, the surfactant of high concentration is added in culture medium, then suppression cell proliferation or inducing cell death.The preferred embodiment of surfactant is alkyl phenoxy Polyethylene Glycol.The concentration (containing time multiple, for its total concentration) of surfactant usually 0.1-100ng/mL, preferably 1-10ng/mL.
The culture medium of the present invention can contain serum in the same manner as conventional mammalian cell culture medium, but by containing serum substitute and pilose antler polysaccharides composition, even the culture medium not containing serum can also make cell high-efficient breed.Therefore, when culture medium is serum-free medium, the effect of the present invention can be played the most well.That is, relative to culture medium total amount, the serum content in the culture medium of the present invention preferably 0%.
The culture medium of the present invention except containing pilose antler polysaccharides composition, further preferably containing above-mentioned antioxidant, serum substitute, somatomedin and surfactant more than one outside, it is also possible to as known mammalian cell culture medium.Therefore basically by adding above two essential component, further preferably more than one above-mentioned preferred compositions in known basal medium, it is possible to obtain the culture medium of the present invention.
The culture medium of the present invention can be containing the known various additives contained in mammalian cell culture medium.Above-mentioned known additive can enumerate: other additive of amino acids, inorganic salts, vitamins and carbon source or antibiotic etc..
As amino acids, can enumerate: glycine, ALANINE, L-arginine, altheine, L-Aspartic acid, Cys, CYSTINE, Pidolidone, L-glutaminate, L-Histidine, ILE, L-Leu, 1B, METHIONINE, L-phenylalanine, L-PROLINE, Serine, L-threonine, L-Trp, TYR and Valine.
As inorganic salts, can enumerate: calcium chloride, copper sulfate, ferric nitrate (III), iron sulfate, magnesium chloride, magnesium sulfate, potassium chloride, sodium bicarbonate, sodium chloride, disodium hydrogen phosphate, sodium dihydrogen phosphate, zinc sulfate and selenic acid.
As vitamins, can enumerate: choline, vitamin A, vitamin B1, vitamin B2, vitamin B3, adenine phosphate, vitamin B5, vitamin B6, vitamin B7, vitamin B12, orotic acid, VITAMIN B15, laetrile, vitamin B h, vitamin B t, PABA, vitamin C, vitamin D, vitamin E, vitaminF, vitamin K, vitamin(e) M and Citrin.
Being added to by these additives in mammalian cell culture medium, this itself is known, and the addition of each additive can also be with known culture medium same way, it is also possible to suitably set by normal experiment.It is about 5mg/L-500mg/L that the addition of such as amino acids is typically every kind of aminoacid, preferably about 10mg/L-400mg/L, it is about 0mg/L-10g/L that the addition of inorganic salts is typically every kind of inorganic salts, preferably about 0.01mg/L-7g/L, the addition of vitamin be every kind of vitamin be about 0.01mg/L-500mg/L, preferably about 0.05mg/L-300mg/L.
As other additive, can enumerate: (1) corticosterone, the somatomedin of progesterone etc., (2) penicillin, streptomycin, the antibiotic such as gentamycin and kanamycin, (3) glucose, galactose, the carbon source such as fructose and sucrose, (4) magnesium, ferrum, zinc, calcium, potassium, sodium, copper, selenium, cobalt, stannum, molybdenum, the minor metallic element such as nickel and silicon and adenosine 5 ' phosphoric acid, corticosterone, ethanolamine, insulin, reduced glutathion, thioctic acid, hypoxanthine, phenol red, progesterone, putrescine, acetone acid, breast glycosides, 3, transferrin, other additive such as lactoferrin.The addition of these additives is the most same, it is also possible to suitably set by normal experiment according to the purpose of each additive.Usually 0.001mg/L-5g/L, specifically for about 0.1-3g/L.
The culture medium of the present invention, in addition to being used for for the cultivation of somatic propagation or holding, when the cell cultivated is stem cell, may be used for the induction of stem cell.When the induction of stem cell, differentiating inducer can be added further.As differentiating inducer, can enumerate: cytokine (various interleukin, erythropoietin, thrombopoietin etc.), colony stimulating factor (such as G-CSF etc.), steroid hormone (dexamethasone etc.), indole (indomethacin), tetrahydrothiazole derivates (rosiglitazone etc.) etc..They one or more can be used.The addition of differentiating inducer is the most same, and the total amount being commonly angled relative to culture medium can add about 1 × 10-10 to 1 × 10-6 weight %.
The culture medium of the present invention can be containing one or more of above-mentioned various additives.Generally can combine containing multiple additives.
Mammalian somatic cell cultivation in the culture medium of the present invention can be carried out according to method same itself, generally at temperature and the 5%CO of 30-37 DEG C2In the environment of and 5-21%O2In the environment of carry out.
The present invention also provides for the additive of the culture medium for constituting the invention described above.Therefore, the additive of the present invention contains pilose antler polysaccharides.Further preferably containing above-mentioned preferred various compositions.Still further preferably contain one or more of above-mentioned various additive.For the additive of the present invention, following additive is easy, thus preferably, and described additive has can form the composition of the invention described above culture medium in water or basal medium by being dissolved in.In this case, the ratio of the content of the mixing proportion composition each with culture medium of contained in additive various compositions is identical.It should be noted that, based on culture medium, the above-mentioned culture medium in the past used in the cultivation of mammalian cell can be enumerated.
Embodiment 1: the cultivation of mononuclearcell (PBMC) in human peripheral
(1-1) preparation of pilose antler active polysaccharide
The pilose antler active polysaccharide of 2mg is dissolved in 1mL PBS, prepares the many sugar additives of pilose antler active (A).
(1-2) propagation of mononuclearcell (PBMC) in human peripheral
Containing amino acids (above-mentioned whole amino acidses), inorganic salts (above-mentioned whole inorganic salts), vitamins (above-mentioned whole vitaminss) and other additives (adenosine 5 ' phosphoric acid, corticosterone, ethanolamine, D-galactose, D-Glucose, insulin, reduced glutathion, thioctic acid, hypoxanthine, phenol red, progesterone, putrescine, acetone acid, breast glycosides, 3, transferrin, sodium selenite, Cys, transforming growth factor, granulocyte colony-stimulating factor, epidermal growth factor) culture medium in add 7 × 10-4% 2 mercapto ethanol (2-Me), 100U/mL penicillin streptomycin (prepared by PBS), obtain serum-free medium (B1).
(1-3) in serum-free medium, add A prepared by (1-1) with various concentration, obtain serum-free medium (C1).
Specifically add A according to following concentration.
The serum-free medium of 0.01mM A is added in culture medium B；
The serum-free medium of 0.1mM A is added in culture medium B；
The serum-free medium of 5mM A is added in culture medium B；
The serum-free medium of 10mM A is added in culture medium B；
The serum-free medium of 100mM A is added in culture medium B；
7 × 10 are added in RPMI-1640 culture medium-4%2-mercaptoethanol (2-ME), 100U/L penicillin streptomycin (prepared by PBS), the somatomedin of 2.5ng/mL, obtain conventional serum-free medium (conventional serum-free medium).
These culture medium of 10mL are added separately in the culture dish of diameter 100mm, inoculate hMADs, at 37 DEG C, 5%CO with 10000 cells of initial cell number/mL2Environment in cultivate.
The cell quantity of every day is counted, the 12 days inner cell population doublings quantity (10 certainly started from cultivation by cell countern) change.Population doublings quantity is the cell total amount observed every day.
Result is expressed as follows:
Fig. 1 is the change (concentration of pilose antler active polysaccharide is 0.01mM) of the population doublings quantity representing mononuclearcell in human blood
Fig. 2 is the change (concentration of pilose antler active polysaccharide is 0.1mM) of the population doublings quantity representing mononuclearcell in human blood
Fig. 3 is the change (concentration of pilose antler active polysaccharide is 5mM) of the population doublings quantity representing mononuclearcell in human blood
Fig. 4 is the change (concentration of pilose antler active polysaccharide is 10mM) of the population doublings quantity representing mononuclearcell in human blood
Fig. 5 is the change (concentration of pilose antler active polysaccharide is 100mM) of the population doublings quantity representing mononuclearcell in human blood
Embodiment 2: from the cultivation of human fat tissue derived stem cells (ADMSCs)
(2-1) preparation of pilose antler active polysaccharide
The pilose antler active polysaccharide of 2mg is dissolved in 1mL PBS, prepares the many sugar additives of pilose antler active (A).
(2-2) propagation of human fat tissue derived stem cells (ADMSCs)
The basal medium of preparation removes from embodiment 1 hypoxanthine and thymidine and a part of inorganic salts (copper sulfate, ferric nitrate (III), iron sulfate, magnesium chloride, disodium hydrogen phosphate, zinc sulfate), in the basal medium of gained, add 100U/mL penicillin streptomycin (PBS preparation), obtain serum-free medium (B2).
(2-3) in serum-free medium, add A prepared by (2-1) with various concentration, obtain serum-free medium (C2).
Specifically add A according to following concentration.
The serum-free medium of 0.01mM A is added in culture medium B2；
The serum-free medium of 0.1mM A is added in culture medium B2；
The serum-free medium of 5mM A is added in culture medium B2；
The serum-free medium of 10mM A is added in culture medium B2；
The serum-free medium of 100mM A is added in culture medium B2；
7 × 10 are added in RPMI-1640 culture medium-4%2-mercaptoethanol (2-ME), 100U/L penicillin streptomycin (prepared by PBS), the somatomedin of 2.5ng/mL, obtain conventional serum-free medium (conventional serum-free medium).
These culture medium of 10mL are added separately in the culture dish of diameter 100mm, inoculate ADMSCs, at 37 DEG C, 5%CO with 10000 cells of initial cell number/mL2Environment in cultivate.
By cell counter counting every day cell quantity, from cultivate from start 12 days inner cell population doublings quantity (10n) change.Population doublings quantity is the cell total amount observed every day.
Result is expressed as follows:
Fig. 6 is the change (concentration of pilose antler active polysaccharide is 0.01mM) of the population doublings quantity representing human fat tissue derived stem cells
Fig. 7 is the change (concentration of pilose antler active polysaccharide is 0.1mM) of the population doublings quantity representing human fat tissue derived stem cells
Fig. 8 is the change (concentration of pilose antler active polysaccharide is 5mM) of the population doublings quantity representing human fat tissue derived stem cells
Fig. 9 is the change (concentration of pilose antler active polysaccharide is 10mM) of the population doublings quantity representing human fat tissue derived stem cells
Figure 10 is the change (concentration of pilose antler active polysaccharide is 100mM) of the population doublings quantity representing human fat tissue derived stem cells
Embodiment 3: from the cultivation of people's bone marrow stem cell (MSC)
(3-1) preparation of pilose antler active polysaccharide
The pilose antler active polysaccharide of 2mg is dissolved in 1mL PBS, prepares the many sugar additives of pilose antler active (A).
(3-2) propagation of people's bone marrow stem cell (MSC)
The basal medium of preparation removes from embodiment 1 transforming growth factor, granulocyte colony-stimulating factor, epidermal growth factor and 3 and a part of inorganic salts (copper sulfate, ferric nitrate (III), iron sulfate, magnesium chloride, disodium hydrogen phosphate, zinc sulfate), in the basal medium of gained, add 100U/mL penicillin streptomycin (PBS preparation), obtain serum-free medium (B3).
(3-3) in serum-free medium, add A prepared by (2-1) with various concentration, obtain serum-free medium (C3).
Specifically add A according to following concentration.
The serum-free medium of 0.01mM A is added in culture medium B3；
The serum-free medium of 0.1mM A is added in culture medium B3；
The serum-free medium of 5mM A is added in culture medium B3；
The serum-free medium of 10mM A is added in culture medium B3；
The serum-free medium of 100mM A is added in culture medium B3；
7 × 10 are added in RPMI-1640 culture medium-4%2-mercaptoethanol (2-ME), 100U/L penicillin streptomycin (prepared by PBS), the somatomedin of 2.5ng/mL, obtain conventional serum-free medium (conventional serum-free medium).
These culture medium of 10mL are added separately in the culture dish of diameter 100mm, inoculate ADMSCs, at 37 DEG C, 5%CO with 10000 cells of initial cell number/mL2Environment in cultivate.
By cell counter counting every day cell quantity, from cultivate from start 12 days inner cell population doublings quantity (10n) change.Population doublings quantity is the cell total amount observed every day.
Result is expressed as follows:
Figure 11 is the change (concentration of pilose antler active polysaccharide is 0.01mM) of the population doublings quantity representing people's bone marrow stem cell
Figure 12 is the change (concentration of pilose antler active polysaccharide is 0.1mM) of the population doublings quantity representing people's bone marrow stem cell
Figure 13 is the change (concentration of pilose antler active polysaccharide is 5mM) of the population doublings quantity representing people's bone marrow stem cell
Figure 14 is the change (concentration of pilose antler active polysaccharide is 10mM) of the population doublings quantity representing people's bone marrow stem cell
Figure 15 is the change (concentration of pilose antler active polysaccharide is 100mM) of the population doublings quantity representing people's bone marrow stem cell.
Claims (15)
1. mammalian cell culture, this culture medium contains pilose antler active polysaccharide.
2. the culture medium described in claim 1, this culture medium is serum-free medium.
3. the serum-free medium described in claim 2, this culture medium contains serum substitute.
4. the serum-free medium according to any one of Claims 2 or 3, wherein, serum substitute includes KSR (a kind of business
The band serum free culture system additive of industry), N2 (a kind of business-like blood serum substituting additive, composition be insulin, transferrins,
Progesterone, putrescine, sodium selenite) and B27 (a kind of serum-free culture additive for cultured nerve cell) etc. for serum free culture system
Additive, but it is several to be not limited to this.
5. the serum substitute according to any one of claim 3 or 4, this culture medium uses N2 (a kind of business-like serum
Substituting additive, composition is insulin, transferrins, progesterone, putrescine, sodium selenite) as serum substitute.
6. the culture medium described in claim 1 or 2, wherein, pilose antler active polysaccharide includes: lentinan, Cordyceps polysaccharide and sulfur
Acid Cornu Cervi Pantotrichum chrondroitin.
7. the culture medium according to any one of claim 1-6, this culture medium contains antioxidant further.
8. the culture medium described in claim 7, wherein, above-mentioned antioxidant is Cys.
9. the culture medium described in claim 8, this culture medium contains somatomedin further.
10. profit requires the culture medium described in 9, and wherein, above-mentioned somatomedin is transforming growth factor (TGF), granular leukocyte colony thorn
Swash the factor (G-CSF) and epidermal growth factor (EGF).
The additive of 11. mammalian cell cultures, this additive is pilose antler active polysaccharide.
Additive described in 12. claim 11, final concentration of in cell culture medium of this pilose antler active polysaccharide
0.01mM-10mM。
Additive according to any one of 13. claim 11-12, this additive contains antioxidant further.
Additive according to any one of 14. claim 11-13, this additive contains somatomedin further.
Additive according to any one of 15. claim 11-14, this additive has by being dissolved in water or basis cultivation
Base and form the composition of the culture medium according to any one of claim 1-10.
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107034189A (en) * | 2017-05-31 | 2017-08-11 | 东莞市保莱生物科技有限公司 | A kind of candidate stem cell cultural method |
| CN109486749A (en) * | 2017-09-11 | 2019-03-19 | 广西索芙特集团有限公司 | Velvet deerhorn cell Serium-free Culture |
| CN110499282A (en) * | 2019-09-03 | 2019-11-26 | 广州赛莱拉干细胞科技股份有限公司 | A kind of culture medium and its application and induction method of the tendon stem cell to Adipocyte Differentiation |
| CN113583949A (en) * | 2021-07-28 | 2021-11-02 | 新疆西部赛澳生物科技有限责任公司 | Cell culture medium and application thereof |
-
2015
- 2015-04-10 CN CN201510166524.2A patent/CN106148269A/en active Pending
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107034189A (en) * | 2017-05-31 | 2017-08-11 | 东莞市保莱生物科技有限公司 | A kind of candidate stem cell cultural method |
| CN109486749A (en) * | 2017-09-11 | 2019-03-19 | 广西索芙特集团有限公司 | Velvet deerhorn cell Serium-free Culture |
| CN110499282A (en) * | 2019-09-03 | 2019-11-26 | 广州赛莱拉干细胞科技股份有限公司 | A kind of culture medium and its application and induction method of the tendon stem cell to Adipocyte Differentiation |
| CN113583949A (en) * | 2021-07-28 | 2021-11-02 | 新疆西部赛澳生物科技有限责任公司 | Cell culture medium and application thereof |
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