CN109362708A - A kind of stem cell refrigerant and its application - Google Patents
A kind of stem cell refrigerant and its application Download PDFInfo
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- CN109362708A CN109362708A CN201811364395.8A CN201811364395A CN109362708A CN 109362708 A CN109362708 A CN 109362708A CN 201811364395 A CN201811364395 A CN 201811364395A CN 109362708 A CN109362708 A CN 109362708A
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N1/00—Preservation of bodies of humans or animals, or parts thereof
- A01N1/02—Preservation of living parts
- A01N1/0205—Chemical aspects
- A01N1/021—Preservation or perfusion media, liquids, solids or gases used in the preservation of cells, tissue, organs or bodily fluids
- A01N1/0221—Freeze-process protecting agents, i.e. substances protecting cells from effects of the physical process, e.g. cryoprotectants, osmolarity regulators like oncotic agents
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Abstract
The present invention provides a kind of stem cell refrigerant, the stem cell refrigerant includes following components in percentage by weight: the human serum albumin of 1%-10%, the Sodium Hyaluronate of 2%-20%, 5%-30% half lift-off sugar and 1%-5% dimethyl sulfoxide.Human serum albumin, Sodium Hyaluronate, half lift-off sugar in stem cell refrigerant of the invention have good biocompatibility, toxicity is relatively low when high concentration, utilize the human serum albumin of the proportion, Sodium Hyaluronate, half lift-off sugar, it can inhibit the formation of intracellular ice crystal to the maximum extent, reduce damage of the low temperature environment to institute's freeze-stored cell, the thus significantly lower toxicity of freeze-stored cell, and improve survival rate of the freeze-stored cell after recovery culture.
Description
Technical field
The present invention relates to field of biomedicine more particularly to a kind of stem cell refrigerant and its applications.
Background technique
Cell freezing preservation is that cell culture is suspended in the solution containing cryoprotector, is dropped with certain rate
To subzero, and the process of long-term preservation at low temperature.Cell freezing, which saves, is generally stored in -80 DEG C or liquid nitrogen -196
DEG C low temperature environment in.In refrigerating process, outer crystallization intracellular can occur for cell, while the solution concentration of intraor extracellular also can
Since corresponding change occurs for crystallization, the cell exposed to the open air at low temperature can also generate different pressure therewith.
So far, two technologies are applied in the freezen protective application of stem cell, one is traditional slow freezing,
It is another then be glass frozen preservation.In both methods, dimethyl sulfoxide (DMSO) is the cryoprotector generallyd use.Visitor
For sight, it is dry thin that DMSO is widely used in mouse embryo stem cell, mankind hemopoietic stem cell, human mesenchymal stem cell, people's fat
In the freezen protective of born of the same parents, and good refrigerating effect is achieved in these cells, however, slightly and more serious adverse reaction
It is all related with defrosting cellular products of the infusion containing DMSO.In addition, its metabolite of the bad smell of DMSO influence patient and
Working environment.
Summary of the invention
There is provided it is an object of the invention to overcome in place of the shortcomings of the prior art a kind of stem cell refrigerant and its
Using.
To achieve the above object, the technical scheme adopted by the invention is as follows: a kind of stem cell refrigerant, stem cell freezing
Agent includes following components in percentage by weight: the human serum albumin of 1%-10%, the Sodium Hyaluronate of 2%-20%, 5%-30%
Half lift-off sugar (Pentaisomaltose) and 1%-5% dimethyl sulfoxide (DMSO).
Human serum albumin, Sodium Hyaluronate, half lift-off sugar in stem cell refrigerant of the invention have good biology
Compatibility, toxicity is relatively low when high concentration, can be most using the human serum albumin of the proportion, Sodium Hyaluronate, half lift-off sugar
Inhibit to limits the formation of intracellular ice crystal, reduce damage of the low temperature environment to institute's freeze-stored cell, thus significantly lower freezes
The toxicity of cell is deposited, and improves survival rate of the freeze-stored cell after recovery culture.The present invention is matched by the simple combination
Than the maximum protection advantage that can play human serum albumin, Sodium Hyaluronate, half lift-off sugar, while in frozen cell program
Have no increase.
Preferably, the range of the molecular weight of the Sodium Hyaluronate is 3.5 × 104-5×104。
Preferably, the DMSO accounts for the 1-3% of the stem cell refrigerant weight.
It is highly preferred that the DMSO accounts for the 1.8% of the stem cell refrigerant weight.
Preferably, the human serum albumin accounts for 2%, 4%, 6% or the 8% of the stem cell refrigerant weight.
Preferably, the Sodium Hyaluronate accounts for 5%, 8%, 12% or the 15% of the stem cell refrigerant weight.
Preferably, the half lift-off sugar accounts for 10%, 15%, 20% or the 25% of the stem cell refrigerant weight.
Preferably, the stem cell refrigerant further includes icariine (Icariin), and the icariine is described dry thin
Content in born of the same parents' refrigerant is 5-100 μM.
More electedly, content of the icariine in the stem cell refrigerant is 15-30 μM.
Preferably, the stem cell refrigerant further includes heparin.
Preferably, the stem cell refrigerant further includes acid-base modifier, and the soda acid regulator is 4- hydroxyethyl piperazine
Ethanesulfonic acid (HEPES) or disodium ethylene diamine tetraacetate (EDTA).
Preferably, the stem cell refrigerant further includes the antioxidant for accounting for the stem cell refrigerant weight 1%-8%.
It is highly preferred that the antioxidant is vitamin C, vitamin E or lecithin.
Stem cell refrigerant of the invention cell freezing and during thaw can effectively stabilizing cell membrane and protein,
And a kind of glassy matrices can be formed in refrigerating process, inhibit the formation of potential fatal intracellular ice crystal with this, by
This reduces cellular damage, and stem cell refrigerant of the invention can effectively protect protein molecule, Ke Yiqing at low ambient temperatures
Except the active oxygen generated due to low temperature, at the same after low temperature stress help freeze-stored cell recovery.
The present invention provides a kind of preferred stem cell refrigerants, and the stem cell refrigerant includes following components,
It accounts for the human serum albumin of the stem cell refrigerant weight 2%-5%, account for the stem cell refrigerant weight 5%-
15% Sodium Hyaluronate, half lift-off for accounting for the stem cell refrigerant weight 10%-20% sugared (Pentaisomaltose),
It accounts for the dimethyl sulfoxide (DMSO) of the stem cell refrigerant weight 1.8%, account for the stem cell refrigerant weight 1%-8%'s
Antioxidant, 20 μM of icariine, acid-base modifier and 1-2.5IE/ml heparin.
The present invention also provides a kind of any one of the above stem cell refrigerants in mescenchymal stem cell or candidate stem cell
In application.
The beneficial effects of the present invention are: the present invention provides a kind of stem cell refrigerant and its application, of the invention is dry
Human serum albumin, Sodium Hyaluronate, half lift-off sugar in cell freezing agent have a good biocompatibility, toxicity when high concentration
It is relatively low, using the human serum albumin of the proportion, Sodium Hyaluronate, half lift-off sugar, it can inhibit to the maximum extent intracellular
The formation of ice crystal, reduces damage of the low temperature environment to institute's freeze-stored cell, thus the significantly lower toxicity of freeze-stored cell, and improves
Survival rate of the freeze-stored cell after recovery culture.The present invention can play the white egg of people's blood by the simple combination matching
The maximum protection advantage of white, Sodium Hyaluronate, half lift-off sugar, while increase is had no in frozen cell program；Of the invention is dry
Cell freezing agent cell freezing and during thaw can effectively stabilizing cell membrane and protein, and can be in refrigerating process
A kind of glassy matrices are formed, inhibits the formation of potential fatal intracellular ice crystal with this, thus reduces cellular damage, this
The stem cell refrigerant of invention can effectively protect protein molecule at low ambient temperatures, can remove the activity generated due to low temperature
Oxygen, at the same after low temperature stress help freeze-stored cell recovery.
Detailed description of the invention
Fig. 1 is the cell viability testing result figure that the stem cell refrigerant of the embodiment of the present invention is applied.
Fig. 2 is the apoptosis rate testing result figure that the stem cell refrigerant of the embodiment of the present invention is applied.
Fig. 3 is the cell viability testing result figure that the stem cell refrigerant of the embodiment of the present invention is applied.
Fig. 4 is the apoptosis rate testing result figure that the stem cell refrigerant of the embodiment of the present invention is applied.
Fig. 5 is the cell viability testing result figure that the stem cell refrigerant of the embodiment of the present invention is applied.
Fig. 6 is the apoptosis rate testing result figure that the stem cell refrigerant of the embodiment of the present invention is applied.
Specific embodiment
To better illustrate the object, technical solutions and advantages of the present invention, below in conjunction with specific embodiment to the present invention
It is described further.
Embodiment 1
A kind of stem cell refrigerant as the embodiment of the present invention comprising following components,
It accounts for the human serum albumin of the stem cell refrigerant weight 4%, account for the transparent of the stem cell refrigerant weight 8%
Matter acid sodium, accounts for the stem cell freezing at half lift-off sugared (Pentaisomaltose) for accounting for the stem cell refrigerant weight 15%
The dimethyl sulfoxide (DMSO) of agent weight 1.8%, the antioxidant for accounting for the stem cell refrigerant weight 5%, 20 μM of excessive sheep
The heparin of leaves of pulse plants glucoside, EDTA and 1.8IE/ml, the range of the molecular weight of the Sodium Hyaluronate are 3.5 × 104-5×104。
Embodiment 2
A kind of stem cell refrigerant as the embodiment of the present invention comprising following components,
It accounts for the human serum albumin of the stem cell refrigerant weight 4%, account for the transparent of the stem cell refrigerant weight 8%
Matter acid sodium, accounts for the stem cell freezing at half lift-off sugared (Pentaisomaltose) for accounting for the stem cell refrigerant weight 15%
The dimethyl sulfoxide (DMSO) of agent weight 1% accounts for 5% antioxidant of the stem cell refrigerant weight, 20 μM of Herba Epimedii
The heparin of glucoside, EDTA and 1.8IE/ml, the range of the molecular weight of the Sodium Hyaluronate are 3.5 × 104-5×104。
Embodiment 3
A kind of stem cell refrigerant as the embodiment of the present invention comprising following components,
It accounts for the human serum albumin of the stem cell refrigerant weight 4%, account for the transparent of the stem cell refrigerant weight 8%
Matter acid sodium, accounts for the stem cell freezing at half lift-off sugared (Pentaisomaltose) for accounting for the stem cell refrigerant weight 15%
The dimethyl sulfoxide (DMSO) of agent weight 2.5%, the antioxidant for accounting for the stem cell refrigerant weight 5%, 20 μM of excessive sheep
The heparin of leaves of pulse plants glucoside, EDTA and 1.8IE/ml, the range of the molecular weight of the Sodium Hyaluronate are 3.5 × 104-5×104。
Embodiment 4
A kind of stem cell refrigerant as the embodiment of the present invention comprising following components,
It accounts for the human serum albumin of the stem cell refrigerant weight 4%, account for the transparent of the stem cell refrigerant weight 8%
Matter acid sodium, accounts for the stem cell freezing at half lift-off sugared (Pentaisomaltose) for accounting for the stem cell refrigerant weight 15%
The dimethyl sulfoxide (DMSO) of agent weight 4% accounts for 5% antioxidant of the stem cell refrigerant weight, 20 μM of Herba Epimedii
The heparin of glucoside, EDTA and 1.8IE/ml, the range of the molecular weight of the Sodium Hyaluronate are 3.5 × 104-5×104。
Experimental example 1
The effect of the refrigerant application of embodiment 1-4 is detected, method is as follows.
Cell culture: the Eagle culture medium that mescenchymal stem cell is improved in Dulbecco, DMEM are supplemented with 10% tire ox blood
It grows, cell line is placed in conical flask, 37 in (FBS) clearly, 1% penicillin/streptomycin and 12.5 μ g/mL puromycins
DEG C, it is kept in 5%CO2/95% air atmosphere, and passage in every 2-3 days is primary.
Cell freezing and defrosting: the stem cell that the cell for passing on 3 times is resuspended in the 1mL embodiment 1-4 in cryovial is cold
Freeze in agent, cell concentration 2 × 106A/mL, the freezen protective in the refrigerator of speed control, freezing procedure: 4 DEG C of initial temperature, -1
DEG C/min to 0 DEG C, -2 DEG C/min to -45 DEG C and -5 DEG C/min to -100 DEG C, cryovial is transferred into liquid nitrogen (- 190 DEG C) simultaneously
Storage is until use.Freezen protective thaws sample after 2 weeks, 3 weeks before them, and by cryovial, quick-thawing is straight at 37 DEG C
Ice is not observed into solution.Fresh culture is slowly added to inhibit infiltration to damage for stem cell freezing agent solution removal
Wound, until removing any remaining CPA solution.
Cell viability detection: by the cell after above-mentioned defrosting with culture medium rinse 3 times, using the cell of non-frozen preservation as
Control group is seeded in 96 orifice plates and cultivates 24 hours.Then, CCK-8 determines that cell viability passes through, and detects and inhales at 450nm
Luminosity.
Apoptosis rate measurement: Annexin V-FITC/PI apoptosis detection kit, experimental group cell and control group is thin
Born of the same parents are resuspended in 500 μ L combination buffers, and 5 μ L annexin V-FITC and PI are added, then, by sample room temperature in the dark
It is lower to incubate 15 minutes, pass through flow analysis apoptosis rate cell count.
Cell viability testing result is as shown in Figure 1, as shown in Figure 1, in other refrigerant components containing embodiment 1-4
Under situation, the content of the dimethyl sub-maple in the refrigerant from 1% increase to 4% when, vigor after cell thaws after freezing
Increase, illustrate that refrigerating effect improves, but the content of dimethyl sub-maple from 1.8% increase to 4% when, the degree of increase is slow,
In view of the toxicity of dimethyl sub-maple, embodiment 1 is preferable.And the cell viability of freezing after three weeks cell viability of two weeks compared with freezing
It is smaller.
Apoptosis rate testing result is as shown in Fig. 2, as shown in Figure 2, in other refrigerant components containing embodiment 1-4
In the case of, the content of the dimethyl sub-maple in the refrigerant from 1% increase to 4% when, it is thin after cell thaws after freezing
Born of the same parents' apoptosis rate reduce, illustrate that refrigerating effect improves, but the content of dimethyl sub-maple from 1.8% increase to 4% when, Apoptosis
The degree that rate reduces is slow, it is contemplated that the toxicity of dimethyl sub-maple, embodiment 1 are preferable.And freezing cell viability after three weeks compared with
The apoptosis rate of freezing two weeks is larger.
Embodiment 5
A kind of stem cell refrigerant as the embodiment of the present invention comprising following components,
It accounts for the human serum albumin of the stem cell refrigerant weight 4%, account for the transparent of the stem cell refrigerant weight 8%
Matter acid sodium, accounts for the stem cell freezing at half lift-off sugared (Pentaisomaltose) for accounting for the stem cell refrigerant weight 5%
The dimethyl sulfoxide (DMSO) of agent weight 1.8%, the antioxidant for accounting for the stem cell refrigerant weight 5%, 20 μM of excessive sheep
The heparin of leaves of pulse plants glucoside, EDTA and 1.8IE/ml, the range of the molecular weight of the Sodium Hyaluronate are 3.5 × 104-5×104。
Embodiment 6
A kind of stem cell refrigerant as the embodiment of the present invention comprising following components,
It accounts for the human serum albumin of the stem cell refrigerant weight 4%, account for the transparent of the stem cell refrigerant weight 8%
Matter acid sodium, accounts for the stem cell freezing at half lift-off sugared (Pentaisomaltose) for accounting for the stem cell refrigerant weight 10%
The dimethyl sulfoxide (DMSO) of agent weight 1.8%, the antioxidant for accounting for the stem cell refrigerant weight 5%, 20 μM of excessive sheep
The heparin of leaves of pulse plants glucoside, EDTA and 1.8IE/ml, the range of the molecular weight of the Sodium Hyaluronate are 3.5 × 104-5×104。
Embodiment 7
A kind of stem cell refrigerant as the embodiment of the present invention comprising following components,
It accounts for the human serum albumin of the stem cell refrigerant weight 4%, account for the transparent of the stem cell refrigerant weight 8%
Matter acid sodium, accounts for the stem cell freezing at half lift-off sugared (Pentaisomaltose) for accounting for the stem cell refrigerant weight 20%
The dimethyl sulfoxide (DMSO) of agent weight 1.8%, the antioxidant for accounting for the stem cell refrigerant weight 5%, 20 μM of excessive sheep
The heparin of leaves of pulse plants glucoside, EDTA and 1.8IE/ml, the range of the molecular weight of the Sodium Hyaluronate are 3.5 × 104-5×104。
Embodiment 8
A kind of stem cell refrigerant as the embodiment of the present invention comprising following components,
It accounts for the human serum albumin of the stem cell refrigerant weight 4%, account for the transparent of the stem cell refrigerant weight 8%
Matter acid sodium, accounts for the stem cell freezing at half lift-off sugared (Pentaisomaltose) for accounting for the stem cell refrigerant weight 25%
The dimethyl sulfoxide (DMSO) of agent weight 1.8%, the antioxidant for accounting for the stem cell refrigerant weight 5%, 20 μM of excessive sheep
The heparin of leaves of pulse plants glucoside, EDTA and 1.8IE/ml, the range of the molecular weight of the Sodium Hyaluronate are 3.5 × 104-5×104。
Embodiment 9
A kind of stem cell refrigerant as the embodiment of the present invention comprising following components,
It accounts for the human serum albumin of the stem cell refrigerant weight 4%, account for the transparent of the stem cell refrigerant weight 8%
Matter acid sodium, accounts for the stem cell freezing at half lift-off sugared (Pentaisomaltose) for accounting for the stem cell refrigerant weight 30%
The dimethyl sulfoxide (DMSO) of agent weight 1.8%, the antioxidant for accounting for the stem cell refrigerant weight 5%, 20 μM of excessive sheep
The heparin of leaves of pulse plants glucoside, EDTA and 1.8IE/ml, the range of the molecular weight of the Sodium Hyaluronate are 3.5 × 104-5×104。
Experimental example 2
The effect of the refrigerant application of embodiment 5-9 is detected, method is the same as experimental example 1.
Cell viability testing result is as shown in figure 3, from the figure 3, it may be seen that in other freezings containing embodiment 5-9, embodiment 1
In the case of agent component, the content of the half lift-off sugar in the refrigerant from 5% increase to 30% when, after cell thaws after freezing
Vigor increase, illustrate that refrigerating effect improves, but the content of half lift-off sugar be greater than 15% after, the degree of increase is slow, consideration
To cost, embodiment 1 is preferably.And the cell viability of freezing after three weeks cell viability of two weeks compared with freezing is smaller.
Apoptosis rate testing result as shown in figure 4, as shown in Figure 4, containing embodiment 5-9, embodiment 1 other are cold
Freeze in the case of agent component, the content of the half lift-off sugar in the refrigerant from 5% increase to 30% when, cell defrosting after freezing
After apoptosis rate afterwards reduces, and illustrates that refrigerating effect improves, but the content of half lift-off sugar is greater than 15%, apoptosis rate subtracts
Small degree is slow, it is contemplated that cost, embodiment 1 are preferable.And the cell viability of freezing after three weeks cell of two weeks compared with freezing withers
It is larger to die rate.
Embodiment 10
A kind of stem cell refrigerant as the embodiment of the present invention comprising following components,
It accounts for the human serum albumin of the stem cell refrigerant weight 4%, account for the transparent of the stem cell refrigerant weight 2%
Matter acid sodium, accounts for the stem cell freezing at half lift-off sugared (Pentaisomaltose) for accounting for the stem cell refrigerant weight 15%
The dimethyl sulfoxide (DMSO) of agent weight 1.8%, the antioxidant for accounting for the stem cell refrigerant weight 5%, 20 μM of excessive sheep
The heparin of leaves of pulse plants glucoside, EDTA and 1.8IE/ml, the range of the molecular weight of the Sodium Hyaluronate are 3.5 × 104-5×104。
Embodiment 11
A kind of stem cell refrigerant as the embodiment of the present invention comprising following components,
It accounts for the human serum albumin of the stem cell refrigerant weight 4%, account for the transparent of the stem cell refrigerant weight 5%
Matter acid sodium, accounts for the stem cell freezing at half lift-off sugared (Pentaisomaltose) for accounting for the stem cell refrigerant weight 15%
The dimethyl sulfoxide (DMSO) of agent weight 1.8%, the antioxidant for accounting for the stem cell refrigerant weight 5%, 20 μM of excessive sheep
The heparin of leaves of pulse plants glucoside, EDTA and 1.8IE/ml, the range of the molecular weight of the Sodium Hyaluronate are 3.5 × 104-5×104。
Embodiment 12
A kind of stem cell refrigerant as the embodiment of the present invention comprising following components,
It accounts for the human serum albumin of the stem cell refrigerant weight 4%, account for the saturating of the stem cell refrigerant weight 12%
Bright matter acid sodium, half lift-off for accounting for the stem cell refrigerant weight 15% sugared (Pentaisomaltose), to account for the stem cell cold
The dimethyl sulfoxide (DMSO) for freezing agent weight 1.8%, the antioxidant for accounting for the stem cell refrigerant weight 5%, 20 μM excessive
The heparin of sheep leaves of pulse plants glucoside, EDTA and 1.8IE/ml, the range of the molecular weight of the Sodium Hyaluronate are 3.5 × 104-5×104。
Embodiment 13
A kind of stem cell refrigerant as the embodiment of the present invention comprising following components,
It accounts for the human serum albumin of the stem cell refrigerant weight 4%, account for the saturating of the stem cell refrigerant weight 15%
Bright matter acid sodium, half lift-off for accounting for the stem cell refrigerant weight 15% sugared (Pentaisomaltose), to account for the stem cell cold
The dimethyl sulfoxide (DMSO) for freezing agent weight 1.8%, the antioxidant for accounting for the stem cell refrigerant weight 5%, 20 μM excessive
The heparin of sheep leaves of pulse plants glucoside, EDTA and 1.8IE/ml, the range of the molecular weight of the Sodium Hyaluronate are 3.5 × 104-5×104。
Embodiment 14
A kind of stem cell refrigerant as the embodiment of the present invention comprising following components,
It accounts for the human serum albumin of the stem cell refrigerant weight 4%, account for the saturating of the stem cell refrigerant weight 20%
Bright matter acid sodium, half lift-off for accounting for the stem cell refrigerant weight 15% sugared (Pentaisomaltose), to account for the stem cell cold
The dimethyl sulfoxide (DMSO) for freezing agent weight 1.8%, the antioxidant for accounting for the stem cell refrigerant weight 5%, 20 μM excessive
The heparin of sheep leaves of pulse plants glucoside, EDTA and 1.8IE/ml, the range of the molecular weight of the Sodium Hyaluronate are 3.5 × 104-5×104。
Experimental example 3
The effect of the refrigerant application of embodiment 10-14 is detected, method is the same as experimental example 1.
Cell viability testing result as shown in figure 5, as shown in Figure 5, containing embodiment 10-14, embodiment 1 other are cold
Freeze in the case of agent component, the content of the Sodium Hyaluronate in the refrigerant from 2% increase to 20% when, cell solution after freezing
Vigor after jelly increases, and illustrates that refrigerating effect improves, but after the content of Sodium Hyaluronate is greater than 8%, the degree of increase is slow,
In view of cost, embodiment 1 is preferable.And the cell viability of freezing after three weeks cell viability of two weeks compared with freezing is smaller.
Apoptosis rate testing result as shown in fig. 6, it will be appreciated from fig. 6 that containing embodiment 10-14, embodiment 1 other
In the case of refrigerant component, the content of the Sodium Hyaluronate in the refrigerant from 2% increase to 20% when, cell after freezing
Apoptosis rate after defrosting reduces, and illustrates that refrigerating effect improves, but after the content of Sodium Hyaluronate is greater than 8%, cell withers
The degree for dying rate reduction is slow, it is contemplated that cost, embodiment 1 are preferable.And the cell viability of freezing after three weeks was compared with freezing two weeks
Apoptosis rate is larger.
Comparative example 1
A kind of stem cell refrigerant as comparative example of the present invention comprising following components,
It accounts for the human serum albumin of the stem cell refrigerant weight 4%, account for the half of the stem cell refrigerant weight 15%
Lift-off sugar (Pentaisomaltose), the dimethyl sulfoxide (DMSO) for accounting for the stem cell refrigerant weight 1.8%, account for it is described
The antioxidant of stem cell refrigerant weight 5%, 20 μM of 20 μM of icariine, EDTA and 1.8IE/ml heparin.
Comparative example 2
A kind of stem cell refrigerant as comparative example of the present invention comprising following components,
It accounts for the human serum albumin of the stem cell refrigerant weight 4%, account for the transparent of the stem cell refrigerant weight 8%
Matter acid sodium, accounts for the stem cell refrigerant weight at the dimethyl sulfoxide (DMSO) for accounting for the stem cell refrigerant weight 1.8%
5% antioxidant, 20 μM of icariine, EDTA and 1.8IE/ml heparin, the model of the molecular weight of the Sodium Hyaluronate
Enclose is 3.5 × 104-5×104。
Comparative example 3
A kind of stem cell refrigerant as comparative example of the present invention comprising following components,
It accounts for the dimethyl sulfoxide (DMSO) of the stem cell refrigerant weight 5%, account for the stem cell refrigerant weight 5%
Antioxidant, EDTA and 1.8IE/ml heparin.
Experimental example 4
The effect of the refrigerant application of comparative example 1-3 is detected, method is the same as experimental example 1.
The refrigerating effect testing result of the refrigerant of comparative example 1-3 is as shown in table 1:
Finally, it should be noted that the above embodiments are merely illustrative of the technical solutions of the present invention rather than protects to the present invention
The limitation of range is protected, although the invention is described in detail with reference to the preferred embodiments, those skilled in the art should
Understand, it can be with modification or equivalent replacement of the technical solution of the present invention are made, without departing from the essence of technical solution of the present invention
And range.
Claims (10)
1. a kind of stem cell refrigerant, which is characterized in that the stem cell refrigerant includes following components in percentage by weight:
The human serum albumin of 1%-10%, the Sodium Hyaluronate of 2%-20%, 5%-30% half lift-off sugar and 1%-5% dimethyl
Sulfoxide.
2. stem cell refrigerant according to claim 1, which is characterized in that it is cold that the dimethyl sulfoxide accounts for the stem cell
Freeze the 1-3% of agent weight.
3. stem cell refrigerant according to claim 1 or 2, which is characterized in that the molecular weight of the Sodium Hyaluronate
Range is 3.5 × 104-5×104, the Sodium Hyaluronate account for the stem cell refrigerant weight 5%, 8%, 12% or
15%.
4. stem cell refrigerant according to claim 1 or 2, which is characterized in that the human serum albumin accounts for described dry thin
2%, 4%, 6% or the 8% of born of the same parents' refrigerant weight.
5. stem cell refrigerant according to claim 1 or 2, which is characterized in that the half lift-off sugar accounts for the stem cell
10%, 15%, 20% or the 25% of refrigerant weight.
6. stem cell refrigerant according to claim 1 or 2, which is characterized in that the stem cell refrigerant further includes excessive
Sheep leaves of pulse plants glucoside, content of the icariine in the stem cell refrigerant are 5-100 μM.
7. stem cell refrigerant according to claim 1 or 2, which is characterized in that the stem cell refrigerant further includes acid
Alkali regulator, the soda acid regulator are 4- hydroxyethyl piperazineethanesulfonic acid or disodium ethylene diamine tetraacetate.
8. stem cell refrigerant according to claim 1 or 2, which is characterized in that the stem cell refrigerant further includes accounting for
The antioxidant of the stem cell refrigerant weight 1%-8%, the antioxidant are vitamin C, vitamin E or lecithin.
9. stem cell refrigerant according to claim 1, which is characterized in that the stem cell refrigerant includes following group
Point,
It accounts for the human serum albumin of the stem cell refrigerant weight 2%-5%, account for the stem cell refrigerant weight 5%-15%
Sodium Hyaluronate, account for the stem cell refrigerant weight 10%-20% half lift-off sugar, account for the stem cell refrigerant weight
1.8% dimethyl sulfoxide, antioxidant, 20 μM of the icariine, soda acid for accounting for the stem cell refrigerant weight 1%-8%
The heparin of regulator and 1-2.5IE/ml.
10. a kind of if any one stem cell refrigerant of claim 1-9 is in mescenchymal stem cell or candidate stem cell
Using.
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109691435A (en) * | 2019-03-09 | 2019-04-30 | 南京温博生物科技有限公司 | A kind of freezing protective agent and simple cryopreservation methods of candidate stem cell |
| CN110199987A (en) * | 2019-07-22 | 2019-09-06 | 深圳市未来细胞生命科技有限公司 | A kind of mesenchyme stem cell preserving fluid and its store method and application |
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| CN115606583A (en) * | 2022-10-26 | 2023-01-17 | 河北生命原点生物科技有限公司 | Cryopreservation protection solution for umbilical cord stem cells, and preparation method and application thereof |
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