CN108013025A - A kind of frozen stock solution and its application - Google Patents
A kind of frozen stock solution and its application Download PDFInfo
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
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- 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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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
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Abstract
The present invention provides a kind of frozen stock solution and its application, the frozen stock solution is included by volume fraction：Dimethyl sulfoxide (DMSO) 20 40% and nutrient solution 60 80%；Wherein, combination of the nutrient solution including any one in sodium citrate, glucose, human serum albumins, glycerophosphatide, γ polyglutamic acids, dexamethasone or polyvinylpyrrolidone or at least two.γ polyglutamic acids and dexamethasone synergistic effect in frozen stock solution of the present invention, coordinate, protect the activity of various cells and cell surface antigen in blood, the haemocyte after recovery is in good condition, realizes whole blood and freezes jointly with dimethyl sulfoxide (DMSO).
Description
Technical field
The invention belongs to blood Cryopreservation Technology field, is related to a kind of frozen stock solution and its application, more particularly to a kind of for anxious
The frozen stock solution of property lymphocytic leukemia marrow and its application.
Background technology
People's whole blood is important clinical sample, is widely used in clinical diagnosis, medical research.After human body gathers blood
Need to carry out freezen protective, to avoid the inactivation or death for causing haemocyte under normal temperature environment.At present, mainly by into blood
The Cord blood that frozen stock solution carries out blood is added, the long-term storage problem of blood is solved, has ensured social blood supply.
102669087 A of CN disclose the frozen stock solution and cryopreservation methods of a kind of peripheral blood mononuclear cells, the frozen stock solution
Comprising 10-12% (V/V) permeability cryoprotector, 10-14% (V/V) beta glucan, 8-10% (V/V) serum or blood plasma,
64-72% (V/V) physiological saline, frozen stock solution using the present invention carry out peripheral blood mononuclear cells and freeze, improve cell and answer
Survival rate after Soviet Union, avoids when peripheral blood mononuclear cells is fed back to patient and adverse reaction occurs.However, the frozen stock solution
It is only used for the nucleus such as protection lymphocyte, monocyte, candidate stem cell, it is impossible to which protection is ripe red thin without nucleus
Born of the same parents.
104430303 A of CN disclose configuration and the application method of human peripheral stem cell cryopreserving liquid, and the stem cell is frozen
Liquid storage is made of HAES-Steril Infusion, recurrence amino acid injection, dimethyl sulfoxide (DMSO) and human serum albumins, is had high multiple
Revive rate, can direct venous re-transfusion, however, the frozen stock solution is only used for protection peripheral hematopoietic stem cells, in blood other are thin
Born of the same parents and growth factor do not have protective effect.
The content of the invention
In view of the deficiencies of the prior art, the present invention provides a kind of frozen stock solution and its application, the frozen stock solution especially for
Acute lymphatic leukaemia whole blood freezes, and carrying out whole blood using the frozen stock solution freezes, and can improve the cell of whole blood after recovery
Survival rate, keeping the antigens c D19 of cell surface has higher activity.
For this purpose, the present invention uses following technical scheme：
In a first aspect, the present invention provides a kind of frozen stock solution, the frozen stock solution is included by volume fraction：Dimethyl sulfoxide (DMSO)
20-40% and nutrient solution 60-80%；
Wherein, the nutrient solution include sodium citrate, glucose, human serum albumins, glycerophosphatide, γ polyglutamic acids,
In dexamethasone or polyvinylpyrrolidone any one or at least two combination.
In the frozen stock solution of the present invention, the γ polyglutamic acids have the function that heat and moisture preserving, and γ polyglutamic acids pass through bag
Haemocyte is wrapped up in, can prevent cell from being ruptured when freezing, the dexamethasone has the work of protection neutral blood granulocyte
With slow down cell expansion of the cell during freezing, γ polyglutamic acids and dexamethasone synergistic effect, with dimethyl sulfoxide (DMSO)
It is common to coordinate, protect the activity of various cells and cell surface antigen in blood.
Preferably, volume ratio of the dimethyl sulfoxide (DMSO) in the frozen stock solution is 20-40%, such as can be 20%,
21%th, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%,
36%th, 37%, 38%, 39% or 40%, it is preferably 30%.
Preferably, volume ratio of the nutrient solution in the frozen stock solution is 60-80%, for example, can be 60%, 61%,
62%th, 63%, 64%, 65%, 66%, 67%, 68%, 69%, 70%, 71%, 72%, 73%, 74%, 75%, 76%,
77%th, 78%, 79% or 80%, it is preferably 70%.
Preferably, the nutrient solution includes sodium citrate, glucose, human serum albumins, glycerophosphatide, γ polyglutamics
In acid, dexamethasone or polyvinylpyrrolidone any one or at least two combination, be preferably sodium citrate, grape
The combination of sugar, human serum albumins, glycerophosphatide, γ polyglutamic acids, dexamethasone and polyvinylpyrrolidone.
Preferably, the nutrient solution includes by mass percentage：Sodium citrate 1-5%, glucose 5-10%, human seralbumin
Albumen 5-20%, glycerophosphatide 5-20%, γ polyglutamic acid 10-20%, dexamethasone 1-5%, polyvinylpyrrolidone 1-
2%, surplus is physiological saline.
Preferably, mass percent of the sodium citrate in the nutrient solution is 1-5%, such as can be 1%,
2%th, 3%, 4% or 5%, it is preferably 3%.
Preferably, mass percent of the glucose in the nutrient solution is 5-10%, such as can be 5%,
6%th, 7%, 8%, 9% or 10%, it is preferably 8%.
Preferably, mass percent of the human serum albumins in the nutrient solution is 5-20%, such as can be
5%th, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19% or 20%,
Preferably 10%.
Preferably, mass percent of the glycerophosphatide in the nutrient solution is 5-20%, such as can be 5%,
6%th, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19% or 20%, preferably
For 10%.
Preferably, mass percent of the γ polyglutamic acids in the nutrient solution is 10-20%, such as can be
10%th, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19% or 20%, it is preferably 15%.
Preferably, mass percent of the dexamethasone in the nutrient solution is 1-5%, such as can be 1%,
2%th, 3%, 4% or 5%, it is preferably 2%.
Preferably, mass percent of the polyvinylpyrrolidone in the nutrient solution is 1-2%, such as can be
1% or 2%, it is preferably 1%.
Preferably, the frozen stock solution is included by volume fraction：Dimethyl sulfoxide (DMSO) 30% and nutrient solution 70%；
Wherein, the nutrient solution includes by mass percentage：Sodium citrate 3%, glucose 8%, human serum albumins
10%, glycerophosphatide 10%, γ polyglutamic acids 15%, dexamethasone 2%, polyvinylpyrrolidone 1%, surplus is physiology salt
Water.
Preferably, the number-average molecular weight of the γ polyglutamic acids is 90-120 ten thousand, such as can be 900,000,950,000,100
Ten thousandth, 1,050,000,1,100,000,1,150,000 or 1,200,000, it is preferably 1,000,000.
Preferably, the number-average molecular weight of the polyvinylpyrrolidone is 3-5 ten thousand, such as can be 30,000,40,000 or 50,000,
Preferably 40,000.
Second aspect, the present invention provides a kind of whole blood to freeze the method with recovery, and the method uses such as first aspect
The frozen stock solution.
Preferably, the described method comprises the following steps：
(1) freeze：After gathering whole blood to cryopreservation tube, the frozen stock solution is added, gradient cooling is to -80 DEG C；
(2) recover：It will be placed in 35-40 DEG C of water-bath, dissolved in the backward whole blood equipped with the cryopreservation tube for treating recovery whole blood
The nutrient solution is added, the whole blood recovered.
Preferably, the volume ratio of step (1) whole blood and the frozen stock solution is 10:(0.5-2), such as can be 10:
0.5、10:0.8、10:1、10:1.2、10:1.5、10:1.8 or 10:2, it is preferably 10:1.
Preferably, the step of step (1) described gradient cooling is：
Keep 5-60min at 4 DEG C of (1 '), for example, can be 5min, 10min, 15min, 20min, 25min, 30min,
35min, 40min, 45min, 50min, 55min or 60min；
(2 ') with the speed of 1-10 DEG C/min, such as can be with 1 DEG C/min, 2 DEG C/min, 3 DEG C/min, 4 DEG C/min, 5
DEG C/min, 6 DEG C/min, 7 DEG C/min, 8 DEG C/min, the speed of 9 DEG C/min or 10 DEG C/min, preferably with the speed of 5 DEG C/min
It is cooled to 0 DEG C；
(3 ') are with the speed of 1-5 DEG C/min, such as can be with 1 DEG C/min, 2 DEG C/min, 3 DEG C/min, 4 DEG C/min or 5
DEG C/speed of min, preferably -20 DEG C are cooled to the speed of 3 DEG C/min；
(4 ') with the speed of 1-2 DEG C/min, such as can be with the speed of 1 DEG C/min or 2 DEG C/min, preferably with 1 DEG C/
The speed of min is cooled to -80 DEG C.
Preferably, the temperature of step (2) described water-bath is 35-40 DEG C, for example, can be 35 DEG C, 36 DEG C, 37 DEG C, 38 DEG C,
39 DEG C or 40 DEG C, be preferably 37 DEG C.
Preferably, the volume ratio of step (2) whole blood and the nutrient solution is 10:(1-5), such as can be 10:1、
10:2、10:3、10:4 or 10:5, it is preferably 10:3.
As optimal technical scheme, the present invention provides a kind of whole blood to freeze the method with recovery, the described method includes with
Lower step：
(1) freeze：It is 10 by whole blood/frozen stock solution volume ratio after gathering whole blood to cryopreservation tube:(0.5-2) adds the jelly
Liquid storage, keeps 5-60min by cryopreservation tube at 4 DEG C after mixing, 0 DEG C is cooled to the speed of 1-10 DEG C/min, with 1-5
DEG C/speed of min is cooled to -20 DEG C, finally -80 DEG C are cooled to the speed of 1-2 DEG C/min；
(2) recover：It will be placed in equipped with the cryopreservation tube for treating recovery whole blood in 35-40 DEG C of water-bath, it is backward described complete to shake dissolving
The volume ratio that whole blood/nutrient solution is pressed in blood is 10:(1-5) adds the nutrient solution, the whole blood recovered.
The third aspect, the present invention provides a kind of method as described in second aspect to freeze the whole blood with recovery.
Preferably, the whole blood is acute lymphoblastic leukemia marrow.
Fourth aspect, is detected the present invention provides a kind of whole blood as described in the third aspect for antigen.
Preferably, the whole blood is acute lymphoblastic leukemia marrow.
Compared with prior art, the present invention has the advantages that：
(1) frozen stock solution of the invention includes γ polyglutamic acids with heat and moisture preserving effect and with protecting neutrality grain thin
The dexamethasone of born of the same parents, both act synergistically, coordinate jointly with dimethyl sulfoxide (DMSO), protect various cells and cell surface in blood
The activity of antigen；
(2) present invention carries out the recovery of frozen blood using nutrient solution, and the cell state after recovery is good, and cell is bright,
Size is homogeneous, and after birth is complete, and survival rate is up to 94%, and the recall rate of cell surface antigen CD19 is up to 36.58%；
(3) frozen stock solution using the present invention, realizes whole blood and freezes, and is of great significance to clinical diagnosis research.
Brief description of the drawings
Fig. 1 (A) is the flow cytomery that whole blood freezes as a result, Fig. 1 (B) is the fluidic cell that freezes after whole blood cracking
Instrument testing result.
Embodiment
The technological means and its effect taken for the present invention is further explained, with reference to embodiments with attached drawing to this hair
It is bright to be further described.It is understood that the specific embodiments described herein are used only for explaining the present invention, rather than
Limitation of the invention.
In the examples where no specific technique or condition is specified, according to the described technology of document in the art or condition,
Or carried out according to product description.Reagents or instruments used without specified manufacturer, be can be by regular channel commercially available from
The conventional products of acquisition.
Embodiment 1
Frozen stock solution (I) is included by volume fraction：Dimethyl sulfoxide (DMSO) 30% and nutrient solution 70%, wherein, the nutrient solution
Formula is shown in Table 1.
Table 1
Whole blood is carried out using frozen stock solution (I) to freeze, and is recovered, and is concretely comprised the following steps：
(1) freeze：It is 10 by whole blood/frozen stock solution volume ratio after gathering whole blood to cryopreservation tube:1 adds the frozen stock solution, mixes
Cryopreservation tube is kept into 30min at 4 DEG C after closing uniformly, 0 DEG C is cooled to the speed of 5 DEG C/min, is cooled down with the speed of 3 DEG C/min
To -20 DEG C, finally -80 DEG C are cooled to the speed of 1 DEG C/min；
(2) recover：It will be placed in 37 DEG C of water-baths, shaken in the backward whole blood of dissolving equipped with the cryopreservation tube for treating recovery whole blood
It is 10 by the volume ratio of whole blood/nutrient solution:3 add the nutrient solution, the whole blood recovered.
Embodiment 2
Frozen stock solution (II) is included by volume fraction：Dimethyl sulfoxide (DMSO) 20% and nutrient solution 80%, wherein, the nutrient solution
Formula is shown in Table 2.
Table 2
Whole blood is carried out using frozen stock solution (II) to freeze, and is recovered, and is concretely comprised the following steps：
(1) freeze：It is 20 by whole blood/frozen stock solution volume ratio after gathering whole blood to cryopreservation tube:1 adds the frozen stock solution, mixes
Cryopreservation tube is kept into 60min at 4 DEG C after closing uniformly, 0 DEG C is cooled to the speed of 1 DEG C/min, is cooled down with the speed of 1 DEG C/min
To -20 DEG C, finally -80 DEG C are cooled to the speed of 1 DEG C/min；
(2) recover：It will be placed in 35 DEG C of water-baths, shaken in the backward whole blood of dissolving equipped with the cryopreservation tube for treating recovery whole blood
It is 10 by the volume ratio of whole blood/nutrient solution:1 adds the nutrient solution, the whole blood recovered.
Embodiment 3
Frozen stock solution (III) is included by volume fraction：Dimethyl sulfoxide (DMSO) 40% and nutrient solution 60%, wherein, the nutrient solution
Formula be shown in Table 3.
Table 3
| Ingredient names | Mass percent % |
| Sodium citrate | 5 |
| Glucose | 5 |
| Human serum albumins | 20 |
| Glycerophosphatide | 5 |
| γ polyglutamic acids (900,000) | 20 |
| Dexamethasone | 1 |
| Polyvinylpyrrolidone (30,000) | 2 |
| Physiological saline | 42 |
Whole blood is carried out using frozen stock solution (III) to freeze, and is recovered, and is concretely comprised the following steps：
(1) freeze：It is 5 by whole blood/frozen stock solution volume ratio after gathering whole blood to cryopreservation tube:1 adds the frozen stock solution, mixes
Cryopreservation tube is kept into 5min at 4 DEG C after closing uniformly, 0 DEG C is cooled to the speed of 10 DEG C/min, is cooled down with the speed of 5 DEG C/min
To -20 DEG C, finally -80 DEG C are cooled to the speed of 2 DEG C/min；
(2) recover：It will be placed in 40 DEG C of water-baths, shaken in the backward whole blood of dissolving equipped with the cryopreservation tube for treating recovery whole blood
It is 10 by the volume ratio of whole blood/nutrient solution:5 add the nutrient solution, the whole blood recovered.
Comparative example 1
Compared with Example 1, γ polyglutamic acids are free of in nutrient solution, the mass percent of physiological saline is 66%, other
Component and mass percent are same as Example 1.
Comparative example 2
Compared with Example 1, dexamethasone is free of in nutrient solution, the mass percent of physiological saline is 52%, other groups
Divide and mass percent is same as Example 1.
Comparative example 3
Compared with Example 1, the mass percent of γ polyglutamic acids is 5%, and the mass percent of physiological saline is 61%,
Other components and mass percent are same as Example 1.
Comparative example 4
Compared with Example 1, the mass percent of γ polyglutamic acids is 25%, and the mass percent of physiological saline is
41%, other components and mass percent are same as Example 1.
Comparative example 5
Compared with Example 1, the number-average molecular weight of γ polyglutamic acids is 500,000, other components and mass percent and implementation
Example 1 is identical.
Comparative example 6
Compared with Example 1, the number-average molecular weight of γ polyglutamic acids is 1,500,000, other components and mass percent and reality
It is identical to apply example 1.
Comparative example 7
Compared with Example 4, the whole blood of collection is subjected to centrifugation splitting erythrocyte processing, is by volume 5:1 adds jelly
Liquid storage.
Comparative example 8
Compared with Example 4, it is 5 by whole blood/frozen stock solution volume ratio into whole blood:After 1 adds frozen stock solution, without gradient
Cooling, is placed directly within -80 DEG C and freezes.
Comparative example 9
Compared with Example 4, it is 100 by whole blood/frozen stock solution volume ratio into whole blood:After 1 adds frozen stock solution.
Comparative example 10
Compared with Example 4, it is 2 by whole blood/frozen stock solution volume ratio into whole blood:After 1 adds frozen stock solution.
Cell state, survival rate and CD19 antigen verification and measurement ratios
Cell state, survival rate and cell surface antigen CD19 in blood after embodiment and comparative example recovery are examined
Survey, blood sample comes from Patients With Acute Lymphoblastic Leukemia, the results are shown in Table 4.
4 cell state of table, survival rate and CD19 antigen testing results
In the whole blood that embodiment 1-3 recovers, cell is bright, size is homogeneous, after birth is complete, and in good shape, cell is deposited
For motility rate more than 90%, the recall rate of cell surface CD19 antigens is up to 36.58%.
Compared with Example 1, the frozen stock solution of comparative example 1 is free of γ polyglutamic acids so that a large amount of whole blood cells are when freezing
Rupture, cell survival rate only 35%, and the cell state survived is poor, it is intensely dark, imperfect without fixed form, after birth,
Cell surface CD19 antigens largely inactivate, recall rate only 4.62%；The frozen stock solution of comparative example 2 is free of dexamethasone so that whole blood
In neutrophil leucocyte inactivation, cell survival rate is relatively low；The mass percent of the γ polyglutamic acids of comparative example 3-4 is unreasonable, no
It can fully be cooperateed with dexamethasone and dimethyl sulfoxide (DMSO), protect the ability of cell weaker；The number of the γ polyglutamic acids of comparative example 5-6
Average molecular weight is unreasonable, without the effect for keeping moisture, protecting cell；Comparative example 7 by the whole blood first choice after collection carry out from
Heart cracking operation, damages a large amount of cells, and cell survival rate is down to 26%, the CD19 antigens of cell surface, recall rate only 8.39%；
Comparative example 8 is without gradient operation, and cell interior forms ice needle during freezing, and cell rupture does not fix form, and after birth is not
Completely, survival rate is relatively low；The addition of the frozen stock solution of comparative example 9-10 is unreasonable, significantly affects and freezes effect.
Shown in flow cytometer detection result such as Fig. 1 (A) and Fig. 1 (B) of the CD19 of embodiment 1 and comparative example 7.Whole blood freezes thin
The recall rate of born of the same parents CD19 be 36.58%, and crack after cell CD19 recall rate only 8.39%.
In conclusion in the frozen stock solution of the present invention, the γ polyglutamic acids are by wrapping up haemocyte, it is therefore prevented that cell exists
Ruptured when freezing, the dexamethasone slow down cell expansion of the cell during freezing, and γ polyglutamic acids and ground are filled in
Rice pine synergistic effect, coordinates, protects the activity of various cells and cell surface antigen in blood jointly with dimethyl sulfoxide (DMSO), multiple
Haemocyte after Soviet Union is in good condition, and cell is bright, and size is homogeneous, and after birth is complete, and survival rate is up to 94%, cell surface antigen
The recall rate of CD19 is up to 36.58%, realizes whole blood and freezes, and is of great significance to clinical diagnosis research.
Applicant states that the present invention illustrates the method detailed of the present invention, but not office of the invention by above-described embodiment
It is limited to above-mentioned method detailed, that is, does not mean that the present invention has to rely on above-mentioned method detailed and could implement.Technical field
Technical staff it will be clearly understood that any improvement in the present invention, equivalence replacement and auxiliary element to each raw material of product of the present invention
Addition, selection of concrete mode etc., all fall within protection scope of the present invention and the open scope.
Claims (10)
1. a kind of frozen stock solution, it is characterised in that the frozen stock solution is included by volume fraction：Dimethyl sulfoxide (DMSO) 20-40% and nutrition
Liquid 60-80%；
Wherein, the nutrient solution include sodium citrate, glucose, human serum albumins, glycerophosphatide, γ polyglutamic acids, fill in
Rice pine or polyvinylpyrrolidone in any one or at least two combination.
2. frozen stock solution according to claim 1, it is characterised in that the nutrient solution includes by mass percentage：Citric acid
Sodium 1-5%, glucose 5-10%, human serum albumins 5-20%, glycerophosphatide 5-20%, γ polyglutamic acid 10-20%, ground plug
Rice pine 1-5%, polyvinylpyrrolidone 1-2%, surplus is physiological saline.
3. frozen stock solution according to claim 1 or 2, it is characterised in that the frozen stock solution is included by volume fraction：Dimethyl
Sulfoxide 30% and nutrient solution 70%；
Wherein, the nutrient solution includes by mass percentage：Sodium citrate 3%, glucose 8%, human serum albumins 10% are sweet
Oily phosphatidase 1 0%, γ polyglutamic acids 15%, dexamethasone 2%, polyvinylpyrrolidone 1%, surplus are physiological saline.
4. according to claim 1-3 any one of them frozen stock solutions, it is characterised in that the number-average molecular weight of the γ polyglutamic acids
It is preferably 1,000,000 for 90-120 ten thousand；
Preferably, the number-average molecular weight of the polyvinylpyrrolidone is 3-5 ten thousand, is preferably 40,000.
5. a kind of whole blood freezes the method with recovery, it is characterised in that the method is used as described in claim any one of 1-4
Frozen stock solution.
6. according to the method described in claim 5, it is characterised in that it includes following steps：
(1) freeze：After gathering whole blood to cryopreservation tube, the frozen stock solution is added, gradient cooling is to -80 DEG C；
(2) recover：It will be placed in equipped with the cryopreservation tube for treating recovery whole blood in 35-40 DEG C of water-bath, dissolve and added in the backward whole blood
The nutrient solution, the whole blood recovered.
7. the method according to claim 5 or 6, it is characterised in that the volume of step (1) whole blood and the frozen stock solution
Than for 10:(0.5-2), is preferably 10:1；
Preferably, the step of step (1) described gradient cooling is：
5-60min is kept at 4 DEG C of (1 ')；
(2 ') are preferably cooled to 0 DEG C with the speed of 1-10 DEG C/min with the speed of 5 DEG C/min；
(3 ') are preferably cooled to -20 DEG C with the speed of 1-5 DEG C/min with the speed of 3 DEG C/min；
(4 ') are preferably cooled to -80 DEG C with the speed of 1-2 DEG C/min with the speed of 1 DEG C/min；
Preferably, the temperature of step (2) described water-bath is 37 DEG C；
Preferably, the volume ratio of step (2) whole blood and the nutrient solution is 10:(1-5), is preferably 10:3.
8. according to claim 5-7 any one of them methods, it is characterised in that comprise the following steps：
(1) freeze：It is 10 by whole blood/frozen stock solution volume ratio after gathering whole blood to cryopreservation tube:(0.5-2) adds the frozen stock solution,
Cryopreservation tube is kept into 5-60min at 4 DEG C after mixing, 0 DEG C is cooled to the speed of 1-10 DEG C/min, with 1-5 DEG C/min
Speed be cooled to -20 DEG C, be finally cooled to -80 DEG C with the speed of 1-2 DEG C/min；
(2) recover：It will be placed in 35-40 DEG C of water-bath, shaken in the backward whole blood of dissolving equipped with the cryopreservation tube for treating recovery whole blood
It is 10 by the volume ratio of whole blood/nutrient solution:(1-5) adds the nutrient solution, the whole blood recovered.
9. a kind of method as described in claim 5-8 freezes the whole blood with recovery；
Preferably, the whole blood is acute lymphoblastic leukemia marrow.
10. a kind of whole blood as claimed in claim 9 is detected for antigen；
Preferably, the whole blood is acute lymphoblastic leukemia marrow.
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201711251065.3A CN108013025A (en) | 2017-12-01 | 2017-12-01 | A kind of frozen stock solution and its application |
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Cited By (3)
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| CN108990964A (en) * | 2018-08-09 | 2018-12-14 | 华东理工大学 | Cells frozen storing liquid |
| CN111418580A (en) * | 2020-05-25 | 2020-07-17 | 山东万能干细胞生物技术有限公司 | Stem cell cryopreservation solution and cryopreservation method |
| CN111642557A (en) * | 2020-05-11 | 2020-09-11 | 广州市博仕奥生化技术研究有限公司 | Antifreezing agent and using method thereof |
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| CN111642557A (en) * | 2020-05-11 | 2020-09-11 | 广州市博仕奥生化技术研究有限公司 | Antifreezing agent and using method thereof |
| CN111418580A (en) * | 2020-05-25 | 2020-07-17 | 山东万能干细胞生物技术有限公司 | Stem cell cryopreservation solution and cryopreservation method |
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