CN107347874A - A kind of frozen stock solution of umbilical hemopoietic stem cell - Google Patents
A kind of frozen stock solution of umbilical hemopoietic stem cell Download PDFInfo
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- CN107347874A CN107347874A CN201710678108.XA CN201710678108A CN107347874A CN 107347874 A CN107347874 A CN 107347874A CN 201710678108 A CN201710678108 A CN 201710678108A CN 107347874 A CN107347874 A CN 107347874A
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- stem cell
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- hemopoietic stem
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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 invention discloses a kind of frozen stock solution of umbilical hemopoietic stem cell, the frozen stock solution is made up of freezing protective agent, hyclone, culture medium；The culture medium is composed of the following components：Basal medium, human serum albumins, myricetin, arasaponin, rheum emodin, linoleic acid, trypsase, Aprotinin, lumican, basic fibroblast growth factor, folic acid, β mercaptoethanols, astragalus polyose, transferrins, vitamin, amino acid.Umbilical hemopoietic stem cell is frozen using frozen stock solution of the present invention, greatly reduce damaging action of the frozen stock solution to umbilical hemopoietic stem cell, be advantageous to umbilical hemopoietic stem cell freeze and resuscitation process in cell viability raising, can preferably keep the biological property of umbilical hemopoietic stem cell.
Description
Technical field
The invention belongs to biological technical field, more particularly to a kind of frozen stock solution of umbilical hemopoietic stem cell.
Background technology
Bleeding of the umbilicus is also Cord blood, and the blood in placenta and umbilical cord is remained in after referring to delivery of baby, ligature of the cord and detachment,
It is typically what is passed into disuse.The research of nearly more than ten years finds, containing can rebuild making for human body hematopoiesis and immune system in bleeding of the umbilicus
Hemocytoblast, available for HSCT, treat a variety of diseases.Therefore, bleeding of the umbilicus is as the important next of candidate stem cell
The source in source, particularly consanguinity-less relation candidate stem cell.A kind of and very important human biological's resource.
Contain substantial amounts of stem cell in bleeding of the umbilicus, stem cell is the seed of life, and it can break up the various cells of adult body, knot
Go out a variety of fruits --- blood cell, nerve cell, bone cellses etc..With the prosperity of science and technology, medical expert grinds
Study carefully the method that disease is treated using the stem cell in Cord blood.Stem cell is with self-renewing, hyperproliferation and multinomial
The cell colony of differentiation potential.These cells can maintain the characteristic and quantity of own cells by dividing, and can further divide again
Various histocytes are turned to, so as to be played a positive role in tissue repair etc..Medical research discovery in the last thirty years, navel
Containing very abundant candidate stem cell (HSC) in blood, human body hematopoiesis and immune system can be rebuild, available for candidate stem cell
Transplanting, treatment hematological system, immune system, and hereditary metabolic disorders and congenital disorders.Therefore, it is thin to have turned into Hematopoietic Stem for bleeding of the umbilicus
The important sources of born of the same parents, have been widely used in clinic, are valuable human biological's resources.
In world wide, at present bleeding of the umbilicus have been used to treat all kinds leukaemia, alpastic anemia, lymthoma,
Huppert's disease, neuroblastoma, mucopolysaccharidosis, thalassemia, osteomyelodysplasia syndrome, primary are immune to be lacked
Fall into more than the 80 kinds of diseases such as disease, chronic granuloma.
The Ministry of Public Health of China issues《Umbilical cord blood hematopoietic stem cell treatment technology management regulation (tentative)》Middle regulation, Cord blood
Candidate stem cell treatment technology indication includes following more than 10 class：Marrow failure, hemoglobinopathy, severe immunologic deficiency disease, generation
Thanking property disease, leukaemia, RAEB, malignant lymphoma, Huppert's disease, malignant tumour, severe regeneration
Aplastic anemia and severe radiation sickness etc..
The content of the invention
It is an object of the invention to provide a kind of frozen stock solution of umbilical hemopoietic stem cell, umbilical hemopoietic is done with reducing frozen stock solution
The damaging action of cell.
To achieve the above object, the technical solution adopted by the present invention is：
A kind of frozen stock solution of umbilical hemopoietic stem cell, the frozen stock solution is by freezing protective agent, hyclone, culture medium group
Into；
The culture medium is composed of the following components：
Basal medium, human serum albumins, myricetin, arasaponin, rheum emodin, linoleic acid, trypsase, suppression peptide
Enzyme, lumican, basic fibroblast growth factor, pyruvic acid, Glu, folic acid, beta -mercaptoethanol, the Radix Astragali are more
Sugar, transferrins, vitamin, amino acid.
Preferably, in the culture medium, the content of each component is as follows：
Basal medium 10-50g/L, human serum albumins 5-20g/L, myricetin 5-20mg/L, arasaponin 5-
20mg/L, rheum emodin 5-20mg/L, linoleic acid 2-10mg/L, trypsase 1-10mg/L, Aprotinin 4-10mg/L, photoprotein gather
Sugared 5-20 μ g/L, basic fibroblast growth factor 1-10 μ g/L, pyruvic acid 5-15mg/L, Glu 5-20mg/L,
Folic acid 1-15mg/L, beta -mercaptoethanol 2-10mg/L, astragalus polyose 50-80mg/L, transferrins 1-10mg/L, vitamin 80-
500mg/L, amino acid 80-500mg/L.
It is furthermore preferred that in the culture medium, the content of each component is as follows：
Basal medium 20-30g/L, human serum albumins 10-15g/L, myricetin 10-15mg/L, arasaponin 10-
15mg/L, rheum emodin 10-15mg/L, linoleic acid 5-8mg/L, trypsase 4-8mg/L, Aprotinin 6-9mg/L, lumican
10-15 μ g/L, HBGH-2-8 μ g/L, pyruvic acid 8-12mg/L, Glu 10-15mg/L, leaf
Sour 5-10mg/L, beta -mercaptoethanol 5-8mg/L, astragalus polyose 60-70mg/L, transferrins 2-8mg/L, vitamin 180-
360mg/L, amino acid/11 80-360mg/L.
Further preferably, in the culture medium, the content of each component is as follows：
Basal medium 24g/L, human serum albumins 12g/L, myricetin 12mg/L, arasaponin 12mg/L, rheum officinale
Plain 13mg/L, linoleic acid 6mg/L, trypsase 5mg/L, Aprotinin 8mg/L, the μ g/L of lumican 13, basic fibroblast are thin
The intracellular growth factor 5 μ g/L, pyruvic acid 11mg/L, Glu 12mg/L, folic acid 8mg/L, beta -mercaptoethanol 6mg/L, the Radix Astragali
Polysaccharide 68mg/L, transferrins 5mg/L, vitamin 280mg/L, amino acid 260mg/L.
Preferably, the human serum albumins behaviour bleeding of the umbilicus human serum albumins；Basal medium is IMDM culture mediums；Institute
It is water soluble vitamin to state vitamin；The amino acid is several amino acids compound.
Preferably, the freezing protective agent is permeability cryoprotector, by glycerine, dimethyl sulfoxide (DMSO), ethylene glycol, the third two
Alcohol, acetamide, one or more compositions of methanol.
Preferably, the freezing protective agent, hyclone, the volume ratio of culture medium are 1：1~3：2~4.
Beneficial effect：Umbilical hemopoietic stem cell is frozen using frozen stock solution of the present invention, greatly reduces frozen stock solution pair
The damaging action of candidate stem cell, be advantageous to umbilical hemopoietic stem cell freeze and resuscitation process in cell viability raising, energy
Enough biological properties for preferably keeping umbilical hemopoietic stem cell.
Embodiment
According to following embodiments, the present invention can be better understood from.It is however, as it will be easily appreciated by one skilled in the art that real
Apply specific material proportion, process conditions and its result described by example and be merely to illustrate the present invention, without that will not also should limit
The present invention described in detail in claims processed.
Embodiment 1
A kind of frozen stock solution of umbilical hemopoietic stem cell, by freezing protective agent, hyclone, culture medium according to volume ratio be 1：
1：4 compositions；
Wherein, in culture medium, the content of each component is as follows：Basal medium 24g/L, human serum albumins 12g/L, red bayberry
Plain 12mg/L, arasaponin 12mg/L, rheum emodin 13mg/L, linoleic acid 6mg/L, trypsase 5mg/L, Aprotinin 8mg/L,
The μ g/L of lumican 13, basic fibroblast growth factor 5 μ g/L, pyruvic acid 11mg/L, Glu 12mg/L, leaf
Sour 8mg/L, beta -mercaptoethanol 6mg/L, astragalus polyose 68mg/L, transferrins 5mg/L, vitamin 280mg/L, amino acid
260mg/L。
Freezing protective agent is dimethyl sulfoxide (DMSO).
Umbilical hemopoietic stem cell is taken, is added in above-mentioned frozen stock solution, piping and druming is uniform, forms cell suspension, is placed in cryopreservation tube,
Umbilical hemopoietic stem cell is frozen.
Embodiment 2
A kind of frozen stock solution of umbilical hemopoietic stem cell, by freezing protective agent, hyclone, culture medium according to volume ratio be 1：
2：2 compositions；
Wherein, in culture medium, the content of each component is as follows：Basal medium 10g/L, human serum albumins 15g/L, red bayberry
Plain 10mg/L, arasaponin 15mg/L, rheum emodin 10mg/L, linoleic acid 2mg/L, trypsase 10mg/L, Aprotinin 10mg/
L, the μ g/L of lumican 10, basic fibroblast growth factor 1 μ g/L, pyruvic acid 5mg/L, Glu 15mg/L,
Folic acid 15mg/L, beta -mercaptoethanol 10mg/L, astragalus polyose 80mg/L, transferrins 1mg/L, vitamin 360mg/L, amino acid
180mg/L。
Freezing protective agent is dimethyl sulfoxide (DMSO), glycerine, the mixture of acetamide.
Umbilical hemopoietic stem cell is taken, is added in above-mentioned frozen stock solution, piping and druming is uniform, forms cell suspension, is placed in cryopreservation tube,
Umbilical hemopoietic stem cell is frozen.
Embodiment 3
A kind of frozen stock solution of umbilical hemopoietic stem cell, by freezing protective agent, hyclone, culture medium according to volume ratio be 1：
3：3 compositions；
Wherein, in culture medium, the content of each component is as follows：Basal medium 50g/L, human serum albumins 5g/L, red bayberry
Plain 5mg/L, arasaponin 20mg/L, rheum emodin 5mg/L, linoleic acid 8mg/L, trypsase 1mg/L, Aprotinin 4mg/L, light
The μ g/L of proteoglycans 20, basic fibroblast growth factor 8 μ g/L, pyruvic acid 8mg/L, Glu 5mg/L, folic acid
5mg/L, beta -mercaptoethanol 2mg/L, astragalus polyose 50mg/L, transferrins 3mg/L, vitamin 500mg/L, amino acid 80mg/
L。
Freezing protective agent is dimethyl sulfoxide (DMSO), ethylene glycol, the mixture of propane diols.
Umbilical hemopoietic stem cell is taken, is added in above-mentioned frozen stock solution, piping and druming is uniform, forms cell suspension, is placed in cryopreservation tube,
Umbilical hemopoietic stem cell is frozen.
Embodiment 4
A kind of frozen stock solution of umbilical hemopoietic stem cell, by freezing protective agent, hyclone, culture medium according to volume ratio be 1：
3：4 compositions；
Wherein, in culture medium, the content of each component is as follows：Basal medium 30g/L, human serum albumins 10g/L, red bayberry
Plain 15mg/L, arasaponin 10mg/L, rheum emodin 15mg/L, linoleic acid 8mg/L, trypsase 8mg/L, Aprotinin 6mg/L,
The μ g/L of lumican 15, HBGH-2 μ g/L, pyruvic acid 12mg/L, Glu 20mg/L, leaf
Sour 5mg/L, beta -mercaptoethanol 8mg/L, astragalus polyose 60mg/L, transferrins 2mg/L, vitamin 180mg/L, amino acid
500mg/L。
Freezing protective agent is methanol, ethylene glycol, the mixture of propane diols.
Umbilical hemopoietic stem cell is taken, is added in above-mentioned frozen stock solution, piping and druming is uniform, forms cell suspension, is placed in cryopreservation tube,
Umbilical hemopoietic stem cell is frozen.
Embodiment 5
A kind of frozen stock solution of umbilical hemopoietic stem cell, by freezing protective agent, hyclone, culture medium according to volume ratio be 1：
2：4 compositions；
Wherein, in culture medium, the content of each component is as follows：Basal medium 50g/L, human serum albumins 5g/L, red bayberry
Plain 20mg/L, arasaponin 5mg/L, rheum emodin 20mg/L, linoleic acid 4mg/L, trypsase 1mg/L, Aprotinin 4mg/L,
The μ g/L of lumican 5, basic fibroblast growth factor 10 μ g/L, pyruvic acid 15mg/L, Glu 10mg/L, leaf
Sour 1mg/L, beta -mercaptoethanol 2mg/L, astragalus polyose 50mg/L, transferrins 10mg/L, vitamin 240mg/L, amino acid
240mg/L。
Freezing protective agent is glycerine.
Umbilical hemopoietic stem cell is taken, is added in above-mentioned frozen stock solution, piping and druming is uniform, forms cell suspension, is placed in cryopreservation tube,
Umbilical hemopoietic stem cell is frozen.
Described above is only the preferred embodiment of the present invention, it is noted that for the ordinary skill people of the art
For member, under the premise without departing from the principles of the invention, some improvements and modifications can also be made, these improvements and modifications also should
It is considered as protection scope of the present invention.
Claims (7)
- A kind of 1. frozen stock solution of umbilical hemopoietic stem cell, it is characterised in that：The frozen stock solution by freezing protective agent, hyclone, Culture medium forms；The culture medium is composed of the following components：Basal medium, human serum albumins, myricetin, arasaponin, rheum emodin, linoleic acid, trypsase, Aprotinin, Lumican, basic fibroblast growth factor, pyruvic acid, Glu, folic acid, beta -mercaptoethanol, astragalus polyose, Transferrins, vitamin, amino acid.
- 2. the frozen stock solution of umbilical hemopoietic stem cell according to claim 1, it is characterised in that：In the culture medium, each group The content divided is as follows：Basal medium 10-50g/L, human serum albumins 5-20g/L, myricetin 5-20mg/L, arasaponin 5-20mg/L, Rheum emodin 5-20mg/L, linoleic acid 2-10mg/L, trypsase 1-10mg/L, Aprotinin 4-10mg/L, lumican 5-20 μ G/L, basic fibroblast growth factor 1-10 μ g/L, pyruvic acid 5-15mg/L, Glu 5-20mg/L, folic acid 1- 15mg/L, beta -mercaptoethanol 2-10mg/L, astragalus polyose 50-80mg/L, transferrins 1-10mg/L, vitamin 80-500mg/ L, amino acid 80-500mg/L.
- 3. the frozen stock solution of umbilical hemopoietic stem cell according to claim 2, it is characterised in that：In the culture medium, each group The content divided is as follows：Basal medium 20-30g/L, human serum albumins 10-15g/L, myricetin 10-15mg/L, arasaponin 10- 15mg/L, rheum emodin 10-15mg/L, linoleic acid 5-8mg/L, trypsase 4-8mg/L, Aprotinin 6-9mg/L, lumican 10-15 μ g/L, HBGH-2-8 μ g/L, pyruvic acid 8-12mg/L, Glu 10-15mg/L, leaf Sour 5-10mg/L, beta -mercaptoethanol 5-8mg/L, astragalus polyose 60-70mg/L, transferrins 2-8mg/L, vitamin 180- 360mg/L, amino acid/11 80-360mg/L.
- 4. the frozen stock solution of umbilical hemopoietic stem cell according to claim 3, it is characterised in that：In the culture medium, each group The content divided is as follows：Basal medium 24g/L, human serum albumins 12g/L, myricetin 12mg/L, arasaponin 12mg/L, rheum emodin 13mg/L, linoleic acid 6mg/L, trypsase 5mg/L, Aprotinin 8mg/L, the μ g/L of lumican 13, basic fibroblast Growth factors 5 μ g/L, pyruvic acid 11mg/L, Glu 12mg/L, folic acid 8mg/L, beta -mercaptoethanol 6mg/L, the Radix Astragali are more Sugared 68mg/L, transferrins 5mg/L, vitamin 280mg/L, amino acid 260mg/L.
- 5. according to the frozen stock solution of any described umbilical hemopoietic stem cells of claim 1-4, it is characterised in that：The human seralbumin Albumen behaviour bleeding of the umbilicus human serum albumins；Basal medium is IMDM culture mediums；The vitamin is water soluble vitamin；It is described Amino acid is several amino acids compound.
- 6. the frozen stock solution of umbilical hemopoietic stem cell according to claim 1, it is characterised in that：The freezing protective agent is to ooze Permeability cryoprotector, it is made up of the one or more of glycerine, dimethyl sulfoxide (DMSO), ethylene glycol, propane diols, acetamide, methanol.
- 7. the frozen stock solution of umbilical hemopoietic stem cell according to claim 1, it is characterised in that：The freezing protective agent, tire Cow's serum, the volume ratio of culture medium are 1：1~3：2~4.
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109913416A (en) * | 2019-04-19 | 2019-06-21 | 天晴干细胞股份有限公司 | Protectant preparation method after a kind of umbilical cord blood hematopoietic stem cell recovery for freezing |
| CN110250165A (en) * | 2019-07-24 | 2019-09-20 | 安徽科门生物科技有限公司 | A kind of umbilical cord mesenchymal stem cells frozen stock solution and cryopreservation methods |
| CN110771599A (en) * | 2019-11-25 | 2020-02-11 | 深圳科学之门生物工程有限公司 | Stem cell refrigerating fluid and refrigerating method thereof |
| CN113331176A (en) * | 2021-06-01 | 2021-09-03 | 北京戴域生物技术有限公司 | Mesenchymal stem cell cryopreservation liquid |
| WO2022099472A1 (en) * | 2020-11-10 | 2022-05-19 | 南通市巨久新材料科技有限公司 | Cryopreservation tube containing aloe-polysaccharide-coated layer for amniotic-fluid-derived mesenchymal stem cells |
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Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109913416A (en) * | 2019-04-19 | 2019-06-21 | 天晴干细胞股份有限公司 | Protectant preparation method after a kind of umbilical cord blood hematopoietic stem cell recovery for freezing |
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| CN110250165A (en) * | 2019-07-24 | 2019-09-20 | 安徽科门生物科技有限公司 | A kind of umbilical cord mesenchymal stem cells frozen stock solution and cryopreservation methods |
| CN110771599A (en) * | 2019-11-25 | 2020-02-11 | 深圳科学之门生物工程有限公司 | Stem cell refrigerating fluid and refrigerating method thereof |
| WO2022099472A1 (en) * | 2020-11-10 | 2022-05-19 | 南通市巨久新材料科技有限公司 | Cryopreservation tube containing aloe-polysaccharide-coated layer for amniotic-fluid-derived mesenchymal stem cells |
| CN113331176A (en) * | 2021-06-01 | 2021-09-03 | 北京戴域生物技术有限公司 | Mesenchymal stem cell cryopreservation liquid |
| CN113331176B (en) * | 2021-06-01 | 2022-04-29 | 零下十八度(北京)生物科技有限公司 | Umbilical cord mesenchymal stem cell cryopreservation liquid |
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