CN110747164A - Preparation method of dental pulp stem cells - Google Patents
Preparation method of dental pulp stem cells Download PDFInfo
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- CN110747164A CN110747164A CN201911129932.5A CN201911129932A CN110747164A CN 110747164 A CN110747164 A CN 110747164A CN 201911129932 A CN201911129932 A CN 201911129932A CN 110747164 A CN110747164 A CN 110747164A
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- dental pulp
- stem cells
- physiological saline
- teeth
- pulp stem
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
- C12N5/06—Animal cells or tissues; Human cells or tissues
- C12N5/0602—Vertebrate cells
- C12N5/0652—Cells of skeletal and connective tissues; Mesenchyme
- C12N5/0662—Stem cells
- C12N5/0664—Dental pulp stem cells, Dental follicle stem cells
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2509/00—Methods for the dissociation of cells, e.g. specific use of enzymes
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- Organic Chemistry (AREA)
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Abstract
The invention discloses a preparation method of dental pulp stem cells. The preparation method comprises the following steps: 1) sterilizing; 2) taking pulp, adding physiological saline containing double antibody (1000UI \ ml streptomycin +1000UI \ ml penicillin + physiological saline), washing, and cutting into 1mm3Small blocks; 3) tissue digestion: the digestive juice is digested by 0.1-0.5% collagenase I + 0.1% Dispase II + normal saline in a sealed manner at the constant temperature of 37 +/-5 ℃ and under the oscillation of 167rpm of 145-; 4) centrifuging, discarding the supernatant, washing with physiological saline, centrifuging and discarding the supernatant; 5) culturing: the cells were cultured in mesenchymal stem cell medium (5% platelet lysate + 95% basal medium) at 37. + -. 5 ℃ in 5% carbon dioxide. The method can effectively obtain the single cell suspension of the dental pulp stem cells or soften the dental pulp tissue, is more beneficial to the climbing out of the dental pulp stem cells, improves the obtaining rate of the dental pulp stem cells, shortens the cell culture period and ensures the cell quality.
Description
Technical Field
The invention belongs to the technical field of stem cell culture in cell biology, and particularly relates to a preparation method of dental pulp stem cells.
Background
Stem Cells (SC) are a type of pluripotent Cells with the ability to self-replicate, being primitive and unspecified Cells. Under certain conditions, it can differentiate into multiple functional cells with the potential function of regenerating various tissues and organs and human body. The source of stem cells is many, including bone marrow, umbilical cord, cord blood, teeth, and fat.
Dental pulp stem cells were first discovered in 2000, and Gronthos et al found dental pulp stem cells in both isolated deciduous and wisdom teeth. This is a cell type with high proliferation capacity, high self-renewal capacity and multiple differentiation capacity. The cells have been studied to successfully construct dental pulp, dentin, and tooth tissue.
Dental Pulp Stem Cells (DPSCs) have many similarities in biological properties to other tissue-derived stem cells. Gronthos et al investigated the clonogenic rate of dental pulp stem cells and found, when compared to Bone Marrow Stromal Cells (BMSC): the clonal formation rate of dental-derived DPSCs was significantly higher than that of bone marrow-derived BMSCs, indicating that DPSCs have higher proliferative and self-renewal capacities than BMSCs. Miura et al found: every 12-20 cells from 1 shed anterior tooth form adherent clonal colonies, which are typical manifestations of mesenchymal stem cell proliferation.
Dental pulp stem cells belong to multipotent adult stem cells and have the potential to differentiate into nerve cells, osteoblasts, adipocytes and chondroblasts. Research shows that the dental pulp stem cells can be used for repairing and regenerating dentin, dental pulp and tooth tissues, can also be used for repairing the damage of skull, jaw bone and facial bone, and has a great deal of application research on the aspect of other diseases. The method has the advantages of rich sources, safe and convenient material taking, no ethical problem, low immunogenicity and the like, makes great progress in the field of regenerative medicine, and has wide application prospect.
At present, for the extraction and culture processes of dental pulp tissues, the traditional methods are a tissue block direct culture method and a collagenase digestion culture method, but the traditional methods have the defects of overlong culture period, less cell amount, easy differentiation, incomplete digestion or excessive digestion and the like.
Because of the wide variety and large quantity of bacteria and fungi in the oral cavity of a human body, dental pulp stem cells are very easy to be polluted in the preparation process, and the probability of pollution is usually reduced by increasing the using amount of antibiotics or increasing the grade of the antibiotics, so that the abuse of the antibiotics is caused.
Therefore, prevention of abuse of antibiotics, improvement of efficiency of separation of dental pulp stem cells, and shortening of growth cycle in the process of preparation of dental pulp stem cells are problems to be solved in the art.
Disclosure of Invention
In order to overcome the defects of the prior art, the invention aims to provide a preparation method of dental pulp stem cells, which improves the yield of the dental pulp stem cells, shortens the cell culture period and ensures the cell quality.
In order to achieve the purpose, the invention adopts the following technical scheme:
the invention provides a preparation method of dental pulp stem cells, which comprises the following steps:
1) sterilizing;
2) taking pulp, adding physiological saline containing double antibody (1000UI \ ml streptomycin +1000UI \ ml penicillin + physiological saline), washing, and cutting into 1mm3Small blocks;
3) tissue digestion: digesting the digestive juice by using 0.1-0.5% collagenase I and 0.1-0.5% Dispase II and normal saline, sealing the mouth of the digestive tube, keeping the temperature at 37 +/-5 ℃, and carrying out oscillatory digestion at 167rpm of 145-;
4) centrifuging, discarding the supernatant, washing with physiological saline, centrifuging and discarding the supernatant;
5) culturing: the cells were cultured in mesenchymal stem cell medium (5% platelet lysate + 95% basal medium) at 37. + -. 5 ℃ in 5% carbon dioxide.
Further, in the step 1), the disinfection is an aseptic operation, and the teeth are soaked in 75% alcohol for 30 s-2 min and are completely immersed.
Further, in step 1), the sterilization further comprises washing the alcohol-soaked teeth 3-5 times with physiological saline containing antibiotics.
Further, the antibiotic is 1000UI \ ml streptomycin +1000UI \ ml penicillin.
Further, in the step 1), the disinfection also comprises a step of irradiating the clean workshop for 30min by using an ultraviolet lamp; also comprises a tooth storage bottle, a sterilized surgical instrument box, a 50ml centrifuge tube, 75 percent medical alcohol, normal saline and a culture dish which are sterilized by 75 percent medical alcohol and then are put into a biological safety cabinet.
Further, in the step 2), the dental pulp is taken out by wrapping the teeth with sterile dust-free cloth, clamping the teeth with tiger forceps and taking out the dental pulp with ophthalmologic forceps.
Further, in step 3), the final concentration of the components of the digestive juice is 0.1% collagenase I + 0.1% Dispase II + physiological saline.
Further, in the step 4), the centrifugation is 1000-1500 r/min for 5-8 min.
Further, in step 5), the culture is performed by using a mesenchymal stem cell culture medium (5% platelet lysate + 95% basal medium) for resuspension, uniformly inoculating the resuspended tissue blocks into a culture dish, and placing the culture dish into a 5% carbon dioxide incubator at 37 +/-5 ℃.
Further, in step 5), the composition of the platelet lysate is as follows:
further, in the step 5), a fluid supplementing step is also included, wherein the fluid is supplemented on the next day (5% platelet lysate and 95% basal medium), and the fluid is replaced half a day after every three days, and the passage is carried out until the cell fusion degree reaches 30% -60%.
Compared with the prior art, the method can effectively obtain the single cell suspension of the dental pulp stem cells or soften the dental pulp tissue, is more beneficial to the climbing out of the dental pulp stem cells, well improves the obtaining rate of the dental pulp stem cells, shortens the cell culture period and ensures the cell quality.
Drawings
FIG. 1 is a drawing showing the culture of primary dental pulp stem cells in example 1;
in the figure, A is the 3 rd day of primary dental pulp stem cell culture; b, culturing the primary dental pulp stem cells on the 8 th day; c is the 10 th day of primary dental pulp stem cell culture; d, culturing the primary dental pulp stem cells on the 15 th day;
FIG. 2 shows dental pulp stem cells cultured in situ for the third day in the experiment;
in the figure, a is the dental pulp stem cell prepared by the control method; b is the dental pulp stem cell prepared by the method.
Detailed Description
The mesenchymal stem cell culture medium used in the present invention comprises 5% platelet lysate + 95% basal medium.
Among them, the components of platelet lysate are shown in table 1 below.
TABLE 1 platelet lysate Components
| Item | Concentration of |
| Globulin Globulin | 1.8g/dL |
| Albumin | 3.3g/dL |
| Cholesterol | 128mg/dL |
| Bilirubin | 0.2mg/dL |
| Glucose | 208mg/dL |
| Triglyceride trigyceride | 80mg/dL |
| Creatine phosphokinase CPK | 159U/L |
| Creatinine | 0.92mg/dL |
| γ-GT | 26U/L |
| Aspartate transferase AST | 43U/L |
| Bicarbonate salt Bicarbonate | 14.2meq/L |
| Sodium | 140meq/L |
| Phosphorus | 3.9mg/dL |
| Calcium | 61.5mg/dL |
| Magnesium | 2.1mg/dL |
| Potassium | 4.4meq/L |
| Chloride | 109meq/L |
| Iron Iron | 64ug/dL |
FoundationThe culture medium is produced by Shenzhen Dake as bioengineering LimitedThe basic medium dedicated to MSC (product 6115021) is a basic medium dedicated to culturing human Mesenchymal Stem cells (Mesenchymal Stem cells). The product has limited chemical components, does not contain heterogeneous animal components, does not contain serum, and needs to be used after being added with serum or serum substitutes to be mixed into a complete culture medium.
The components of the composition comprise: L-Alanine (L-Alanine), L-Arginine HCl (L-Arginine HCl), L-asparagine H2O(L-Asparagine H2O), L-aspartic acid (L-aspartic acid), L-cysteine HCl H2O(L-CysteineHCl H2O), L-Cystine (L-cysteine), L-Glutamic acid (L-Glutamic acid), L-Glutamine (L-Glutamine), Glycine (Glycine), L-histidine HCl H2O(L-Histidine HCl H2O), L-Isoleucine (L-Isoleucine), L-Leucine (L-Leucine), L-Lysine HCl (L-Lysine HCl), L-Methionine (L-Methionine), L-Phenylalanine (L-Phenylalanine), L-Proline (L-Proline), L-Serine (L-Serine), L-Threonine (L-Threonine), L-Tryptophan (L-Tryptophan), L-Tyrosine (L-Tyrosine), L-Valine (L-Valine), (+) -L-Sodium Ascorbate ((+) -Sodium L-ascorbic acid), Folic acid (Folic acid), Sodium chloride (Sodium chloride), Sodium dihydrogen phosphate (Sodium dihydrogen phosphate), Sodium hydrogen carbonate (Sodium hydrogen carbonate), D-Glucose (D-Glucose), Lipoic acid (Lipoic acid), sodium pyruvate (sodimu pyruvate).
The present invention will now be described in further detail with reference to the accompanying drawings and specific examples, but the present invention is not limited to the following examples.
Example 1
A method of preparing dental pulp stem cells, comprising the following apparatus and steps:
1. the instrument equipment comprises:
biological safety cabinet, carbon dioxide incubator, electronic pipettor, centrifuge, constant temperature shaking table.
2. Reagent consumables:
sterilized surgical instruments, 75% medical alcohol, normal saline, dispase enzyme, collagenase, 35mm petri dishes, pipettes (5ml, 10ml), culture medium, 50ml syringes, waste liquid tanks.
3. The method comprises the following operation steps:
1) and opening an ultraviolet lamp of the clean workshop to irradiate for more than 30min, and opening a clean area fan.
2) Receiving the tooth storage bottle into a mesenchymal stem cell preparation chamber, checking whether the storage bottle has a liquid leakage phenomenon, whether the storage bottle is tightly sealed, whether the storage bottle has a crack or not, whether the label is complete and accurate, and whether the tooth data is complete and meets the preparation requirements. Then wiping the bottle with alcohol gauze, and placing the storage bottle into a biological safety cabinet which normally runs.
3) Sterilizing surgical instrument box, 50ml centrifuge tube, 75% medical alcohol, normal saline, culture dish, etc. with 75% medical alcohol, and placing into biological safety cabinet.
4) The collection bottle is opened aseptically, the preservation solution is poured into a waste liquid jar, and the teeth are soaked in 75% alcohol for 2min (the teeth should be completely immersed).
5) The teeth were placed in a petri dish and the surface was washed 3-5 times with alcohol in physiological saline containing antibiotics. Wrapping teeth with sterile and dustless cloth, clamping with vise, taking out dental pulp with ophthalmologic forceps, washing with physiological saline containing double antibody (1000 UI/ml streptomycin +1000 UI/ml penicillin + physiological saline), transferring into new 35mm culture dish, and shearing into 1mm culture dish with ophthalmologic scissors3And (5) small blocks.
6) Transferring the cut tissue blocks to a 2ml centrifuge tube, adding 1ml digestive juice (0.1% collagenase I + 0.1% Dispase II + normal saline), sealing the tube opening, placing in a constant temperature shaking table at 37 ℃, and performing shake digestion at 145rpm for 15 min.
7) After digestion is finished, centrifuging at 1500r/min for 5min, discarding the supernatant, adding 1ml of physiological saline, blowing, washing, centrifuging at 1500r/min for 5min, discarding the supernatant, and adding 0.5ml of mesenchymal stem cell culture medium (5% platelet lysate + 95% basal medium) for resuspension. The resuspended tissue pieces were evenly plated onto 35mm petri dishes.
8) The culture dish was placed in a 5% carbon dioxide incubator at 37 ℃.
9) The next day, 0.5ml of fluid (5% platelet lysate + 95% basal medium) was replenished, half the fluid changes every three days later, and passages were performed until the degree of cell fusion reached 30% -60%.
The experimental results are as follows: as shown in fig. 1(a-D), it can be seen that the primary dental pulp stem cells obtained on days 3, 8, 10, and 15 have a large initial number of dental pulp stem cells, uniform cell plating, and good expansion state, thus greatly shortening the culture time of dental pulp stem cells and better maintaining the dryness of dental pulp stem cells.
Comparative example 1
The preparation method of the control group comprises the following operation steps:
1) and opening an ultraviolet lamp of the clean workshop to irradiate for more than 30min, and opening a clean area fan.
2) Receiving the tooth storage bottle into a mesenchymal stem cell preparation chamber, checking whether the storage bottle has a liquid leakage phenomenon, whether the storage bottle is tightly sealed, whether the storage bottle has a crack or not, whether the label is complete and accurate, and whether the tooth data is complete and meets the preparation requirements. Then wiping the bottle with alcohol gauze, and placing the storage bottle into a biological safety cabinet which normally runs.
3) Sterilizing surgical instrument box, 50ml centrifuge tube, 75% medical alcohol, normal saline, culture dish, etc. with 75% medical alcohol, and placing into biological safety cabinet.
4) The collection bottle is opened aseptically, the preservation solution is poured into a waste liquid jar, and the teeth are soaked in 75% alcohol for 2min (the teeth should be completely immersed).
5) The teeth were placed in a petri dish and the surface was washed 3-5 times with alcohol in physiological saline containing antibiotics. Wrapping teeth with sterile and dustless cloth, clamping with vise, taking out dental pulp with ophthalmologic forceps, washing with physiological saline containing double antibody (1000 UI/ml streptomycin +1000 UI/ml penicillin + physiological saline), transferring into new 35mm culture dish, and shearing into 1mm culture dish with ophthalmologic scissors3And (5) small blocks.
6) Transferring the cut tissue blocks to a 2ml centrifuge tube, adding 1ml of digestive juice (0.1% collagenase I + normal saline), sealing the tube opening, and placing the tube opening into a constant temperature shaker at 37 ℃ and 145rpm for shaking digestion for 15 min.
7) After digestion is finished, centrifuging at 1500r/min for 5min, discarding the supernatant, adding 1ml of physiological saline, blowing, washing, centrifuging at 1500r/min for 5min, discarding the supernatant, and adding 0.5ml of mesenchymal stem cell culture medium (5% platelet lysate + 95% basal medium) for resuspension. The resuspended tissue pieces were evenly plated onto 35mm petri dishes.
8) The culture dish was placed in a 5% carbon dioxide incubator at 37 ℃.
9) The next day, 0.5ml of fluid (5% platelet lysate + 95% basal medium) was replenished, half the fluid changes every three days later, and passages were performed until the degree of cell fusion reached 30% -60%.
The experimental results are as follows: the third day culture conditions of the dental pulp stem cells prepared in comparative example 1 and example 1 of the method are shown, as shown in fig. 2a-2b, the dental pulp tissue digestion is more thorough, the stem cell yield is better, and the proliferation, the cell morphology and the phenotype of the cells are not influenced.
The present invention is not limited to the above-described embodiments, and various changes and modifications of the present invention are intended to be included within the scope of the claims and the equivalent technology of the present invention if they do not depart from the spirit and scope of the present invention.
Claims (10)
1. A method for preparing dental pulp stem cells is characterized by comprising the following steps:
1) sterilizing;
2) taking pulp, adding physiological saline containing double antibody (1000UI \ ml streptomycin +1000UI \ ml penicillin + physiological saline), washing, and cutting into 1mm3Small blocks;
3) tissue digestion: digesting the digestive juice by using 0.1-0.5% collagenase I and 0.1-0.5% Dispase II and normal saline, sealing the mouth of the digestive tube, keeping the temperature at 37 +/-5 ℃, and carrying out oscillatory digestion at 167rpm of 145-;
4) centrifuging, discarding the supernatant, washing with physiological saline, centrifuging and discarding the supernatant;
5) culturing: the cells were cultured in mesenchymal stem cell medium (5% platelet lysate + 95% basal medium) at 37. + -. 5 ℃ in 5% carbon dioxide.
2. The method of claim 1, wherein: in the step 1), the disinfection is aseptic operation, the teeth are soaked in 75% alcohol for 30 s-2 min, and the teeth are completely immersed.
3. The method of claim 2, wherein: in step 1), the disinfection further comprises washing the alcohol-soaked teeth 3-5 times with physiological saline containing antibiotics.
4. The method of claim 1, wherein: in the step 1), the disinfection also comprises a step of irradiating the clean workshop for 30min by using an ultraviolet lamp; also comprises a tooth storage bottle, a sterilized surgical instrument box, a 50ml centrifuge tube, 75 percent medical alcohol, normal saline and a culture dish which are sterilized by 75 percent medical alcohol and then are put into a biological safety cabinet.
5. The method of claim 1, wherein: in the step 2), the dental pulp is taken out by wrapping the teeth with sterile dust-free cloth, clamping the teeth with tiger forceps and taking out the dental pulp with ophthalmologic forceps.
6. The method of claim 1, wherein: in the step 3), the final concentration of the components of the digestive juice is 0.1% collagenase I + 0.1% Dispase II + normal saline.
7. The method of claim 1, wherein: in the step 4), the centrifugation is 1000-1500 r/min for 5-8 min.
8. The method of claim 1, wherein: in the step 5), the culture is performed by using a mesenchymal stem cell culture medium (5% platelet lysate and 95% basal medium) for heavy suspension, the heavy-suspended tissue blocks are uniformly inoculated on a culture dish, and the culture dish is placed in a 5% carbon dioxide incubator at the temperature of 37 +/-5 ℃ for culture.
9. The method of claim 1, wherein: in step 5), the composition of the platelet lysate is as follows:
10. the method of claim 1, wherein: and step 5), a fluid supplementing step is further included, fluid is supplemented the next day (5% of platelet lysate and 95% of basal medium), the fluid is replaced half a day every three days later, and passage is carried out when the cell fusion degree reaches 30% -60%.
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112522188A (en) * | 2020-11-25 | 2021-03-19 | 广东圆康再生医学科技开发有限公司 | Extraction of dental pulp stem cells |
| CN114642683A (en) * | 2022-02-25 | 2022-06-21 | 宁波一棵芽生物科技有限公司 | Preparation method of dental pulp mesenchymal stem cell lysate with photoaging resistance |
| CN115521906A (en) * | 2021-06-24 | 2022-12-27 | 东莞宣冠干细胞再生医学有限公司 | A method for preparing high-quality human dental pulp stem cells |
| CN116240167A (en) * | 2022-12-30 | 2023-06-09 | 海南优尼科尔生物科技有限公司 | A preparation method for extracting dental pulp stem cells |
Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103387959A (en) * | 2012-10-19 | 2013-11-13 | 夏佑红 | Stem cell from deciduous teeth, and preparation method and application thereof |
| CN104560872A (en) * | 2014-12-29 | 2015-04-29 | 深圳市北科生物科技有限公司 | In-vitro separation and cultivation method for tooth-sourced mesenchymal stem cells |
| CN104694464A (en) * | 2015-02-09 | 2015-06-10 | 安徽新生命干细胞科技有限公司 | Clinical dental pulp stem cell and preparation method thereof |
| CN105907711A (en) * | 2016-06-27 | 2016-08-31 | 安徽新生命干细胞科技有限公司 | Preparation method of deciduous tooth mesenchymal stem cells and used kit |
| CN107663514A (en) * | 2017-06-29 | 2018-02-06 | 刘劼 | A kind of serum free medium of human umbilical cord mesenchymal stem cells |
| CN108949682A (en) * | 2018-08-22 | 2018-12-07 | 广东唯泰生物科技有限公司 | A kind of preparation, culture and the purification process of dental pulp mescenchymal stem cell |
| CN109321522A (en) * | 2018-11-20 | 2019-02-12 | 潍坊市康华生物技术有限公司 | A method of preparing dental pulp stem cell |
-
2019
- 2019-11-18 CN CN201911129932.5A patent/CN110747164A/en active Pending
Patent Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103387959A (en) * | 2012-10-19 | 2013-11-13 | 夏佑红 | Stem cell from deciduous teeth, and preparation method and application thereof |
| CN104560872A (en) * | 2014-12-29 | 2015-04-29 | 深圳市北科生物科技有限公司 | In-vitro separation and cultivation method for tooth-sourced mesenchymal stem cells |
| CN104694464A (en) * | 2015-02-09 | 2015-06-10 | 安徽新生命干细胞科技有限公司 | Clinical dental pulp stem cell and preparation method thereof |
| CN105907711A (en) * | 2016-06-27 | 2016-08-31 | 安徽新生命干细胞科技有限公司 | Preparation method of deciduous tooth mesenchymal stem cells and used kit |
| CN107663514A (en) * | 2017-06-29 | 2018-02-06 | 刘劼 | A kind of serum free medium of human umbilical cord mesenchymal stem cells |
| CN108949682A (en) * | 2018-08-22 | 2018-12-07 | 广东唯泰生物科技有限公司 | A kind of preparation, culture and the purification process of dental pulp mescenchymal stem cell |
| CN109321522A (en) * | 2018-11-20 | 2019-02-12 | 潍坊市康华生物技术有限公司 | A method of preparing dental pulp stem cell |
Non-Patent Citations (1)
| Title |
|---|
| 侯宗柳等: "《围产期成体干细胞基础与临床应用》", 31 August 2016 * |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112522188A (en) * | 2020-11-25 | 2021-03-19 | 广东圆康再生医学科技开发有限公司 | Extraction of dental pulp stem cells |
| CN115521906A (en) * | 2021-06-24 | 2022-12-27 | 东莞宣冠干细胞再生医学有限公司 | A method for preparing high-quality human dental pulp stem cells |
| CN114642683A (en) * | 2022-02-25 | 2022-06-21 | 宁波一棵芽生物科技有限公司 | Preparation method of dental pulp mesenchymal stem cell lysate with photoaging resistance |
| CN114642683B (en) * | 2022-02-25 | 2023-10-10 | 宁波一棵芽生物科技有限公司 | Preparation method of dental pulp mesenchymal stem cell lysate with photoaging resistance |
| CN116240167A (en) * | 2022-12-30 | 2023-06-09 | 海南优尼科尔生物科技有限公司 | A preparation method for extracting dental pulp stem cells |
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