A kind of human vascular endothelial cell culture medium and culture method thereof
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A vascular endothelium and cell culture technology, which is applied in the directions of vascular endothelial cells, cell dissociation methods, cell culture active agents, etc. high effect
Active Publication Date: 2019-11-05
上海瑞鹿生物技术有限公司
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At present, the relatively mature endothelial cell culture media on the market mainly include EGM and EGM-2 series of Lonza Company, and ECM culture medium of Sciencell. effect, but the formula is secret and expensive
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Embodiment 1
[0063] The present embodiment adopts the culture solution of the present invention, the M199 complete medium containing 10% fetal bovine serum and the ECM serum-containing medium of sciencecell to carry out comparative experiments. The three culture solutions are respectively:
[0064] Culture solution (improved culture medium) of the present invention:
[0065] On the basis of every 100ml DMEM / F12 medium, add the following components: 10ml fetal bovine serum, 100U / ml penicillin, 100mg / ml streptomycin, 1.25ml L-glutamine, 90μg / ml heparin, 5ng / ml ml human recombinant epidermal growth factor, 10ng / ml human recombinant fibroblast growth factor, 0.5ng / ml vascular endothelial growth factor, 20ng / ml insulin-like growth factor, 1μg / ml vitamin C, 1μg / ml cortisol. 100 ng / ml cholera toxin, 10 μM aminoethanol, 100 pM phosphoethanolamine.
[0066] Control medium A: M199 complete medium containing 10% fetal bovine serum
[0067] Control Medium B: Sciencecell’s ECM Serum-Containing Medium ...
Embodiment 2
[0076] Recovery, subculture, cryopreservation and rethrowing of human microvascular endothelial cells HMEC-1. See Example 1 for details of the three cell culture media used.
[0077] First, put the improved culture medium of the present invention, control culture medium A and control culture medium B three kinds of culture medium into 3 petri dishes according to the medium consumption of 1ml / 5cm, put into 37 ± 1 ℃, 5% CO 2 , Preheat in an incubator with a humidity of 90±2% for 30 minutes.
[0078] Then three cryopreservation tubes of the same batch of frozen HMEC-1 cells were taken out from the liquid nitrogen, and quickly put into a 37°C water bath for thawing. After the cryopreservation solution is completely thawed, transfer 1ml of the frozen cells in the cryopreservation tube to a centrifuge tube containing 9ml of serum-free medium, centrifuge at 800rpm / min for 5min, and resuspend the cell pellet with 1ml of the corresponding complete medium Then add the cell suspension ...
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Abstract
The invention provides a human vascular endothelial cell culture solution. The human vascular endothelial cell culture solution is prepared from a basic culture medium, fetal calf serum, human recombined epidermal growth factors, human recombined fibroblast growth factors, vascular endothelial growth factors, insulin-like growth factors, vitamin C, cortisol, cholera toxin, aminoethanol and ethanolamine phosphate. Compared with a traditional vascular endothelial cell culture medium, the human vascular endothelial cell culture solution has the advantages that the culture solution can be suitable for multiple kinds of vascular endothelial cells, cell proliferation is effectively promoted, the division time is effectively shortened, the characteristics of the vascular endothelial cells are maintained, the passage number of the in-vitro human vascular endothelial cells is greatly increased, and the survival time of the in-vitro human vascular endothelial cells is greatly prolonged; meanwhile, all the ingredients of the cell culture solution can be obtained in a general laboratory, preparation is convenient, a special finished product culture medium is not needed, therefore, the cost for culturing the endothelial cells is greatly reduced, and the human vascular endothelial cell culture solution is suitable for application and popularization.
Description
technical field [0001] The invention relates to the technical field of cell culture, in particular to a human vascular endothelial cell culture medium and a culture method thereof. Background technique [0002] Vascular endothelial cells not only participate in the mediation of vascular permeability and blood coagulation, but also play an important role in many processes such as immune mediation, transplant rejection, tumor metastasis, and inflammation. In scientific research, in vitro culture is often used to study vascular endothelial cells. Since most people still do not recognize the use of immortalized vascular endothelial cell lines, it is still routine to use purified vascular endothelial cells from humans for research. However, in the culture of vascular endothelial cells, the cell proliferation is often slow, and the properties of the cells change or die after several generations, which increases the research cost and prolongs the research time. [0003] The in vi...
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