Serum-free tumor stem cell culture medium and application thereof

A tumor stem cell, serum-free technology, applied in the field of cell biology research, can solve the problem of time-consuming and high cost, and achieve the effects of high safety, good cell activity and physiological characteristics, and stable growth

Active Publication Date: 2017-11-21
上海揽微赛尔生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The above method for separating and enriching tumor stem cells requires expensive instruments and equipment, which is time-consuming and expensive

Method used

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  • Serum-free tumor stem cell culture medium and application thereof
  • Serum-free tumor stem cell culture medium and application thereof
  • Serum-free tumor stem cell culture medium and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0027] This example is the preparation method of compound 4(15)-cineole-5,7-dihydroxy-1-one

[0028] The preparation method of compound 4(15)-cineole-5,7-dihydroxy-1-one comprises the following steps:

[0029] 1) Fermentation culture of the mangrove fungus Curvularia sp.3e-GC-1CGMCC NO: KY048419: use solid rice medium for fermentation, the medium is rice: seawater = 1:1 (3% sea salt) ;Pick the cultured strains in the slant into the fermentation medium, and culture them statically at room temperature 28°C for 30 days;

[0030] 2) Extract the above-mentioned cultivated cells with chloroform / methanol 1:1 for 3 times, concentrate to obtain the extract, use ethyl acetate: water to distribute and extract the extract for 3 times, concentrate the ethyl acetate to obtain the organic phase of the cells Extract, wherein the mass ratio of ethyl acetate to water is 1:1;

[0031] 3) With petroleum ether-ethyl acetate-methanol as the elution system, the extract is roughly separated by sili...

Embodiment 2

[0037] This example is for the preparation of tumor stem cell culture medium.

[0038] The MCDB153 medium is used as the basal medium, and its composition is as follows:

[0039]

[0040]

[0041] Dissolve the above components in pyrogen-free ultrapure water and add 2 mg / L ethanolamine, 4.7 μg / L oleic acid, 4.7 μg / L linoleic acid, 5 μg / L selenium, 1.2 g / L sodium bicarbonate, and use 1N hydrochloric acid (HCl) or 1N sodium hydroxide (NaOH) to adjust the pH value to 7.4, then add 0.5mg / L albumin, 0.125μg / L epidermal growth factor, 0.33mg / L insulin-like growth factor 1, 0.33mg / L bovine pituitary extract, 10mg / L insulin, 5.5mg / L transferrin, 0.33mg / L hydrocortisone, 0.39mg / L epinephrine, 4(15)-cineole-5, 3.6mg / L of 7-dihydroxy-1-ketone, 100u / L of penicillin and 100u / L of streptomycin, the culture solution is filtered and sterilized, which is the serum-free tumor stem cell liquid medium of the present invention. The basal medium of the present invention contains nutrients r...

Embodiment 3

[0043] In this example, the tumor stem cell culture medium prepared in Example 2 was used to culture the prostate cancer Du145 cell line.

[0044] Prostate cancer stem cells can be enriched by culturing the prostate cancer Du145 cell line with the tumor stem cell culture medium of the present invention. In the experiment of this example, the prostate cancer Du145 cells were first digested into single cells, and 1×10 3 The density of cells / ml inoculated the cells into CorningTM ultra-low adherent 24-well cell culture plates, cultured them in the tumor stem cell culture medium of the present invention, and observed the cells under the microscope on the 1st, 3rd, 6th and 9th day of culture respectively The growth conditions of the obtained results are as follows figure 1 shown.

[0045] Depend on figure 1 It can be seen that the prostate cancer stem cells that form tumor microspheres (Tumor Spheroid) can be enriched by Du145 cells cultured in the stem cell medium of the presen...

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Abstract

The invention discloses a serum-free tumor stem cell culture medium. The serum-free tumor stem cell culture medium is prepared from an MCDB153 basic culture medium, cell factors, protein polypeptides, antibiotics, epinephrine, selenium, sodium hydrogen carbonate, lipids and mangrove endophytic fungus metabolite 4(15)-eudesmene-5,7-dyhydroxyl-1-ketone. The tumor stem cell culture medium contains no serum, the formula compositions are definite, the safety is relatively high, the serum-free tumor stem cell culture medium can efficiently concentrate tumor stem cells from a malignant tumor cell line and tumor tissues, the tumor stem cells are enabled to grow stably, good cell activity and physiological property can be maintained, and the serum-free tumor stem cell culture medium is very applicable to the field of relevant researches for tumor cells and the tumor stem cells.

Description

technical field [0001] The present invention belongs to the field of cell biology research. Specifically, the present invention relates to a serum-free tumor stem cell culture medium and its application. More specifically, the present invention relates to a method of adding mangrove endophytic fungus metabolite 4(15)-eucalyptus Folene-5,7-dihydroxy-1-one serum-free medium for tumor stem cells and its application for culturing tumor cell lines. Background technique [0002] Stem cells are a kind of slow-cycle cells with self-renewal, multi-directional differentiation potential, and unlimited proliferation and division. The related research plays an increasingly important role in biological development, gene therapy, drug screening and organ transplantation. In recent years, with the deepening of the research on the physiological characteristics of stem cells, researchers have found that the progressive growth, recurrence and metastasis of malignant tumors are very similar to ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/095
CPCC12N5/0695C12N2500/12C12N2500/25C12N2500/30C12N2500/36C12N2500/46C12N2500/84C12N2500/90C12N2501/105C12N2501/11C12N2501/39C12N2501/81C12N2501/998
Inventor 郑希黄华容刘婷李冬利张焜周仁平顾为望
Owner 上海揽微赛尔生物科技有限公司
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