Organoid culture medium and organoid culture method

A culture method and organoid technology, which can be applied to the medium for organoid culture and the field of organoid culture, can solve problems such as low clinical practicability, and achieve the effects of shortening the time required, reducing the cost requirement, and increasing the proliferation rate.

Inactive Publication Date: 2019-10-25
CAPITALBIO CORP
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] In order to solve the problem of low clinical practicability of organoids used in the evaluation of anti-tumor drugs in vitro, the presen

Method used

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  • Organoid culture medium and organoid culture method
  • Organoid culture medium and organoid culture method

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Embodiment 1

[0041] The medium for evaluating antitumor drugs of the present invention includes DMEM medium, pleural fluid supernatant with a volume concentration of 10%, enzymolysis casein with a mass percentage of 1%, and a concentration of 25 ng / kg in the DMEM medium. mL of Hydrocortisone, 2mM L-glutamine, 2μM Insulin, 70nM Transferrin, 0.1nM Cholera toxin, 10μM Y-27632, 0.5ng / mL of EGF, amphotericin B at a concentration of 2.5 μg / mL, penicillin at a concentration of 100 U / mL, and streptomycin at a concentration of 100 μg / mL.

[0042] Preparation of culture medium for antitumor drug evaluation: centrifuge the pleural effusion samples, take the supernatant, and filter through a 0.22 μm filter to obtain the pleural effusion supernatant. 1% Enzymatic Casein, Hydrocortisone at 25ng / mL, L-glutamine at 2mM, Insulin at 2μM, Transferrin at 70nM, Cholera toxin at 0.1nM, Concentration The ratio of Y-27632 at 10 μM, EGF at a concentration of 0.5 ng / mL, amphotericin B at a concentration of 2.5 μg...

Embodiment 2

[0050] The culture medium for antitumor drug evaluation of the present invention includes DMEM / F12 medium, pleural water supernatant with a volume concentration of 40% in DMEM / F12 medium, enzymatic hydrolyzed casein with a mass percentage of 5%, Hydrocortisone at 100 ng / mL, L-glutamine at 10 mM, Insulin at 5 μM, Transferrin at 200 nM, Cholera toxin at 1 nM, Y-27632 at 50 μM, EGF at 100 ng / mL, amphotericin B at 10 μg / mL, penicillin at 100 U / mL, and streptomycin at 100 μg / mL.

[0051] Preparation of culture medium for evaluation of antitumor drugs: centrifuge the pleural effusion samples, take the supernatant, and filter through a 0.22 μm filter to obtain the pleural effusion supernatant. 5% enzymatic casein, hydrocortisone at 100 ng / mL, L-glutamine at 10 mM, Insulin at 5 μM, transferrin at 200 nM, Cholera toxin at 1 nM, The ratios of 50 μM Y-27632, 100 ng / mL EGF, 10 μg / mL amphotericin B, 100 U / mL penicillin, and 100 μg / mL streptomycin were added to DMEM / F12 medium, mixed eve...

Embodiment 3

[0059] The medium for evaluating antitumor drugs of the present invention includes MEM medium, pleural fluid supernatant with a volume concentration of 5%, enzymolysis casein with a mass percentage of 0.5%, and a concentration of 1 ng / mL of hydrocortisone, L-glutamine at 1 mM, Insulin at 1 μM, transferrin at 1 nM, Cholera toxin at 0.1 nM, Y-27632 at 1 μM, 0.5 ng / mL of EGF, amphotericin B at a concentration of 1 μg / mL, penicillin at a concentration of 100 U / mL, and streptomycin at a concentration of 100 μg / mL.

[0060] Preparation of culture medium for evaluation of antitumor drugs: centrifuge the pleural effusion samples, take the supernatant, and filter through a 0.22 μm filter to obtain the pleural effusion supernatant. 0.5% Enzymatic Casein, Hydrocortisone at 1 ng / mL, L-glutamine at 1 mM, Insulin at 1 μM, Transferrin at 1 nM, Cholera toxin at 0.1 nM, Concentration The above components were added in the ratio of 1 μM Y-27632, 0.5 ng / mL EGF, 1 μg / mL amphotericin B, 100 U / m...

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Abstract

The invention relates to the field of biotechnology, in particular to an organoid culture medium and an organoid culture method. The organoid culture medium comprises a base medium, hydrothorax supernatant, enzyme hydrolyzed casein, hydrocortisone, l-glutamic acid, insulin, transferrin, cholera viruses, Y-27632, EGF, amphotericin B, penicillin and streptomycin. The organoid culture method includesperforming two-dimensional culture on tumor cells, and performing three-dimensional culture on the cultured cells to form organoid tissue. The medium formula can promote the rapid proliferation of tumor tissue while avoiding hypoxia injuries, and the culture method can greatly shorten the time required for a subsequent drug sensitivity test; the accuracy of the drug sensitivity test is improved,and the test cost is reduced.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a culture medium for organoid culture and an organoid culture method. Background technique [0002] At present, there are two main methods for in vitro evaluation of antitumor drugs. One is to establish tumor xenograft models (patient-derived xenograft tumor models) by xenografting patient-derived tumor tissues or cells into immunodeficient mice. model, PDX model), and then conduct drug animal experiments. The advantages of this method are that the tissue type and genetic characteristics of tumor patients are preserved, and the drug sensitivity test results are highly consistent with the patient's response. However, the disadvantage is that the model establishment cycle requires several month, the success rate is about 30%, the cost is high, and the clinical practicability is not high. Another method is to culture patient tissue in vitro. This method does not require animal experime...

Claims

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Application Information

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IPC IPC(8): C12N5/09C12N5/078C12N5/071C12Q1/02
CPCC12N5/0697C12N2500/02C12N2500/25C12N2500/32C12N2500/84C12N2501/01C12N2501/11C12N2501/39C12N2501/727C12N2501/998C12N2502/11C12N2502/1323C12N2503/02C12N2513/00G01N33/5011G01N2500/10
Inventor 郭弘妍张怡然隋锡朝王俊邢婉丽程京
Owner CAPITALBIO CORP
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