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An immune cell culture medium, culture method and application

An immune cell and cell culture technology, applied in the field of cell culture, can solve the problems of inability to stably expand and cultivate CD8-positive NK cells, and achieve the effects of promoting differentiation and maturation, reducing death, and enhancing killing effect.

Active Publication Date: 2021-03-23
国家卫生健康委科学技术研究所
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0006] Therefore, the technical problem to be solved by the present invention is that in the prior art, a large amount of CD8-positive NK cells cannot be stably expanded and cultivated in vitro, and a large amount of CD8-positive NK cells can be stably expanded and cultivated in vitro. Immune cell culture medium, culture method and use of

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  • An immune cell culture medium, culture method and application
  • An immune cell culture medium, culture method and application
  • An immune cell culture medium, culture method and application

Examples

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Embodiment 1

[0039] This embodiment provides an immune cell culture medium and a culture method, culturing CD8+ NK cells from 8 cases of healthy human PBMC:

[0040] 1. Preparation of CD8+ NK cell culture stimulation solution and medium:

[0041] The formula of cell culture stimulation solution (reagent A) is: serum-free medium X-VIVO 10 (Lonza), recombinant human interleukin 2 (IL2), recombinant human interleukin 15 (IL15) and recombinant human interleukin 21 (IL21); The concentration of interleukin 2 (IL2) was 500 U / mL, the concentration of recombinant human interleukin 15 (IL15) was 20 ng / mL, and the concentration of recombinant human interleukin 21 (IL21) was 5 ng / mL. The preparation method is as follows: take the above-mentioned cytokines and use serum-free medium X-VIVO 10 (Lonza) to prepare CD8+ NK cell in vitro culture stimulation solution with corresponding concentration to obtain recombinant human interleukin 2 (IL2) at a concentration of 500 U / mL, Serum-free medium X-VIVO10 wit...

Embodiment 2

[0052] In the immune cell culture medium and culture method provided in this example, CD8+ NK cells were cultured from 8 cases of healthy human PBMC.

[0053] 1. Preparation of CD8+ NK cell culture stimulation solution and medium:

[0054] The formula of cell culture stimulation solution (reagent A) is: serum-free medium X-VIVO 10 (Lonza), recombinant human interleukin 2 (IL2), recombinant human interleukin 15 (IL15) and recombinant human interleukin 21 (IL21); The concentration of interleukin 2 (IL2) was 500 U / mL, the concentration of recombinant human interleukin 15 (IL15) was 20 ng / mL, and the concentration of recombinant human interleukin 21 (IL21) was 5 ng / mL. The preparation method is as follows: take the above-mentioned cytokines and use serum-free medium X-VIVO 10 (Lonza) to prepare CD8+ NK cell in vitro culture stimulation solution with corresponding concentration to obtain recombinant human interleukin 2 (IL2) at a concentration of 500 U / mL, Serum-free medium X-VIVO...

Embodiment 3

[0066] In the immune cell culture medium and culture method provided in this embodiment, CD8+ NK cells are cultured from human umbilical cord blood mononuclear cells, and the specific steps are as follows:

[0067] 1. Preparation of CD8+ NK cell culture stimulation solution and medium:

[0068] The formula of cell culture stimulation solution (reagent A) is: serum-free medium X-VIVO 10 (Lonza), recombinant human interleukin 2 (IL2), recombinant human interleukin 15 (IL15) and recombinant human interleukin 21 (IL21); The concentration of interleukin 2 (IL2) was 500 U / mL, the concentration of recombinant human interleukin 15 (IL15) was 20 ng / mL, and the concentration of recombinant human interleukin 21 (IL21) was 5 ng / mL. The preparation method is as follows: take the above-mentioned cytokines and use serum-free medium X-VIVO 10 (Lonza) to prepare CD8+ NK cell in vitro culture stimulation solution with corresponding concentration to obtain recombinant human interleukin 2 (IL2) a...

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Abstract

The invention discloses an immune cell culture medium, a culture method and application. The immune cell culture medium comprises a base cell culture medium, cell factors and glucocorticoid; the immune cell culture method comprises the following steps: S1, collecting and activating immune cells to be cultured; S2, inoculating the activated immune cells into the immune cell culture medium for culturing the cells; S3, harvesting the immune cells. A cell culture stimulating liquid comprises the base cell culture medium and the cell factors; the cell factors comprise interleukin 2, interleukin 15and / or interleukin 21. According to the immune cell culture medium and the culture method provided by the invention, the proportion of CD8+NK cells in the cultured NK cells can be effectively increased.

Description

technical field [0001] The invention belongs to the field of cell culture, and in particular relates to an immune cell culture medium, a culture method and application. Background technique [0002] Natural killer cells (natural killer cells, NK) are very important innate immune cells in the body's immune cells. the first barrier. NK cells were discovered in the 1970s. With the continuous deepening of research on NK cells in the past 40 years, NK cells have been identified as a group of heterogeneous cells with cell surface markers CD56+CD16+CD3-. Studies have shown that compared with T cells and B cells, the specificity of NK cell surface markers is relative. Human NK cells mIg-, some NK cells are CD2, CD3 and CD8 positive, express IL-2 receptor β chain (P75, CD122), CD11b / CD18 positive. Commonly used surface markers for detecting NK cells include CD16, CD56, CD57, CD59, CD11b, CD94 and LAK-1. NK cells can be divided into many cell subgroups according to their cell surf...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/0783C12N5/0784
CPCC12N5/0636C12N5/0638C12N5/0639C12N5/0646C12N2500/38C12N2501/2302C12N2501/2315C12N2501/2321C12N2501/30
Inventor 马旭高建恩郭昌龙
Owner 国家卫生健康委科学技术研究所
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