Culture solution and differentiation method for differentiating pluripotent stem cells into natural killer cells

A technology of natural killer cells and pluripotent stem cells, applied in the field of stem cell therapy, can solve the problems of long differentiation cycle, weak cell killing power, and complicated operation, and achieve the effects of low labor cost, easy industrial automation, and simple operation.

Active Publication Date: 2021-04-16
SHANGHAI AISAER BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0018] Aiming at the technical problems existing in the existing NK cell differentiation technology, such as containing animal-derived components, cumbersome operation, long differentiation cycle, low yield of NK cells, immaturity, and weak

Method used

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  • Culture solution and differentiation method for differentiating pluripotent stem cells into natural killer cells
  • Culture solution and differentiation method for differentiating pluripotent stem cells into natural killer cells
  • Culture solution and differentiation method for differentiating pluripotent stem cells into natural killer cells

Examples

Experimental program
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Effect test

Embodiment 1～5

[0049] Examples 1-5 Utilize pluripotent stem cells to differentiate and cultivate NK cells

[0050] 1. Culture medium

[0051] The culture solutions of Examples 1-5 include a basal medium formed by mixing DMEM / F12 medium and blood cell culture medium stempro-34, and additional components added to the basal medium. Table 1 shows the mixing volume ratio, added components and contents of DMEM / F12 medium and blood cell culture medium stempro-34 of each embodiment.

[0052] Table 1 The composition and content of the culture medium

[0053]

[0054]

[0055] Remarks: All substances mentioned in Table 1 are purchased from the market

[0056] Among them, the trace element A is from the manufacturer CORNING, the article number is 25-021-CI; the trace element B is from the manufacturer CORNING, the article number is 25-022-CI.

Embodiment 1

[0057] In the culture solution of Example 1, any induction factor in IGF-1, SR-1, VPA, and AS1842856 was not added; in the culture solution of Example 2, only IGF-1 was added; in the culture solution of Example 3, IGF-1 and SR-1, IGF-1, SR-1 and VPA were added to the culture solution of Example 4, and four induction factors of IGF-1, SR-1, VPA and AS1842856 were added to the culture solution of Example 5.

[0058] 2. Training process

[0059] Examples 1-5 respectively use the corresponding culture medium shown in Table 1 to differentiate and culture NK cells according to the following steps:

[0060] 1. Inoculate pluripotent stem cells into a matrigel-coated 6-well plate, and when the cell density reaches 10-50%, replace the NK differentiation culture medium (2mL / well) to induce cell differentiation (if the culture medium is stored in a refrigerator at 4°C, Need to take it out in advance and preheat at room temperature for 20 minutes), at 37°C, 5% CO 2 Adhesive differentiati...

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Abstract

The invention discloses a culture solution and a differentiation method for differentiating pluripotent stem cells into natural killer cells. The culture solution comprises a DMEM/F12 culture medium and a blood cell culture medium in a volume ratio of 1: (0.9-1.3), and further comprises an induction factor, L-ascorbic acid, L-glutamine, L-leucine, human serum albumin, transferrin, beta-mercaptoethanol, sodium selenite, ethanolamine, a trace element A and a trace element B. The culture solution provided by the invention can induce differentiation of the pluripotent stem cells towards the NK cells more efficiently, and the pluripotent stem cells can be directionally differentiated and cultured in a short time by using the method provided by the invention to obtain the NK cells. The NK cell differentiation method is free of animal-derived components, high in NK cell yield and simple in differentiation process, a differentiation period is shortened to 14 days, NK cell yield is greatly increased, the NK differentiation period is shortened, the NK differentiation process is simplified, and preparation cost is reduced.

Description

technical field [0001] The invention belongs to stem cell differentiation technology in the technical field of stem cell therapy, and in particular relates to a culture medium and a differentiation method for pluripotent stem cells to differentiate into natural killer cells. Background technique [0002] Natural killer cells (NK cells) are the main lymphocytes in the innate immune system and have anti-tumor and anti-viral effects. Clinical studies in recent years have shown that NK cell adoptive immunotherapy has curative effect on a variety of cancers and will not cause graft-versus-host disease, so it has broad application prospects. [0003] NK cell immunotherapy requires a large number of NK cells. At present, the main sources of NK cells are: ① NK cells isolated from autologous / allogeneic peripheral blood (PB-NK), ② NK cells isolated from autologous / allogeneic umbilical cord blood (UCB-NK) , ③ NK cells differentiated from embryonic stem cells / induced pluripotent stem c...

Claims

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Application Information

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IPC IPC(8): C12N5/0783
Inventor 高歌
Owner SHANGHAI AISAER BIOTECH CO LTD
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