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Medium addition kit for culturing nk cells and nk cell culturing method using kit

A technology of NK cells and culturing method, which is applied in the field of medium addition kits for NK cell culture, can solve the problem of difficulty in obtaining NK cells, etc., and achieve the effect of easy culture

Active Publication Date: 2018-10-23
申东赫 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] In particular, immature progenitor cells will mature into T cells if CD3 antibody is strongly stimulated from the beginning, so the conventional method that is usually carried out is to use the method of slightly stimulating CD3 antibody at the beginning, but it depends on the surrounding environment such as individual differences essential, so it is difficult to obtain activated and proliferate NK cells

Method used

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  • Medium addition kit for culturing nk cells and nk cell culturing method using kit
  • Medium addition kit for culturing nk cells and nk cell culturing method using kit
  • Medium addition kit for culturing nk cells and nk cell culturing method using kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0071] Prepare the medium addition kit for NK cell culture as follows.

[0072] 1. Unit B preparation steps

[0073] Add IL-2 2.2mg and 500mM L-glutamine (L-glutamine) solution 100mL into the basic medium for suspension cell culture (if there is already IL-2 or L-glutamine in the basic medium, then Adjust the addition amount to adjust the final concentration) and dissolve to finally make 10 L to prepare a base solution (B solution). Using 3.8 L of this solution, 36 mL was put into each container, the stopper was closed, and the label "B solution (solution), 7 days (day)" prepared in advance was affixed to prepare Unit B, which was refrigerated and stored. The remaining 6.2 L was used to prepare other units.

[0074] 2. C1 unit preparation steps

[0075] A cytokine solution (solution C) was prepared by dissolving 20 ug of IL-12 and 125 ug of IL-18 in distilled water to make 10 mL. 1 mL of solution C was dissolved in 500 mL of the base solution (solution B) to prepare a firs...

Embodiment 2

[0089] After preparing lymphocytes and autologous plasma from the patient's blood, NK cells were cultured using the medium addition kit for NK cell culture obtained in Example 1 as follows.

[0090] 1. Lymphocyte extraction and self-plasma preparation steps

[0091] Utilizing the property that mononuclear cells such as human lymphocytes or monocytes have a specific gravity lower than 1.077, the blood of the patient to be treated is overlapped in a lymphocyte separation solution (Ficoll-Paque Plus) with a specific gravity of 1.077 and made it with a certain centrifugal force. According to the difference in specific gravity, the layer of red blood cells and granulocytes with a specific gravity greater than 1.077 is downward, while the layer of monocytes and platelets with a specific gravity below 1.077 is upward and separated to obtain a mononuclear cell layer containing lymphocytes. From the separated Only lymphocytes were extracted from the monocyte layer.

[0092] More speci...

Embodiment 3

[0112] Except that the blood used was not from patient A but from patient B, the same method as in Example 2 was performed, and the number of immune cells obtained after culturing for 8 days was 17.5×10 7 , the number of cells obtained by culturing the lymphocytes isolated from 30mL of patient B's blood was 4.33×10 9 indivual.

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Abstract

The present invention relates to a method for culturing natural killer cells (NK cells) applied to immune therapy and, more particularly, to a medium addition kit for culturing NK cells with which lymphocytes derived from human peripheral blood can be cultured to effectively amplify and activate NK cells, which have an outstanding therapeutic effect on malignant tumors, while remarkably increasingthe share of NK cells, and an NK cell culturing method using the kit.

Description

technical field [0001] The present invention relates to a natural killer cell (Natural Killer cell; NK cell) culture method suitable for immunotherapy, and more specifically, to a medium addition kit for NK cell culture and NK cells using the kit The culture method, by culturing lymphocytes from human peripheral blood, can efficiently expand and activate NK cells with excellent effects in the treatment of malignant tumors, and at the same time, the proportion of NK cells can be significantly increased. Background technique [0002] Immunotherapy, which uses various stimulators after extracting natural killer cells (NK cells: Natural Killer cells), dendritic cells (DC), B cells, and T cells, which are the most important immune cells in cancer treatment, from the patient's blood , the method of cultivating immune cells that have a strong effect on cancer and then injecting them into the patient again, because the patient's own blood is used, compared with the existing chemothe...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/0783C07K16/28
CPCC12N2500/38C12N2501/515C12N2501/599C07K16/2803C07K16/2809A61P35/00A61K35/17C07K16/283C12N5/0646C12N2501/2302C12N2501/2318
Inventor 申东赫
Owner 申东赫
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