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Culture solution and culture method of umbilical cord blood NKT cells

A technology of NKT cells and culture methods, which is applied in the direction of blood/immune system cells, animal cells, vertebrate cells, etc., can solve the problems of weak cell killing activity, low purity of NKT cell content, and low cell expansion rate, etc., to achieve Strong effect of killing tumor cells, conducive to rapid growth, and high rate of killing activity

Active Publication Date: 2022-07-12
GUANGDONG XIANKANGDA BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] At present, NKT cell culture is to treat isolated peripheral blood mononuclear cells (PBMC) with IFN-γ for 24 hours, then add CD3 monoclonal antibody, IL-1α and IL-2 factors to stimulate and induce a certain number of NKT cells, but the final NKT cells obtained have low purity of effective cell content, weak cell killing activity, and relatively low cell expansion rate

Method used

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  • Culture solution and culture method of umbilical cord blood NKT cells
  • Culture solution and culture method of umbilical cord blood NKT cells
  • Culture solution and culture method of umbilical cord blood NKT cells

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0093] 1), prepare the first medium, the second medium and the third medium

[0094] The first medium: add CD3 with a final concentration of 300ng / ml, CD137 with a final concentration of 200ng / ml, IL-2 with a final concentration of 2500IU / ml, and a final concentration of 40ng / ml in X-VIVO 15 basal medium IL-15 at a final concentration of 25ng / ml, SCF at a final concentration of 65ng / ml, FGF-2 at a final concentration of 700ug / ml and sodium ferulate at a final concentration of 50ng / ml;

[0095] Second medium: add IL-2 with a final concentration of 1600IU / ml, IL-18 with a final concentration of 20ng / ml, sodium ferulate with a final concentration of 55ng / ml and 550ug / ml in X-VIVO 15 basal medium ml of soybean lecithin;

[0096] The third medium: IL-2 with a final concentration of 1600 IU / ml, sodium ferulate with a final concentration of 55 ng / ml and soybean phospholipid with a final concentration of 550 ug / ml were added to the X-VIVO 15 basal medium.

[0097] 2) Inoculation of ...

Embodiment 2

[0103] 1), prepare the first medium, the second medium and the third medium

[0104] The first medium: KBM581 basal medium was supplemented with CD3 at a final concentration of 200ng / ml, CD137 at a final concentration of 300ng / ml, IL-2 at a final concentration of 3000IU / ml, and IL-2 at a final concentration of 20ng / ml. 15. IL-1β with a final concentration of 20ng / ml, SCF with a final concentration of 50ng / ml, FGF-2 with a final concentration of 600ug / ml and sodium ferulate with a final concentration of 30ng / ml;

[0105] The second medium: KBM581 basal medium was supplemented with IL-2 with a final concentration of 2000IU / ml, IL-18 with a final concentration of 10ng / ml, sodium ferulate with a final concentration of 30ng / ml and soybeans with a final concentration of 300ug / ml Phospholipids;

[0106] The third medium: IL-2 with a final concentration of 2000 IU / ml, sodium ferulate with a final concentration of 30 ng / ml and soybean phospholipid with a final concentration of 300 ug / ...

Embodiment 3

[0113] 1), prepare the first medium, the second medium and the third medium

[0114] The first medium: CD3 with a final concentration of 500ng / ml, CD137 with a final concentration of 100ng / ml, IL-2 with a final concentration of 2000IU / ml, and a final concentration of 50ng / ml were added to the AIM-V basal medium. IL-15, IL-1β with a final concentration of 50ng / ml, SCF with a final concentration of 80ng / ml, FGF-2 with a final concentration of 800ug / ml and sodium ferulate with a final concentration of 80ng / ml;

[0115] Second medium: Add IL-2 with a final concentration of 1000IU / ml, IL-18 with a final concentration of 30ng / ml, sodium ferulate with a final concentration of 80ng / ml and 800ug / ml to the AIM-V basal medium of soybean lecithin;

[0116] The third medium: IL-2 with a final concentration of 1000 IU / ml, sodium ferulate with a final concentration of 80 ng / ml and soybean phospholipid with a final concentration of 800 ug / ml were added to the AIM-V basal medium.

[0117] 2)...

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Abstract

The invention discloses a culture solution and a culture method of umbilical cord blood NKT cells. The culture solution comprises the following components: a first culture medium, a second culture medium and a third culture medium, wherein the first culture medium is formed by adding cytokines CD3, CD137, IL-2, IL-15, IL-1beta, SCF, FGF-2 and sodium ferulate into a basic culture medium; the second culture medium is formed by adding cytokines IL-2, IL-18, sodium ferulate and soybean lecithin into the basic culture medium; the third culture medium is composed of sodium ferulate and soybean lecithin. The cell factor sodium ferulate and soybean lecithin are added into the culture solution, so that the cell culture period is shortened by 37.5%, and meanwhile, the amplification multiple, the purity and the tumor killing effect of the NKT cells are improved.

Description

technical field [0001] The present invention relates to the preparation and culture method of cell culture fluid, in particular to a culture fluid and culture method of umbilical cord blood NKT cells. Background technique [0002] NKT cells (natural killer T Cells) are a type of killer cells induced by a variety of cytokines. They are immune cells with the strongest cytotoxic activity. They have both the high tumoricidal activity of T lymphocytes and the non-primary NK cells. Limited tumoricidal effect of tissue-compatible complexes (MHC). NKT cells have the advantages of fast proliferation and high tumoricidal activity. [0003] At this stage, NKT cell culture is to treat the isolated peripheral blood mononuclear cells (PBMC) with IFN-γ for 24 hours, then add CD3 monoclonal antibody, IL-1α and IL-2 factors for stimulation and induction to obtain a certain number of NKT cells, but the effective cell content in the final NKT cells is low in purity, weak in cell killing acti...

Claims

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Application Information

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IPC IPC(8): C12N5/0783
CPCC12N5/0646C12N2501/515C12N2501/599C12N2501/2302C12N2501/2301C12N2501/2315C12N2501/125C12N2501/115C12N2501/2318C12N2500/36C12N2501/999
Inventor 谢海涛谢炜豪薛卫巍
Owner GUANGDONG XIANKANGDA BIOTECH CO LTD