Culture solution and culture method of umbilical cord blood NK cells

A technology of NK cells and culturing method, which is applied in the culture medium and culture field of umbilical cord blood NK cells, can solve the problems of low proportion of effector cells, long culture period and high production cost, and can improve the purity, reduce a large number of deaths, and improve the killing activity. rate effect

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

However, in the prior art, the umbilical cord blood NK cell expansion and culture period is long, the proportion of effector cells is low (low purity), the operation is complicated, the umbilical cord blood NK cell culture kit is expensive, and the production cost is high.

Method used

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

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0088] 1), prepare the first medium, the second medium and the third medium

[0089] The first medium: IL-2 with a final concentration of 2000IU / ml, IL-15 with a final concentration of 50ng / ml, IL-12 with a final concentration of 75ng / ml, and IL-12 with a final concentration of 75ng / ml were added to the X-VIVO 15 basal medium IL-3 at 42ng / ml, OK432 at 1.2ug / ml final, SCF at 30ng / ml, TGF at 28ng / ml, PDGF at 16ng / ml, 3μM at final StemRegenin 1 and 0.15ng / ml Hasinide;

[0090] Second medium: add IL-2 with a final concentration of 1800IU / ml, IL-21 with a final concentration of 22ng / ml, SCF with a final concentration of 16ng / ml, and 550ug / ml of X-VIVO 15 basal medium Soy lecithin and 3ug / ml thymopentin;

[0091] The third medium: IL-2 with a final concentration of 1150 IU / ml, SCF with a final concentration of 16 ng / ml, 550 ug / ml of soybean phospholipid and 3 ug / ml of thymopentin were added to the X-VIVO 15 basal medium.

[0092] 2) Inoculation of umbilical cord blood PBMC and cu...

Embodiment 2

[0098] 1), prepare the first medium, the second medium and the third medium

[0099] The first medium: IL-2 with a final concentration of 1500IU / ml, IL-15 with a final concentration of 80ng / ml, IL-12 with a final concentration of 50ng / ml, and a final concentration of 20ng / ml were added to KBM581 basal medium. ml of IL-3, final concentration of 0.1ug / ml OK432, final concentration of 20ng / ml SCF, final concentration of 40ng / ml TGF, final concentration of 5ng / ml PDGF, final concentration of 2μM StemRegenin 1 and 0.05ng / ml Hasinide;

[0100] Second medium: KBM581 basal medium was supplemented with IL-2 with a final concentration of 1500IU / ml, IL-21 with a final concentration of 10ng / ml, SCF with a final concentration of 10ng / ml, soybean lecithin with a final concentration of 300ug / ml and 1ug / ml of thymopentin;

[0101] The third medium: IL-2 with a final concentration of 1000 IU / ml, SCF with a final concentration of 10 ng / ml, 300 ug / ml of soybean phospholipid and 1 ug / ml of thym...

Embodiment 3

[0108] 1), prepare the first medium, the second medium and the third medium

[0109] The first medium: IL-2 with a final concentration of 3000 IU / ml, IL-15 with a final concentration of 30 ng / ml, and IL-12 with a final concentration of 100 ng / ml were added to the AIM-V basal medium. 50ng / ml IL-3, 2ug / ml final OK432, 40ng / ml final SCF, 20ng / ml final TGF, 20ng / ml final PDGF, 5μM final StemRegenin 1 and 1ng / ml Hasinide;

[0110] Second medium: Add IL-2 with a final concentration of 2000IU / ml, IL-21 with a final concentration of 30ng / ml, SCF with a final concentration of 20ng / ml, and soybean with a final concentration of 20ng / ml to the AIM-V basal medium Phospholipids and thymopentin at 5ug / ml;

[0111] The third medium: IL-2 with a final concentration of 1500 IU / ml, SCF with a final concentration of 20 ng / ml, 800 ug / ml of soybean phospholipid and 5 ug / ml of thymopentin were added to the AIM-V basal medium.

[0112] 2) Inoculation of umbilical cord blood PBMC and culture

[01...

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Abstract

The invention discloses a culture solution and a culture method of umbilical cord blood NK cells. The culture solution comprises the following components: a first culture medium and a second culture medium, wherein the first culture medium is formed by adding cell factors IL-2, IL-15, IL-12, SCF, TGF, PDGF, OK432, StemRegenin 1 and halcinonide into a basic culture medium; the second culture medium is formed by adding cell factors IL-2, IL-21, SCF, soyabean lecithin, soyabean lecithin and thymopentin into the basic culture medium; the third culture medium is formed by adding cell factors IL-2, SCF, soybean lecithin and thymopentin into the basic culture medium. The cell factors 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 NK 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 NK cells. Background technique [0002] Natural killer cells (NK cells) are one of the important components of the human immune system and are effector cells of the natural immune system. They are derived from bone marrow lymphoid stem cells, differentiate and mature in the bone marrow and thymus, and are distributed in the blood, lymph nodes, bone marrow, etc. Unlike T and B cells, NK cells can non-specifically recognize and kill tumor cells and virus-infected cells without antigen sensitization and without MHC restriction. Due to this important function, the research and application of NK cells in the field of tumor immunotherapy have been paid more and more attention. [0003] NK cells have high requirements on quantity and purity in clinical application, especially the purity, which...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/0783
CPCC12N5/0646C12N2501/2302C12N2501/2303C12N2501/2312C12N2501/2315C12N2501/125C12N2500/30C12N2501/998C12N2501/135C12N2501/15C12N2501/148
Inventor 谢海涛谢炜豪薛卫巍
Owner GUANGDONG XIANKANGDA BIOTECH CO LTD
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