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Method for resuscitating mononuclear cells and inducing into multi-cytokine-induced killer cells

A cytokine and cell-killing technology, applied in the field of bioengineering, can solve the problems of clinical application of unsuitable cell products, and achieve the effect of high induction differentiation efficiency, good cell activity, and fast proliferation rate

Inactive Publication Date: 2018-07-06
SOUTH CHINA INSTITUDE OF BIOMEDICINE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Since fetal bovine serum is of animal origin, it is not suitable for clinical application of cell products

Method used

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  • Method for resuscitating mononuclear cells and inducing into multi-cytokine-induced killer cells
  • Method for resuscitating mononuclear cells and inducing into multi-cytokine-induced killer cells
  • Method for resuscitating mononuclear cells and inducing into multi-cytokine-induced killer cells

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0055] 1. PBMC recovery

[0056] (1) Peripheral blood mononuclear cells (PBMCs) cryopreserved in liquid nitrogen were taken out and rapidly thawed in a water bath at 37-42°C.

[0057] (2) The cells were treated with an equal volume of X-VIVO15 medium (referred to as group P) and an equal volume of recovery solution containing 2.5% HSA and 5% Dextran40 (referred to as group F1).

[0058] (3) Centrifuge, take the cells separately, count with a cell viability analyzer, and measure the cell viability.

[0059] 2. Induced into CIK cells

[0060] Cells in group P and group F1 were cultured statically for 24 hours in X-VIVO15 medium, respectively. Then, 1000U / ml IFN-γ and 2.5% Supgrow were added to the cell culture medium of the P group and the F1 group, after continuing to culture for 24 hours, 100ng / ml CD3, 1000U / ml IL-2, 1000U / ml IL-1α were added, Perform in vitro induction amplification.

[0061] 3. Calculate

[0062] Since the number of initial cells is inconsistent when re...

Embodiment 2

[0089] 1. Cord blood MNC resuscitation

[0090] (1) The cryopreserved umbilical cord blood mononuclear cells (MNC) were taken out from liquid nitrogen and rapidly thawed in a water bath at 37-42°C.

[0091] (2) Use equal volumes of X-VIVO15 medium (referred to as P group), and equal volumes of resuscitation fluid containing 2.5% HSA and 5% Dextran40 (referred to as F1 group), 2.5% plasma, 5% Dextran40 Cells were treated in four ways with resuscitation solution (referred to as F2 group), resuscitation solution with 5% plasma and 5% Dextran40 (referred to as F3 group).

[0092] (3) Centrifuge, take the cells separately, and use a cell viability analyzer to count and measure the cell viability.

[0093] 2. Induced into CIK cells

[0094] The cells treated in groups P and F1-3 were cultured statically for 24 hours in X-VIVO15 medium, respectively. The cells treated in the P and F1-3 groups were treated by adding 1000U / ml IFN-γ and 2.5% Supgrow, and continued to culture for 24 h...

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Abstract

The present invention discloses a method for resuscitating mononuclear cells and inducing into multi-cytokine-induced killer cells. The method comprises: placing cryopreserved mononuclear cells in a water bath with a temperature of 37-42 DEG C, and thawing to obtain thawed mononuclear cells; mixing the thawed mononuclear cells and a freezing resuscitation liquid for multi-cytokine-induced killer cells to obtain a resuscitation cell mixing liquid, wherein the freezing resuscitation liquid contains albumin and dextran; carrying out centrifugation treatment on the resuscitation cell mixing liquidto obtain resuscitated cells; and carrying out directional induction treatment on the resuscitated cells to obtain the induced multi-cytokine-induced killer cells. According to the present invention,the multi-cytokine-induced killer cells obtained through the induction and amplification of the mononuclear cells obtained through the resuscitating have advantages of rapid proliferation, high induction differentiation efficiency and good cell viability. .

Description

technical field [0001] The invention relates to the field of bioengineering, specifically, a method for recovering mononuclear cells and inducing their expansion into killer cells induced by various cytokines. Background technique [0002] Cytokine-induced killer cells (CIK) are a group of heterogeneous cells obtained from mononuclear cells in human blood induced and expanded by various cytokines in vitro. The cells express both CD3+ and CD56+ cell membrane surface markers, so they are called NK cell-like T lymphocytes, which have T lymphocyte activity and non-MHC restriction of NK cells. Therefore, it can be used to activate the body's immune system and improve the body's immunity; CIK cells can secrete various cytokines such as IL-2, IL-6, IFN-γ, and treat leukemia, lymphoma, lung cancer, gastric cancer, and intestinal cancer. It plays an important role in preventing tumor recurrence and metastasis and improving the quality of life of patients. Effector cells CD3+CD56+ c...

Claims

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Application Information

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IPC IPC(8): C12N5/0783
CPCC12N5/0037C12N5/0646C12N2500/34C12N2501/24C12N2501/998
Inventor 裴雪涛陈琳陈海明岳文姚海雷习佳飞何丽娟张博文贾雅丽南雪谢小燕
Owner SOUTH CHINA INSTITUDE OF BIOMEDICINE
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