Inducing culture reagent for CIK (Cytokines Induced Killer) cells and culture method thereof

A culture method and reagent technology, applied in the direction of cell culture active agent, tissue culture, animal cells, etc., to achieve the effect of improving anti-tumor effect and increasing the number of proliferation

Inactive Publication Date: 2017-08-01
北京焕生汇生物技术研究院有限公司
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] The object of the present invention is to provide a CIK cell induction culture reagent and its culture method, mainly to solve the stability problems of cell induction quality and cell proliferation quantity in the CIK cell culture process

Method used

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  • Inducing culture reagent for CIK (Cytokines Induced Killer) cells and culture method thereof
  • Inducing culture reagent for CIK (Cytokines Induced Killer) cells and culture method thereof
  • Inducing culture reagent for CIK (Cytokines Induced Killer) cells and culture method thereof

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Embodiment 1

[0022] A CIK cell induction culture reagent, the reagent includes reagent A: 50ng / ml anti-CD3 monoclonal antibody; reagent B: 1000IU / ml IFN-γ; reagent C: 1000IU / ml IL-2 and 20ng / ml IL-15 ; Reagent D: 100IU / ml IL-1; Reagent E: 40ml of lymphocyte separation liquid; Utilize above-mentioned reagent to induce culture method to CIK cell, comprise the following steps:

[0023] 1) Take Reagent A and coat it in a T-175 culture flask, overnight at 4°C;

[0024] 2) Take 30ml of peripheral blood collected by machine, dilute to 50ml with D-PBS, and use density gradient centrifugation to harvest PBMC with reagent E, 1500rpm for 30 minutes;

[0025] 3), wash the collected PBMC with D-PBS twice, 1350rpm for 5 minutes, resuspend the cleaned PBMC in 100ml X-VIVO15 medium according to 1-2*10^6 / ml, add reagent B, set Incubate in a carbon dioxide incubator;

[0026] 4), after 24 hours, add reagent C 100ul and reagent D;

[0027] 5), on the third day, add 150ml of medium and 150ul of reagent C; ...

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Abstract

The invention discloses an inducing culture reagent for CIK (Cytokines Induced Killer) cells. The reagent is prepared from a reagent A: 50ng/ml anti-CD3 monoclonal antibody; a reagent B: 1000IU/ml IFN-gamma; a reagent C: 1000IU/ml IL-2 and 20ng/ml IL-15; a reagent D: 100IU/ml IL-1; a reagent E: 40ml of lymphocyte separation medium. The invention also discloses an inducing culture method for the CIK cells by using the reagent. The reagent and the method, provided by the invention have the advantages that simple, high-efficient and stable induced CIK cells can be realized; by adding IL-15 in a classic preparation solution, multiplication number of the CIK cells and secretion of gamma-IFN can be better improved, and the anti-tumor effect of the CIK cells is improved; the cell inducing quality during CIK cell culture is high and the stability of cell multiplication number is good.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a CIK cell induction culture reagent and a culture method thereof. Background technique [0002] As the fourth new method of treating cancer after surgery, radiotherapy and chemotherapy, immune cell therapy has made great progress in recent years. Currently, immune cells have been widely recognized by the medical community with their efficient tumor-killing activity and no side effects. CIK As a relatively mature treatment method in immunotherapy, there are currently many induction methods, and their quality and quantity are also uneven; there are also researches on the preparation of CIK in different directions, but in practical applications, due to many cytokines and preparations, Due to the long period and different preparation standards, the preparation process is cumbersome and unstable, the quality and quantity of cells cannot be effectively controlled, and even pol...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/0783
CPCC12N5/0638C12N2501/2301C12N2501/2302C12N2501/2315C12N2501/24C12N2501/515
Inventor 姚玲玲杨苗杨敏孟得龙宋丹
Owner 北京焕生汇生物技术研究院有限公司
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