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CIK (cytokine induced killer) cell and culture method and application thereof

A cell culture and culture method technology, applied in the field of CIK cells and their culture, can solve the problems of unsatisfactory CIK cytotoxicity, unsatisfactory tumor treatment effect, insufficient culture expansion multiples, etc. Proliferative ability, the effect of avoiding exogenous infection

Inactive Publication Date: 2016-08-24
SHENZHEN RUI XIANG YUAN SCI & TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] The purpose of the present invention is to overcome the above-mentioned deficiencies of the prior art, provide a kind of CIK cell and its culture method, to solve the existing CIK cell toxicity unsatisfactory, its culture has insufficient expansion multiple, high cost a

Method used

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  • CIK (cytokine induced killer) cell and culture method and application thereof
  • CIK (cytokine induced killer) cell and culture method and application thereof
  • CIK (cytokine induced killer) cell and culture method and application thereof

Examples

Experimental program
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Example Embodiment

[0050] Example 1

[0051] This embodiment provides an in vitro preparation method of CIK cells with high cytotoxic activity and CIK cells cultured by the method. The CIK cell body culture method includes the following steps:

[0052] S11. Antibody coating:

[0053] Coat 10μg / ml anti-CD3 monoclonal antibody and 10μg / ml anti-CD28 monoclonal antibody in a T-75 cell culture flask, and leave it in a 37°C incubator for 5-8 hours, or a 4°C refrigerator overnight;

[0054] S12. PMBC preparation:

[0055] A sterile blood collection tube containing sodium heparin was used to collect 50 ml of peripheral blood, and the anticoagulated peripheral blood was diluted equally with 0.9% normal saline. After mixing, the diluted blood was slowly added to the human lymphatic separation fluid, and centrifuged at 2000 rpm for 20 min. Aspirate the milky white mononuclear cell layer at the interface of the separation solution, and centrifuge and wash twice to calculate the total number of mononuclear cells; fi...

Example Embodiment

[0062] Example 2

[0063] This embodiment provides an in vitro preparation method of CIK cells with high cytotoxic activity and CIK cells cultured by the method. The CIK cell body culture method includes the following steps:

[0064] S21. Antibody coating:

[0065] Coat 10μg / ml of anti-CD3 monoclonal antibody in a T-75 cell culture flask, and leave it in a 37°C incubator for 5-8 hours, or a 4°C refrigerator overnight;

[0066] S22. PBMC preparation:

[0067] A sterile blood collection tube containing sodium heparin was used to collect 50 ml of peripheral blood, and the anticoagulated peripheral blood was diluted equally with 0.9% normal saline. After mixing, the diluted blood was slowly added to the human lymphatic separation fluid, and centrifuged at 2000 rpm for 20 min. Aspirate the milky white mononuclear cell layer at the interface of the separation solution, and centrifuge and wash twice to calculate the total number of mononuclear cells; finally resuspend the PBMC with KBM581 me...

Example Embodiment

[0074] Example 3

[0075] This embodiment provides an in vitro preparation method of CIK cells with high cytotoxic activity and CIK cells cultured by the method. The CIK cell body culture method includes the following steps:

[0076] S31. Antibody coating:

[0077] Coat 5μg / ml anti-CD3 monoclonal antibody and 5μg / ml anti-CD28 monoclonal antibody in a T-75 cell culture flask, and leave it in a 37°C incubator for 5-8 hours, or a 4°C refrigerator overnight;

[0078] S32. PBMC preparation:

[0079] A sterile blood collection tube containing sodium heparin was used to collect 50 ml of peripheral blood, and the anticoagulated peripheral blood was diluted equally with 0.9% normal saline. After mixing, the diluted blood was slowly added to the human lymphatic separation fluid, and centrifuged at 2000 rpm for 20 min. Aspirate the milky white mononuclear cell layer at the interface of the separation liquid, and centrifuge and wash twice to calculate the total number of mononuclear cells; finall...

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Abstract

The invention discloses a CIK (cytokine induced killer) cell and a culture method and an application thereof. The culture method of the CIK cell comprises the following steps of collecting a peripheral blood mononuclear cell of a patient, adding the peripheral blood mononuclear cell, an anti-CD3 antibody, an anti-CD28 antibody and cell factors into a serum-free medium, and performing induced culture. The culture method of the CIK cell has the advantages that the stimulation activity for stimulating and inducing the mononuclear cell is improved under the synergetic action of the anti-CD28 antibody and the anti-CD3 antibody; the CIK cell is jointly induced by the cell factors IL-2 and IL-21, so that the joint application of the cell factors can generate the synergetic action on the amplification of lymphocyte, the killing activity of the CIK cell is improved, and the toxic or side effect caused by the application of multiple single factors is reduced; the culture time is shortened, the culture cost is reduced, and the purity of the CIK cell is guaranteed.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a CIK cell, a culture method thereof and an application thereof. Background technique [0002] Malignant tumors are currently one of the major diseases that endanger human health. In countries where infectious diseases are under control, cardiovascular and cerebrovascular diseases and malignant tumors have become the first and second causes of death, respectively. According to statistics in 2005, malignant tumors are the leading cause of death among urban residents in my country, and the second leading cause of death among rural residents. [0003] Traditional treatments for cancer include surgery, chemotherapy, and radiation. However, these therapies have limitations: surgery often cannot eradicate cancer cells due to their infiltration into adjacent or distant tissues; chemotherapy and radiotherapy are limited by the toxicity and damage to other normal tissues in the b...

Claims

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Application Information

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IPC IPC(8): C12N5/0783A61K35/17A61P35/00
CPCA61K35/17C12N5/0646C12N2500/90C12N2501/2302C12N2501/2321C12N2501/998
Inventor 张文张诺琳陈伟雄
Owner SHENZHEN RUI XIANG YUAN SCI & TECH CO LTD
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