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Enhanced NK cell with strong immune regulation and strong killing capacity on tumor cells and virus infected cells, preparation method and kit thereof

A technology of NK cells and tumor cells, applied in the field of cell culture, can solve the problems of cumbersome operation, high cost, and increased risk of clinical application, and achieve the effect of enhancing proliferation ability and enhancing killing activity

Inactive Publication Date: 2020-07-10
北京荟科柘生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, these methods have obvious disadvantages
The use of feeder cells undoubtedly introduces exogenous cells, which increases the risk of clinical application; while magnetic bead sorting and flow sorting operations are cumbersome, which undoubtedly increases the risk of cell contamination, and the cost is also high

Method used

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  • Enhanced NK cell with strong immune regulation and strong killing capacity on tumor cells and virus infected cells, preparation method and kit thereof
  • Enhanced NK cell with strong immune regulation and strong killing capacity on tumor cells and virus infected cells, preparation method and kit thereof
  • Enhanced NK cell with strong immune regulation and strong killing capacity on tumor cells and virus infected cells, preparation method and kit thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0068] test group

[0069] Experimental operations may include the following steps:

[0070] Step 1: NK cell culture flask pretreatment: Dissolve CD16 monoclonal antibody in DPBS, transfer to a T75 culture flask, and store in the dark at 37°C for 3 hours, or overnight at 4°C.

[0071] Step 2: PBMC Isolation

[0072] Sub-step 2.1: Take the lymphocyte separation solution out of the 4°C refrigerator 30 minutes in advance and place it at room temperature, and use it after the temperature rises to room temperature.

[0073] Sub-step 2.2: Pour 30ml of fresh heparin-anticoagulated peripheral blood into a 50ml centrifuge tube, balance, centrifuge at 700g / min for 20min (slowest speed drop), collect the upper layer, inactivate at 56°C for 30min, then place at 4°C for 15min , and finally centrifuged at 2000g for 20min, then sucked the upper layer liquid for later use.

[0074] Sub-step 2.3: Add DPBS to the lower layer after centrifugation of the above-mentioned peripheral blood, and m...

Embodiment 2

[0095] control group

[0096] Experimental operations may include the following steps:

[0097] Step 1: NK cell culture flask pretreatment: Dissolve CD16 monoclonal antibody in 20ml DPBS, transfer to a T75 culture flask, and store in the dark at 37°C for 3 hours, or overnight at 4°C.

[0098] Step 2: PBMC isolation.

[0099] Sub-step 2.1: Take the lymphocyte separation solution out of the 4°C refrigerator 30 minutes in advance and place it at room temperature, and use it after the temperature rises to room temperature.

[0100] Sub-step 2.2: Pour 100ml of fresh heparin-anticoagulated peripheral blood into a 50ml centrifuge tube, balance, centrifuge at 700g / min for 20min (slowest speed drop), collect the upper layer, inactivate at 56°C for 30min, then place at 4°C for 15min , and finally centrifuged at 2000g for 20min, then sucked the upper layer liquid for later use.

[0101] Sub-step 2.3: Add DPBS to the lower layer after centrifugation of the above-mentioned peripheral bl...

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Abstract

The invention discloses an enhanced NK cell with strong immune regulation and strong killing capacity on tumor cells and virus infected cells, a preparation method and a preparation kit thereof. A Transwell culture system is adopted to use mesenchymal stem cells to incubate and acclimate NK cells before activation, so that the Transwell culture system co-cultures the mesenchymal stem cells and theNK cells, and the immune regulation ability of the NK cells is acclimated. In addition, the NK cells are activated and cultured by CD16 monoclonal antibodies combined with IFN-gamma, OK432, IL-2, IL-21 and IL-15, and the NK cells are amplified and cultured by IL-2 and nicotinamide, so that the proliferation ability of the NK cells is enhanced, and the killing activity of the NK cells on the tumorcells and the virus infected cells is promoted.

Description

technical field [0001] This description relates to the field of cell culture, in particular to an enhanced NK cell capable of strong immune regulation and strong killing of tumor cells and virus-infected cells, a preparation method and a kit thereof. Background technique [0002] Natural killer cells (Natural Killer cells, NK cells) are the third largest type of lymphocytes except T cells and B cells. They belong to the first line of defense of our human immunity and are also an important member of the human immune cell family. Fighting on the front line of protecting human health all the year round, it is known as the core cell of natural immunity. NK cells are not only related to anti-tumor, anti-viral infection and immune mediation, but even participate in the occurrence of hypersensitivity and autoimmune diseases. [0003] Natural killer cells are important immune cells in the body. They are not only related to anti-tumor, anti-viral infection and immune regulation, but...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/0783C12N5/0775
CPCC12N5/0646C12N2500/38C12N2501/06C12N2501/2302C12N2501/2315C12N2501/2321C12N2501/24C12N2502/1352
Inventor 不公告发明人
Owner 北京荟科柘生物科技有限公司
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