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A cryoprotectant for maintaining high lethality of cik cells

A cryopreservation and lethality technology, applied in the field of immunity, can solve the problems of decreased lethality, inapplicability, and inability to maintain the lethality of leukemia cells, and achieve the effect of continuing lethality

Active Publication Date: 2020-07-14
苏州药明生物技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The applicant found that CIK cells with low expression of miRNA-146a had very high lethality to leukemia cells before cryopreservation, but the lethality decreased significantly after cryopreservation and recovery
This indicates that traditional cell cryopreservation agents may not be suitable for the cryopreservation of CIK cells with low expression of miRNA-146a. Although they can maintain their recovery rate, they cannot maintain their lethality against leukemia cells

Method used

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  • A cryoprotectant for maintaining high lethality of cik cells
  • A cryoprotectant for maintaining high lethality of cik cells
  • A cryoprotectant for maintaining high lethality of cik cells

Examples

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Effect test

Embodiment 1

[0017] Example 1: Effect of cryopreservation on the lethality of CIK cells with low expression of miRNA-146a

[0018] 1. Experimental materials

[0019] miRNA-146a inhibitor and inhibitor negative control were provided by Shanghai Gemma Pharmaceutical Technology Co., Ltd. Tumor cells are leukemia cells, including K562 / A02, THP-1 and HL-60 cells. Other reagents or instruments are commonly used reagents or instruments in molecular biology and immunology.

[0020] 2. Experimental method

[0021] 1. Isolation and culture of effector cell CIK

[0022] (1) Separation of mononuclear cells: collect 20 mL of peripheral blood from healthy volunteers, dilute it with pre-cooled PBS 1:1, slowly add the upper layer of lymphocyte separation medium, centrifuge at 650g, 4°C for 20 min, collect the white cell layer, and separate mononuclear cells , 1640 medium to resuspend cells and place at 37°C, 5% CO 2 Incubate for 2 hours in the incubator.

[0023] (2) Cultivation of CIK cells: collec...

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Abstract

The invention discloses a cryopreservation protection agent for keeping the high lethality of CIK cells. The cryopreservation protection agent comprises RPMI 1640, fetal calf serum, dimethyl sulfoxide and cyclohexapeptide Cyclo(-Gly-Thr-Phe-Leu-Tyr-Ala-); the cryopreservation protection agent is prepared through mixing the RPMI 1640, the fetal calf serum and the dimethyl sulfoxide according to a volume ratio of 6:3:1; and the concentration of the cyclohexapeptide Cyclo(-Gly-Thr-Phe-Leu-Tyr-Ala-) is 2-4 [mu]g / ml. The cryopreservation protection agent maximally keeps the high lethality of the CIK cells based on guaranteeing the cryopreservation recovery rate of the CIK cells, and continues the strong lethality brought by miRNA-146a.

Description

technical field [0001] The invention belongs to the field of immunization and relates to the cultivation of CIK cells, in particular to a cryopreservation protective agent for maintaining high lethality of CIK cells. Background technique [0002] The applicant submitted a patent application for CIK cell culture on December 22, 2016. The application disclosed the application of miRNA-146a and its inhibitors in CIK cell culture. miRNA-146a can be used as a drug target to improve the ability of CIK cells to lethality of tumor cells. The specification of this application discloses that CIK cells with down-regulated expression of miRNA-146a have higher lethality than CIK cells with normal expression of miRNA-146a, and the difference is very significant. Applicants also found that the cyclic hexapeptide Cyclo (-Gly-Thr-Phe-Leu-Tyr-Ala-) is a potent inhibitor of miRNA-146a. [0003] For experimental needs, researchers usually need to freeze cells for later recovery. The applican...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A01N1/02
CPCA01N1/0231
Inventor 不公告发明人
Owner 苏州药明生物技术有限公司