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Cryoprotective agent for maintaining high killing ability of DC-CIK (dendrite cell-cytokine induced killer) cells

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

Active Publication Date: 2017-03-08
武汉光谷中源协和细胞基因科技有限公司
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The applicant found that although the DC-CIK cells with low expression of miRNA-155 had very high lethality to leukemia cells before cryopreservation, the lethality decreased significantly after cryopreservation and recovery
This indicates that traditional cell cryopreservation agents may not be suitable for the cryopreservation of DC-CIK cells with low expression of miRNA-155. Although they can maintain their recovery rate, they cannot maintain their lethality against leukemia cells

Method used

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  • Cryoprotective agent for maintaining high killing ability of DC-CIK (dendrite cell-cytokine induced killer) cells
  • Cryoprotective agent for maintaining high killing ability of DC-CIK (dendrite cell-cytokine induced killer) cells
  • Cryoprotective agent for maintaining high killing ability of DC-CIK (dendrite cell-cytokine induced killer) cells

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Effect test

Embodiment 1

[0019] Example 1: Effect of cryopreservation on the lethality of DC-CIK cells with low expression of miRNA-155

[0020] 1. Experimental materials

[0021] 1. The miRNA-155 inhibitor was provided by Shanghai Gemma Pharmaceutical Technology Co., Ltd. (sequence 1 below); the inhibitor negative control was provided by Shanghai Gemma Pharmaceutical Technology Co., Ltd. (sequence 2 below).

[0022] Sequence 1: 5'-ACCCCUAUCACGAUUAGCAUUAA-3';

[0023] Sequence 2: 5'-CAGUACUUUUGUGUAGUACAA-3'.

[0024] 2. The tumor cells are leukemia cells, including K562 / A02, THP-1 and HL-60 cells.

[0025] 2. Experimental method

[0026] 1. Isolation and culture of effector cells DC and CIK

[0027] (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 ...

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Abstract

The invention discloses a cryoprotective agent for maintaining the high killing ability of DC-CIK (dendrite cell-cytokine induced killer) cells. The cryoprotective agent comprises RPMI 1640, fetal calf serum, dimethyl sulfoxide and heterophyllin B or C. The RPMI 1640, the fetal calf serum and the dimethyl sulfoxide are mixed with one another according to a volume ratio of 6:3:1; the concentration of the heterophyllin B is 7-9 micro-g / ml; the concentration of the heterophyllin C is 4-6 micro-g / ml. The cryoprotective agent has the advantages that the high killing ability of the DC-CIK cells can be maintained to the greatest extent on the basis that the cryopreservation resuscitation rates of the DC-CIK cells are guaranteed, and the strong killing ability owing to low expression of miRNA [micro-RNA (ribonucleic acid)]-155 can be lengthened.

Description

technical field [0001] The invention belongs to the field of immunization and relates to the cultivation of DC-CIK cells, in particular to a cryoprotectant for maintaining high lethality of DC-CIK cells. Background technique [0002] The applicant submitted a patent application for DC-CIK cell culture on December 22, 2016, which disclosed the application of miRNA-155 and its inhibitors in DC-CIK cell culture, miRNA-155 can be used as a drug target Application in improving the lethality of CIK cells enhanced by DC cell co-culture to tumor cells. The specification of the application discloses that DC-CIK cells with down-regulated expression of miRNA-155 have higher lethality than DC-CIK cells with normal expression of miRNA-155, and DC cells with down-regulated expression of miRNA-155 are more The lethality of DC cells was basically the same without significant difference. This indicated that down-regulation of miRNA-155 could not directly improve the lethality of DC cells a...

Claims

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Application Information

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IPC IPC(8): A01N1/02
CPCA01N1/0221
Inventor 不公告发明人
Owner 武汉光谷中源协和细胞基因科技有限公司
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