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a tonsil-derived t γδ Cell and preparation method and application thereof

A tonsil and cell technology, applied in the field of Tγδ cell preparation, can solve the problems of poor tumor cell killing ability, short cell survival time, short expansion period, etc., and achieve excellent anti-tumor or virus, excellent anti-tumor effect.

Active Publication Date: 2022-08-02
CELARTICS BIOPHARMA CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, it is reported that the amplification period of this method is short (generally about 14 days, and the number of amplifications is limited), and the obtained T γδ The survival time of the cells is short (the amplified cells can continue to survive for about 7 days), the cells have poor anti-apoptosis ability, and the ability to kill tumor cells is poor.

Method used

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  • a tonsil-derived t <sub>γδ</sub> Cell and preparation method and application thereof
  • a tonsil-derived t <sub>γδ</sub> Cell and preparation method and application thereof
  • a tonsil-derived t <sub>γδ</sub> Cell and preparation method and application thereof

Examples

Experimental program
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Effect test

preparation example

[0040] Lentiviral plasmid vector construction and lentiviral preparation method, comprising the following steps:

[0041] (1) The gene fragment encoding the Tax protein of STLV4 virus was inserted into the lentiviral vector, and the recombinant plasmid vector was transformed into E.coli. After the sequencing was correct and the bacteria were shaken, the recombinant plasmid vector was extracted and used.

[0042](2) Mix the obtained STLV4 virus lentiviral plasmid vector with the mixed packaging plasmid (the mixed packaging plasmid includes the expression plasmids of VSV-G, Gag-Pol and Rev), and add transfection reagent; the transfection reagent can be phosphoric acid Calcium transfection reagent, liposome transfection reagent or polymer transfection reagent. Subsequently, the lentiviral plasmid vector of STLV4 virus and the mixed packaging plasmid are co-transfected into host cells for co-culture, and the host cells can be 293 cells, 293T cells or 293FT cells.

[0043] (3) Con...

Embodiment 1

[0055] A tonsil-derived T γδ The preparation method of cells includes the following steps:

[0056] S1. Isolation of immune cells from tonsils

[0057] A method for isolating immune cells from tonsils, comprising the following steps:

[0058] S11. Sample transportation and pretreatment: extract fresh tonsils from individuals undergoing routine tonsillectomy, put the fresh tonsils in RPMI1640 medium containing penicillin, and transport them to the laboratory under the condition of cold chain 2~8℃ , and the extraction work is completed within 1~3h.

[0059] Inside the biosafety cabinet, using sterile forceps, place the fresh tonsil specimen on a sterile 10 cm first cell culture dish and keep the tissue moist with 2 mL of HANK'S buffer containing antibiotics; then use scissors to remove the fresh tonsil specimen Cut into 3-5mm tissue pieces.

[0060] S12. Prepare immune cell suspension:

[0061] S121. Put the sterilized stainless steel mesh (200 mesh) into a sterile 10cm sec...

Embodiment 2

[0082] The difference between this example and Example 1 is that the operation of step S23 is: resuspend the cells with RPMI1640 medium containing 10wt% serum, add 200unit / mL IL2 to continue culturing the cells, and change the medium every day for the first 2 weeks. Others are the same as in Example 1.

[0083] The experimental results are: compared with Example 1, the medium is more likely to turn yellow, indicating that the cells grow vigorously. However, after 2 weeks, the growth rate of cells was significantly reduced, and a large number of dead cells appeared, and the overall state of the cells was worse than that of Example 1, resulting in a lower success rate of final preparation of such DC cells than Example 1.

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Abstract

This application relates to T γδ The technical field of cell preparation, specifically discloses a tonsil-derived T γδ Cells and preparation methods and uses thereof. T γδ A cell preparation method includes the following steps: S1, separating immune cells from tonsils; S2, contacting the immune cells with the Tax protein of STLV4 virus, and preparing dendritic cells after cell culture; S3, taking a new step S1 to prepare The immune cells and the dendritic cells are mixed and cultured to prepare the T cells present in the cell culture. γδ cell. T of this application γδ Cells can be used to prepare anti-tumor or anti-viral cell products; T of this application γδ The cells still had excellent antitumor effect after 3 weeks of culture in vitro.

Description

technical field [0001] This application relates to T γδ The technical field of cell preparation, more specifically, it relates to a tonsil-derived T γδ Cells and preparation methods and uses thereof. Background technique [0002] The tonsils are immunocompetent organs and the immune system's first line of defense against ingested or inhaled foreign pathogens, so they are often congested to aid in the immune response to common diseases such as respiratory viral infections. However, recurrent tonsillitis seriously affects the quality of life. Therefore, tonsillectomy has become the most commonly performed operation in otolaryngology to treat recurrent chronic tonsillitis. It is known that tonsil mucosa contains a large amount of lymphoid tissue, which is the gathering place of various immune cells. The immune cells in the excised tonsils can be fully utilized, cultured and activated, and can be used for the treatment of autologous or allogeneic cancer, or viral infectious d...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/0783
CPCC12N5/0636C12N5/0639C07K14/005C12N2740/00022C12N2501/2302C12N2509/00C12N2509/10
Inventor 张欢于洋程铧
Owner CELARTICS BIOPHARMA CO LTD
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