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Highly active T cell in-vitro culture kit and culture method

A kit and high-activity technology, applied in the direction of cell culture active agent, culture process, tissue culture, etc., can solve the problems of increasing operator's operation time and cell culture cycle, increasing the use of reagent consumables, increasing the risk of experimental operation, etc., to achieve Reduce pain and medical costs, increase consumption, and have no toxic and side effects

Active Publication Date: 2020-09-29
珠海贝索细胞科学技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This method of mixed reinfusion after separate culture increases the risk of experimental operation, increases the operating time of the operator and the cell culture cycle, greatly increases the use of reagent consumables, and increases the cost of treatment

Method used

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  • Highly active T cell in-vitro culture kit and culture method
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  • Highly active T cell in-vitro culture kit and culture method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0047] This example is used to introduce a kit for in vitro culture of highly active T cells of the present invention and its preparation and quality testing of the kit. The highly active T cells (HAT) of the present invention specifically refer to the γδT and CIK cells. In combination, the in vitro culture kit of the present invention can facilitate the simultaneous cultivation of two kinds of cells (γδT cells and CIK cells) at one time in clinical practice, reduce the risk of experimental operation, shorten the operation time of operators and cell culture cycle, and effectively reduce the cost of reagent consumables use.

[0048] A high-activity T cell in vitro culture kit, including the main components of zometa (zoledronic acid), PHA (phytohemagglutinin, phytohemagglutinin), her-2 (human epidermal growth factor receptor 2, human epidermal growth factor receptor 2) , INF-γ (Interferon-γ) induction solution, the main component is IL-2 (Interleukin 2) stimulating solution, the m...

Embodiment example 1

[0084] Implementation case 1 results:

[0085] Test items Inducer Stimulant Activation solution Amplification solution determination skills requirement Endotoxin<0.06EU / ml

<0.06EU / ml

<0.06EU / ml

<0.06EU / ml

qualified<0.25EU / ml

Mycoplasma Negative Negative Negative Negative qualified Negative Fungus Negative Negative Negative Negative qualified Negative bacterial Negative Negative Negative Negative qualified Negative

[0086] Table 1

[0087] As another realization, a high-activity T cell in vitro culture kit includes an induction solution with main components of zometa, PHA, her-2, and INF-γ, a stimulating solution with IL-2 as the main component, and IL -2. The activation solution of IL-1α and IL-7, the main components of which are the amplification solution of IL-2 and IL-21; in the induction solution, the effective concentrations of zometa, PHA, her-2, and INF-γ are respectively 5ug / ml, 1ug / ml, 10ug / ml and 100IU / ml; in the stimulation solution, the effective concent...

Embodiment 2

[0090] Collect human peripheral blood from a healthy volunteer, separate mononuclear cells, culture them with the kit for highly active T cells (γδT and CIK cells) described in Example 1 of the present invention, and analyze the number of cell expansion, cell viability and cell proliferation multiples .

[0091] The culture method of the present invention includes the following steps:

[0092] 1. Preparation of PBMC (peripheral blood mononuclear cells): 40 ml of human peripheral blood was collected using a heparin sodium anticoagulation tube, and the blood was separated according to density gradient centrifugation. Separate the plasma and mononuclear cells in sequence, and inactivate the plasma at 56°C for 30 minutes and centrifuge for use to prepare mononuclear cells: Dilute the separated plasma cells with PBS (phosphate buffered saline) at a ratio of 1:1, and inject them into lymphocytes Slowly rise and fall on the separating fluid by centrifugation, and carefully aspirate monon...

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Abstract

The invention relates to a highly active T cell in-vitro culture kit. The in-vitro culture kit comprises an induction solution with zometa, PHA, her-2 and INF-gamma as the main components, a stimulation solution with IL-2 as the main component, an activation solution with IL-2, IL-1 alpha and IL-7 as the main components, and an amplification solution with IL-2 and IL-21 as the main components. According to the in-vitro culture kit, two cells (gamma delta T cell and CIK cell) can be conveniently and clinically cultured and transfused back at the same time, the experimental operation risk is reduced, the operation time of operators is shortened, the cell culture period is shortened, and the use of reagent consumables is effectively reduced.

Description

Technical field [0001] The invention belongs to the technical field of immune cell therapy, and in particular relates to a highly active T cell (Highly active T cells, HAT) in vitro culture kit and a culture method. Background technique [0002] Cellular immunotherapy is a new type of biotherapeutic technology with significant curative effect. This new type of treatment method that uses the transformation of autoimmunity to fight cancer mainly uses biotechnology and biological agents to culture and transform the patient's own immune cells collected in vitro. Then it is injected into the patient's body to stimulate the recognition and guidance of immunity to kill tumor cells to enhance the body's own immune function and achieve the effect of treating tumors. [0003] γδ T cells are T cells that perform innate immune functions, and their TCR is composed of γ and δ chains. It is mainly used to kill cancer cells and help DC cells recognize and find cancer cell antigens, and then kill ...

Claims

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Application Information

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IPC IPC(8): C12N5/0783
CPCC12N5/0638C12N5/0636C12N2500/50C12N2501/24C12N2501/2302C12N2501/2301C12N2501/2307C12N2501/2321C12N2500/30C12N2501/11
Inventor 孔伟圣蓝欣冉红陈智妍
Owner 珠海贝索细胞科学技术有限公司
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