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A kind of culture method of specific T cell

A culture method and specific technology, applied in the field of specific T cell culture, can solve the problems of less TCM content, low TCM purity, high cost, etc., and achieve the effect of long storage time, high cell purity and fast proliferation

Active Publication Date: 2020-10-02
SHANGHAI BIOMED UNION BIOTECHNOLOGY CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] 1. The TCM in peripheral blood "memorizes" different antigens, and responds to different diseases and infections. Only a very small part of TCM can recognize tumors. Therefore, TCM separated from peripheral blood has limited tumor killing ability, basically The amplified cells are all miscellaneous cells;
[0009] 2. TCM traditional culture methods often use TCR matching, tumor antigens, immunomagnetic beads, monoclonal selection, etc. These culture methods are very time-consuming, labor-intensive, costly, and difficult to promote;
[0010] 3. The content of TCM in peripheral blood is very small, and it is difficult to identify, isolate and amplify;
[0011] 4. In the process of TCM in vitro culture, TCM is easy to differentiate into effector memory T cells, which makes the purity of TCM lower

Method used

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  • A kind of culture method of specific T cell
  • A kind of culture method of specific T cell
  • A kind of culture method of specific T cell

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] Embodiment 1 cultivates TCM

[0039] 1.1 Experimental method

[0040] 1.1.1 Preparation before experiment

[0041] Preparation of lymphocytes: take the peripheral blood of healthy donors as blood samples, collect 20ml of blood samples and evenly distribute them into four 50ml centrifuge tubes containing 20ml of lymphocyte separation medium, centrifuge at 2200rpm for 20min; after standing still, use a Pasteur pipette Aspirate the middle buffy coat into a new 50ml centrifuge tube, then centrifuge with normal saline at 1800rpm for 5min, wash twice; use CD56 magnetic beads to remove other lymphocytes that do not specifically kill the tumor, centrifuge with normal saline at 1800rpm for 5min, wash for 2 times to obtain lymphocytes.

[0042] Tumor tissue preparation: Inoculate 3×106 melanoma cells A375 subcutaneously into nude mice to make tumor models; when the subcutaneous tumors in nude mice grow to 6-10cm2, kill the mice and remove them from the mice. The tumor tissue i...

Embodiment 2

[0073] Embodiment 2 activity verification experiment

[0074] 2.1 Experimental scheme

[0075] Take the cells cultured to the 7th day in Example 1, divide them into three groups A, B, and C on average, and culture them to the 8th day according to the following culture method, and detect the FACS in the A group, B group, and C group respectively, and the results are as follows: Figure 8 .

[0076] Group A: the cell volume is 5×10 7 , add expansion medium and 5 x 10 5 Lysates of A375 cells;

[0077] Group B: the cell volume is 5×10 7 , add expansion medium and 5 x 10 5 A375 cells;

[0078] Group C: the cell volume is 5×10 7 , add expansion medium.

[0079] 2.2 Result Analysis

[0080] Figure 8 The upper right quadrant is the content of TCM, and the lower right quadrant is the content of effector T cells. combine figure 2 and Figure 8 It can be seen that the A375 lysate added to the cells in group A, that is, the specificity-related antigen, can be seen that TCM ...

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Abstract

The invention relates to a culture method of specific T cells. The culture method comprises the following steps of (1) performing activation and differentiation on PBMC to obtain effector T cells; (2)using an anaplasia culture medium, and performing anaplasia on the effector T cells to obtain TCM; and (3) amplifying the TCM with an amplification culture medium. The cultured cells are high in purity and long in preservation time, have tumor specificity and are high in activity. The culture method is high in proliferation speed, low in cost and easy to operate.

Description

technical field [0001] The invention relates to a method for cultivating specific T cells, in particular to a method for cultivating tumor-specific central memory T cells. Background technique [0002] "2012 China Cancer Registration Annual Report" once reported that the incidence rate of cancer in my country is 285.91 / 100,000, and the urban and rural incidence rates are 303.91 / 100,000 and 249.98 / 100,000, respectively. It is estimated that by 2049, the incidence of cancer in my country may reach the level of 400 / 100,000. The main reason for the increase in the incidence of cancer is the aging of the population. Today, the proportion of my country's elderly population is 13.6%, and it may reach 16% in 20-30 years. Correspondingly, we can learn that the incidence of cancer has indeed been on a straight-line upward trend in recent years, and despite the continuous development of current cancer treatment measures, the physical condition of patients with advanced malignant tumo...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/0783
CPCC12N5/0636
Inventor 柴勋钱程
Owner SHANGHAI BIOMED UNION BIOTECHNOLOGY CO LTD
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