Reagent kit and method for cultivating immune cells and application thereof

A technology of immune cells and culture methods, applied in the direction of blood/immune system cells, animal cells, vertebrate cells, etc., can solve the problems of lack of anti-tumor cell group synergy, limited effect, etc., to improve systemic immune suppression and prolong survival long-term, good synergistic effect

Inactive Publication Date: 2016-01-06
SUN YAT SEN UNIV +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although these single types of anti-tumor immune lymphocytes are used for experimental research to obtain the anti-tumor mechanism of this type of cell population, the clinical treatment of tumor patients often has limited effects due to the lack of synergistic effects of other anti-tumor cell populations

Method used

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  • Reagent kit and method for cultivating immune cells and application thereof
  • Reagent kit and method for cultivating immune cells and application thereof
  • Reagent kit and method for cultivating immune cells and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] A kit for culturing immune cells, which includes culture medium A and coating fluid.

[0043] Among them, the culture medium A is prepared according to the following method: add rhIFN-γ, rhIL-2, rhIL-15 and human transferrin into the serum-free lymphocyte culture medium and mix well, wherein, the serum-free lymphocyte culture medium used Specifically, KBM561 medium (TaKaRa company, Japan); rhIFN-γ (trade name: Clone Gamma), rhIL-2 (Beijing Shuanglu Pharmaceutical Co., Ltd.), rhIL-15 and human transferrin (Sigma company, U.S.A.) The final concentrations in culture solution A were 1000U / mL, 1000U / mL, 5ng / mL, 5ug / ml respectively.

[0044] The coating solution was prepared as follows: OKT3 antibody and Retronectin were added to PBS buffer and mixed evenly. Among them, the final concentrations of OKT3 antibody (Janssen Pharmaceutical Company, Japan) and Retronectin (TaKaRa Company, Japan) in the coating solution were both 100 ng / mL.

Embodiment 2

[0046] A method for culturing immune cells, carried out according to the following steps:

[0047] 1) Mononuclear cells (PBMC) were isolated from the peripheral blood of head and neck cancer patients who had not undergone chemotherapy by Ficoll–Hypaque density gradient centrifugation;

[0048] 2) Inoculate the mononuclear cells in step 1) into the culture flasks coated with the coating solution of Example 1, and culture them with activation medium, at 37°C, 5% CO 2 4 days in the cell culture box; Wherein, the activation medium used is to add the plasma (autologous plasma) of this head and neck tumor patient to the culture medium A of embodiment 1 to make, and this plasma is in this activation medium The mass percentage is 2%;

[0049] 3) Transfer the cells cultured in step 2) to a large culture bottle or a culture bag, and continue culturing with the culture solution A in Example 1. During the culture process, replace the culture solution A every 2 to 3 days.

[0050] 4) The...

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Abstract

The invention provides a reagent kit for cultivating immune cells, a method for cultivating the immune cells by the aid of the reagent kit and application of the immune cells cultivated by the aid of the method to preparing preparations for preventing or treating tumors. The reagent kit comprises cultivation liquid A. RhIFN-gamma (recombinant human interferon-gamma), rhIL-2 (recombinant human interleukin-2), rhIL-15 and human transferrin are added into serum-free lymphocyte cultivation liquid to obtain the cultivation liquid A. The reagent kit, the method and the application have the advantages that the quantities of the immune cells cultivated by the aid of the method can reach approximately 10 billions, and the immune cells include large quantities of NK (natural killer) cell and NKT (natural killer T) cell masses besides CTL (cytotoxic T lymphocyte) masses; the immune cells cultivated by the aid of the method can be applied to HNSCC (head and neck squamous cell carcinoma) treatment, so that immunosuppression due to chemotherapy can be improved, and the lifetime of patients can be prolonged.

Description

technical field [0001] The invention belongs to the technical field of biomedicine, and in particular relates to a kit for culturing immune cells, a method for cultivating immune cells and its application. Background technique [0002] As we all know, malignant tumors are currently the number one killer that endangers human health. Taking HNSCC (head and neck cancer) as an example, according to statistics, about 650,000 new HNSCC patients appear every year in the world, and about 350,000 cancer patients die of HNSCC. At present, surgical resection, radiotherapy, and chemotherapy are still commonly used methods for HNSCC, but these treatments have not achieved satisfactory results. Surgical resection alone has disadvantages such as poor prognosis, high recurrence rate and tumor metastasis rate, and is not suitable for patients with advanced cancer. Chemotherapy before or after radical resection did not improve the prognosis of HNSCC patients. Chemotherapy is often accompan...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/078C12N5/0786A61K35/17A61P35/00
Inventor 汪华蒋盼张雁张海霞
Owner SUN YAT SEN UNIV
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