Method for culturing NK cells in vitro by removing mononuclear cells

Abstract

The invention relates to the technical field of cell culture, in particular to a method for culturing NK cells in vitro by removing mononuclear cells. The method for culturing the NK cells in vitro byremoving the mononuclear cells comprises the following steps: (1) separating serum from whole blood cells; (2) inactivating blood; (3) obtaining the mononuclear cells; (4) carrying out adherent culture of the mononuclear cells; (5) activating the mononuclear cells; (6) inducing the NK cells; and (7) amplifying the NK cells. According to the method, coating is not needed, the operation is simple,the NK cells can grow dominantly by activating the culture medium and proliferating the culture medium, the amplification multiple can reach 1000-1300 times after 10-20 days of culture, detection by aflow cytometry shows that the content of CD3-C56+ is in a range between 80% and 98%, and industrial production is facilitated.

Description

technical field [0001] The invention relates to the technical field of cell culture, in particular to a method for culturing NK cells in vitro without monocytes. Background technique [0002] As a major disease, tumor has always troubled human health. With the advancement of medicine, the cure rate and survival rate of cancer have been significantly improved, but at present, due to the complexity, diversity, variability and heterogeneity of tumors, a better treatment method cannot be found. At present, the treatment methods for cancer in the medical industry include surgical resection, chemotherapy and radiation therapy, but the damage to the human body is relatively large. The advantages of adoptive immunotherapy, such as outstanding targeting, less side effects, and simple intervention methods, have become a hot spot for research on tumor treatment today, and have been widely used in clinical practice. [0003] NK cells have become one of the outstanding cells of adoptiv...

AI Technical Summary

Problems solved by technology

[0006] The purpose of the present invention is to overcome factors such as the instability of NK cell culture in vitro, the use of exogenous serum, the use of feeder cells, the low flow cytometry results, the cumbersome coating operation and the increased risk of bacterial infection, etc., to inhibit NK cells through adherent removal. Cell-activated monocytes are then stimulated with IL-2, IL-12, IL-15, IL-18, and CD16 monoclonal antibodies to induce NK cells, and CTLA-4 and CD28 are used to competitively bind to T cell activation receptors to inhibit T cells are activated and then expanded with serum-free complete medium containing IL-2 and IL-15 to obtain a large number of stable and high-purity NK cells

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