Human primary cell culture medium and application thereof

Abstract

The invention relates to a human primary cell culture medium, and the culture medium is used for the culture of human primary cells in vitro and the culture of human primary B acute lymphocyte leukemia (B-ALL) in vitro. The culture medium also comprises cell factors: human FLT3L, human IGF1, human IL-7 and human IL-6. The culture medium does not contain serum, and can prevent the co-culture of primary cells and matrix cells, and also solves a series of problems in conventional schemes of serum and matrix cell culture. Compared with a conventional culture mode, a mode of employing the culture medium is higher in amplification efficiency, and is longer in time of culture in vitro.

Description

technical field [0001] The present invention relates to the technical field of cell culture, in particular to a primary cell culture medium, the use of the medium for culturing human primary cells in vitro, and the use of the culture medium for culturing primary human acute lymphoblastic leukemia (B- ALL) cell method. Background technique [0002] B-cell acute lymphoblastic leukemia (B-ALL, B-cellacutelymphoblasticleukemia), also known as precursor B-cell (preB-cell) acute lymphoblastic leukemia, is a malignant tumor derived from B-cell progenitor cells. B-ALL is mainly a high-incidence cancer in children, and its incidence rate decreases in adults. Prognosis is good in juvenile patients with B-ALL, and the long-term survival rate (EFS, event-free survival) can reach 90%, but it is poor prognosis and low survival rate in B-ALL adults. In addition, in adult B-ALL patients, the effect of traditional chemotherapy is poor, and the mortality rate is about 60%. Therefore, there...

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Problems solved by technology

[0008] In 2009, an article entitled "Long–termcultureofprimaryhumanlymphoblasticleukemiacellsintheabsenceofserumorhematopoieticgrowthfactors" mentioned the in vitro medium of primary lymphocytic leukemia cells, which was a serum-free IMDM medium supplemented with cytokines IL-3, IL-7, and SCF; Although the medium does not contain serum, the inventors of the present invention conducted repeated experiments using the medium and found that the culture effect described in the article cannot be achieved, that is, its stability needs to be studied;

[0010] It should be noted that although serum is necessary for the survival and culture of traditional cells in vitro, when human primary cells are cultured in vitro, serum can accelerate the differentiation and aging of primary cells, especially hematopoietic stem cells and progenitor cells; Moreover, as far as the serum itself is concerned, even if it is the serum of the same brand, there may be obvious differences between different batches of serum, which makes the culture effect also have obvious differences, so the use of serum-containing medium has unclear medium components. The problem is that the practicability is relatively insufficient

[0011] However, co-cultivation with stromal cells is not only cumbersome, but also the growth rate of stromal cells (especially immortalized cell lines) far exceeds that of primary cells during the co-cultivation process, which competes with primary cells for nutrients in the medium. , thereby affecting the growth of primary cells; in addition, stromal cells activate the survival and expansion of primary cells in vitro through protein secretion or receptor binding, but these activation factors are uncontrollable with the cell state, differentiation, aging, and passage times of stromal cells. Factors are related, resulting in obvious differences in the culture effect of primary cells cultured by the stromal cell co-culture method, which is difficult to repeat and is not conducive to popularization and application

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