Preparation method of double-layer cell ball

A production method and technology of cell spheroids, applied in biochemical equipment and methods, microorganisms, tissue culture, etc., can solve the problems that cannot truly reflect the living environment of cells

Inactive Publication Date: 2011-11-23
ZHEJIANG UNIV
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Problems solved by technology

Tumor cells are always in close contact with the surrounding mesenchymal cells during the development process. When the effect factors in the local microenvironment reach a certain concentration, the interaction between the two can lead to a series of structural, functional and metabolic changes in tumor cells. Therefore, this change cannot be ob...

Method used

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Embodiment Construction

[0015] In this embodiment, the production of double-layered cell spheres of tumor cells and normal cells is taken as an example, and the specific steps are as follows:

[0016] a. Adopt RPMI1640 containing 10% fetal bovine serum as culture medium (i.e. 100 milliliters of culture medium contains 10 milliliters of fetal bovine serum and 90 milliliters of RPMI1640), and culture normal cells and tumor cells respectively according to conventional culture methods; DMEM can also be used Or α-MEM instead of RPMI1640.

[0017] b. Select one of the cells to make a single cell suspension, that is, pour out the culture medium in the culture bottle, add 1-2ml PBS (Phosphate-buffered saline or phosphate buffer solution, the Chinese name is phosphate buffer saline, and mix the PBS powder according to the instructions. Dissolve in double-distilled water for high-pressure disinfection, and prepare a phosphate buffer solution with a concentration of 0.01M and a pH of 7.2-7.4) gently wash it for...

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Abstract

The invention relates to a preparation method of a double-layer cell ball. The invention aims at providing a preparation method of the double-layer cell ball, comprehensively and truly reflecting the survival environment of in-vivo cells, and visually reproducing the interaction between two kinds of cells under certain conditions. The technical schemes for achieving the aims are as follows: the preparation method of the double-layer cell ball comprises the steps of: a. separately culturing two different kinds of cells; b. preparing a single cell suspension from one kind of the cells; c. controlling the single cell concentration in a bottle at 1*10<5>-1*10<6> cells/ml; d. standing the single cell suspension in a 37 DEG C incubator containing 5% of CO2 for 1-4 hours; e. carrying out rotation culture on the single cell suspension for rolling the single cell suspension into balls, and changing the single cell suspension once every 10-14 hours; and f. preparing a single cell suspension of the other kind of cells, and adding the single cell suspension to the culture liquid containing the cell balls obtained in step e, continuing rotation culture so that the two kinds of cells in a tube form a spherical cell mass with a clear boundary and a double-layer structure, and changing the suspension once every 10-14 hours. The preparation method of the double-layer cell ball provided by the invention is mainly used for three-dimensional culture of cells.

Description

technical field [0001] The invention relates to a method for making double-layer cell spheres, which is mainly suitable for three-dimensional culture of cells. Background technique [0002] At present, the traditional culture of adherent cells is a two-dimensional culture model, a model of cell monolayer culture, which lacks the support of the microenvironment in vivo, and the experimental results obtained cannot be consistent with the actual situation in vivo. Since the parenchymal cells are in close contact with the surrounding mesenchymal cells in the body, there is an interaction between them. The normal interaction can maintain the homeostasis and the normal operation of the body, while the abnormal interaction can lead to diseases, such as tumors. Studies have found that the occurrence of tumors is not only caused by tumor cells, its local microenvironment also plays a vital role, and even determines the fate of tumors. Tumor cells are always in close contact with the...

Claims

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Application Information

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IPC IPC(8): C12N5/02
Inventor 邓红赵志敏许晶晶饶翠来茂德
Owner ZHEJIANG UNIV
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