Cell culture layer and application in culture of human primary cancer cells thereof

Abstract

The invention discloses a cell culture layer which is prepared by the following methods: culturing BALB3T3 cells by adopting a DMEM culture medium, performing cell adherent culture until 70-80 percent of a culture bottle is full, and irradiating; performing X-ray irradiation by taking 3000cGy as a radiation dose; culturing by using an improved F culture medium after irradiation; culturing for 8 hours, and obtaining the culture, namely the cell culture layer. The cell culture layer can be used for culturing human primary cancer cells or cancer stem cells. The invention also discloses improvement of cell types of the cell culture layer as well as a preparation method of the cell culture layer. Compared with J2 cells, the 3T3 cells for preparing the culture layer are easily obtained, and the cell concentration and state can be easily controlled in the culture process. When the cell culture layer is used for culturing the human primary cancer cells or cancer stem cells, growth of the primary cancer cells can be well promoted, and human cancer stem cells in a higher ratio are obtained (the number of CD44 positive cells is improved by about 80 percent).

Description

technical field [0001] The invention relates to a cell feeder layer, a preparation method thereof, and an application in culturing human primary tumor cells or tumor stem cells. Background technique [0002] At present, the commonly used methods of isolation and culture of human tumor stem cells mostly adopt the serum-free suspension culture method, which isolates and cultures stem cells with very small content from cell lines or directly purchases stem cell lines for suspension culture, while the method of adherent culture of primary tumor cells It is difficult to successfully isolate and culture human tumor stem cells. [0003] The use of feeder layer in cell culture: There are foreign reports that use J2 (a subtype of 3T3 cell) cells as a feeder layer for primary culture of tumor cells, but there is no mention of using it for tumor stem cell culture. Moreover, the concentration and state of J2 cells need to be strictly controlled in the process of serving as a feeder lay...

AI Technical Summary

Problems solved by technology

[0002] At present, the commonly used methods of isolation and culture of human tumor stem cells mostly adopt the serum-free suspension culture method, which isolates and cultures stem cells with very small content from cell lines or directly purchases stem cell lines for suspension culture, while the method of adherent culture of primary tumor cells Difficult to successfully isolate and culture human tumor stem cells

[0003] The use of feeder layer in cell culture: There are foreign reports using J2 (a subtype of 3T3 cell) cells as a feeder layer for primary culture of tumor cells, but there is no mention of using it for tumor stem cell culture

Moreover, the concentration and state of J2 cells need to be strictly controlled in the process of serving as a feeder layer, which is not easy to operate, and is currently not available in China.

[0004] Currently commonly used feeder layer preparation methods: Co60 or mitomycin is often used to treat cells to prepare feeder layers, which has the following defects: it is difficult to find Co60 radioactive sources in China at present; use mitomycin to treat feeder layers, different batches of silk from different It is difficult to grasp the titer concentration of lysomycin, and there are drug-resistant cells, and even cause malignant transformation of fibroblasts

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