Separation and culture of mouse mesenchymal stem cells

Abstract

The invention provides a separation and culture method of mouse mesenchymal stem cells. The separation and culture method comprises the following steps: (1) an ICR mouse is anesthetized and killed; (2) thighbones and tibiae are taken and placed in ethyl alcohol of 75% to be soaked and disinfected; (3) washing is conducted for three times through PBS, and the epiphysis ends of the thighbones and the tibiae are cut off; (4) a culture medium containing double resistants is sucked through a needle tube of 10 ml to rush out bone marrow; (5) a percoll separating medium is used for centrifuging through a density gradient method, and a middle white layer is taken; (6) washing is conducted for two times through the PBS; (7) the cell activity is detected; (8) the cells are counted; (9) the cell morphology is identified; (10) detecting is conducted through an ELISA method; and (11) RT-PCR detection is conducted. The separation and culture method of the mouse mesenchymal stem cells is easy to operate, large in number of the obtained cells, high in survival rate and ideal, and provides the reliable cell resource for experiments.

Description

technical field [0001] The invention belongs to the technical field of cell culture of modern biotechnology, and specifically relates to a method for separating and culturing mouse bone marrow mesenchymal stem cells. . Background technique [0002] Bone marrow mesenchymal stem cells are important members of the stem cell family, derived from the mesoderm and ectoderm in the early stages of development. Bone marrow mesenchymal stem cells were originally discovered in the bone marrow, and have attracted increasing attention because of their multi-lineage differentiation potential, hematopoietic support and promotion of stem cell implantation, immune regulation and self-replication. For example, mesenchymal stem cells can be differentiated into fat, bone, cartilage, muscle, tendon, ligament, nerve, liver, heart muscle, endothelial and other tissue cells under specific induction conditions in vivo or in vitro, after continuous subculture and cryopreservation It still has multi...

AI Technical Summary

Problems solved by technology

[0004] The whole bone marrow method, also called the adherent method, is the earliest method for the isolation of bone marrow mesenchymal stem cells. The whole bone marrow method is a culture method in which all the flushing fluid washed out is centrifuged and washed directly, and then inoculated directly. Blood cells and other non-adherent cells are removed by changing the liquid. cells, this method is simple to operate, but the resulting cell components are complex and the cell purity is insufficient

Immunomagnetic bead method and flow cytometry method, the cells obtained are of high purity, but these two methods are difficult to operate and costly, and the sorting process also has a certain impact on the activity of cells

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