Separation culture method of human mesenchymal stem cells by density gradient centrifugation method

Abstract

The invention provides a separation culture method of bone marrow mesenchymal stem cells (BMSCs) by a density gradient centrifugation method. The separation culture method comprises the steps of BMSCs separation, BMSCs primary culture, BMSCs subculture and amplification, cultured cell surface antigen detection and growing curve determination. According to the method, a density gradient centrifugation method is adopted, a lymphocyte separation medium with density of 1.077 is used for separating BMSCs, and most erythrocyte, adipocyte, soterocyte and the like are removed after gradient centrifuge to obtain BMSCs cells with relatively high purity; and the BMSCs cells are cultured for 2 to 3 days to find that the BMSCs basically grow by adhering walls according to the characteristic that the BMSCs grows by adhering to walls; therefore, BMSCs with good uniformity can be obtained in the final phase of primary culture. The method has the advantages of simpleness in operation, quickness and practicality and the like, and is an effective BMSCs separation purification method.

Description

technical field [0001] The invention relates to a method for isolating and culturing human bone marrow mesenchymal stem cells in the field of biotechnology, in particular to a method for isolating and culturing human bone marrow mesenchymal stem cells by density gradient centrifugation. Background technique [0002] Bone marrow mesenchymal stem cells (Bone Marrow Stromal Cells, BMSCs), also known as bone marrow stromal stem cells, is another type of stem cells that exist in the bone marrow. Since Friendenstein used the adhesion characteristics of BMSCs to isolate, culture and study their multi-differentiation potential in 1968, BMSCs have become a hot spot in the fields of modern biology and medicine. It is now generally believed that BMSCs are dry-stage cells of mesoderm development and have multi-directional differentiation potential. They can not only differentiate into hematopoietic parenchyma and support hematopoiesis, but also differentiate into a variety of tissue cel...

AI Technical Summary

Problems solved by technology

The adherent separation and screening method is based on the characteristics of BMSCs adherent growth and hematopoietic cell suspension growth, but the obtained cell components are complex, and the acquisition rate of BMSCs is low; flow cytometry separation and immunomagnetic bead separation experiments The conditions are relatively high, and the amount of bone marrow required is large. However, in actual operation, the amount of bone marrow extracted from each part should generally not exceed 2ml, so it cannot meet the amount of bone marrow required for flow cytometry experiments.

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