Method for isolation and serum gradient switching culture of human umbilical cord mesenchymal stem cells

Abstract

The invention discloses a method for isolation and serum gradient switching culture of human umbilical cord mesenchymal stem cells. The method comprises the following steps: digesting human umbilical cord with collagenase I until Wharton's jelly is fully digested, collecting digested single cells, inoculating the single cells into alpha-MEM medium, carrying out suspension culture until 80-90% of cells are fused, and digesting with trypsin; inoculating the suspension of trypsin-digested cells into alpha-MEM medium, carrying out suspension culture until 80-90% of cells are fused, digesting with trypsin, subcuturing and inoculating the subcultured cells into alpha-MEM medium, and carrying out subculture repeatedly. The concentration of fetal bovine serum in alpha-MEM medium in the steps of subculture decreases with the increase of the passage number until fetal bovine serum disappears. According to the method, the material human umbilical cords are easily accessible; the preparation of human umbilical cord mesenchymal stem cells is rapid and safe, and is not subject to ethical restrictions; and the subsequent culture is independent on the characteristics of fetal calf serum, resulting in greatly reduced culture cost and risk in clinical use. The method of the invention has a good application prospect.

Description

Technical field: [0001] The invention belongs to the field of biomedicine, and in particular relates to a method for separating and switching culture of human umbilical cord mesenchymal stem cells. Background technique: [0002] In recent years, stem cell research has made major breakthroughs. In 1999 and 2000, the world's most authoritative American magazine "Science" listed stem cells and the Human Genome Project as the top 10 scientific breakthroughs of the year for two consecutive years, especially in 1999, it even listed stem cell research as the first. Stem cells are likely to lead to revolutionary progress in the medical field, and thus have immeasurable medical value and have attracted widespread attention and research around the world. The American "Time" magazine believes that stem cells and the Human Genome Project will simultaneously become the most promising development and application in the new century. field. [0003] Mesenchymal stem cells (MSCs for short)...

AI Technical Summary

Problems solved by technology

Many research institutions have found pluripotent stem cells in other tissues, including mobilized peripheral blood, cord blood, deciduous teeth, and umbilical cord mesenchymal cells, however, the number of cells obtained from these tissues is very limited

[0004] At present, the medium supplemented with fetal bovine serum is considered to be the most effective medium for the expansion of mesenchymal stem cells in vitro. However, the composition of the serum is not clear, and it may contain known or unknown pathogens carried by animals. In clinical applications Potentially dangerous, and fetal bovine serum is expensive

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