Method for preparing human adipose-derived mesenchymal stem cell exosomes

Abstract

The invention relates to a method for preparing human adipose-derived mesenchymal stem cell exosomes. The method comprises the following steps: adding a basic culture medium into stem cells, performing centrifuging, re-suspending a cell precipitation, and inoculating the cells into a culture flask and performing culturing for 72 h; then sub-culturing the adipose-derived mesenchymal stem cells after 72 h of culture with a culture medium made of the basic medium, a serum substitute and other additives, and performing culturing for 72 h; transferring the culture flask to a biological safety cabinet, and collecting a supernatant; performing ultrafiltration and concentration to obtain a first-level concentrated solution; and centrifuging the first-level concentrated solution by using a 30% sucrose density gradient, and collecting a concentrated solution. The method of the invention can effectively reduce the possibility of cells being infected by bacteria during the culture process, and atthe same time can promote the proliferation of the stem cells and the secretion of the exosomes.

Description

technical field [0001] The invention relates to the technical field of mesenchymal stem cell exosomes, in particular to a method for preparing human adipose mesenchymal stem cell exosomes. Background technique [0002] Exosomes are tiny vesicles secreted by a variety of cells under normal or pathological conditions, with a diameter of 30-150 nm and a lipid bilayer membrane structure. Studies have shown that stem cell-derived secretions and target cells can transport biologically active substances to target cells through membrane fusion or endocytosis, mediate intercellular communication and affect target cell functions. In addition, secretions can also inhibit inflammation, immune Regulatory, angiogenesis, functions involved in tissue repair. Compared with stem cells, stem cell exosomes are more stable, safer and more convenient to store. [0003] The Chinese patent with the existing application publication number CN109880797A discloses a method for preparing exosomes from...

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Problems solved by technology

[0004] The medium in the above-mentioned published patent contains fetal bovine serum, which is easily infected by bacteria and mycoplasma, and then infects stem cells. In addition, in the process of cell culture, even if the staff carefully operates according to the experimental rules, and the experimental equipment is well-equipped However, during the experiment, cells were still contaminated by bacteria and stopped proliferating or secreting exosomes, resulting in lower content and purity of the prepared exosomes

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