Method for extracting human bone marrow mesenchymal stem cells by using implant cavity bone fragments

Abstract

The invention relates to a method for extracting human bone marrow mesenchymal stem cells by using implant cavity bone scraps, which comprises the following steps of: aseptically collecting the implant cavity bone scraps attached to the walls of a reamer and a tapping drill, immediately putting the implant cavity bone scraps into an alpha-MEM culture solution, and sealing; transferring the crushed bone scraps in the centrifugal tube together with the culture solution into a culture dish in a super clean bench, and incubating in a constant-temperature incubator; after 3 days, half of the fresh culture solution is replaced for the first time, and then full of the culture solution is replaced every 3 days, and non-adherent cells are removed; after the long fusiform cells are observed to climb to the periphery by taking the bone scraps as the center, replacing the culture solution with an alpha-MEM culture solution for continuous culture; when the cells climb to the dish bottom and reach 80% of density, adding 1 mL of 0.25% trypsin/EDTA for digestion, performing passage at 1: 3, recording as P1, and performing passage at 1: 3 to P3, so as to obtain the human mesenchymal stem cells.

Description

technical field [0001] The invention belongs to the field of medicine, and in particular relates to a method for extracting human bone marrow mesenchymal stem cells by using bone scraps in planting holes. Background technique [0002] Mesenchymal stem cells have self-renewal ability, multi-directional differentiation potential and immunoregulatory properties, and are widely used in experimental research in the fields of tissue defect repair, immune regulation, gene therapy, and drug screening. Mesenchymal stem cells can be isolated from various tissues, such as umbilical cord, endometrial polyps, menstrual blood, bone marrow, adipose tissue, etc. [Ding D-C, Shyu W-C, Lin S-Z. Mesenchymal Stem Cells [J]. Cell Transplantation, 2011 , 20(1): 5-14.]. Among them, bone marrow-derived mesenchymal stem cells are easily isolated and can differentiate into various cell subsets such as osteoblasts, adipocytes, chondrocytes, and nerve cells [Sasaki H, Takeuchi I, Okada M, et al. Labe...

AI Technical Summary

Problems solved by technology

These extraction methods are cumbersome, and the density gradient centrifugation method and the adherence method all involve multiple filtration steps and centrifugation steps, which greatly increase the probability of contamination and the loss of cells, and a large number of red blood cells and stem cells are mixed together after centrifugation. Affects the attachment behavior of stem cells, which may reduce the proportion of stem cells in primary cells

In addition, the technical sensitivity of extraction using magnetic bead sorting method is high, and individual operation differences have a greater impact on the extraction results

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