Isolation and cultivation method of prokaryotic cells of human umbilical cord blood mesenchyme stem cells

A technology of mesenchymal stem cells and culture methods, applied in the field of nuclear cell isolation and culture, dimethyl sulfoxide induction process, can solve the problems of lack of utilization, ethical and moral restrictions, etc.

Inactive Publication Date: 2012-07-04
江苏迈健生物科技发展股份有限公司
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Problems solved by technology

Neural stern cells (NSCs) derived from embryonic tissue were once considered to be a hope for the treatment of central nervous s

Method used

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  • Isolation and cultivation method of prokaryotic cells of human umbilical cord blood mesenchyme stem cells
  • Isolation and cultivation method of prokaryotic cells of human umbilical cord blood mesenchyme stem cells

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Embodiment Construction

[0011] Such as figure 1 Shown is a flow chart of the steps of the dimethyl sulfoxide induction process of human umbilical cord blood mesenchymal stem cells differentiated into neuronal cells of the present invention, and the specific steps of the method of the embodiment are as follows:

[0012] S1. Collection of umbilical cord blood and isolation of mononuclear cells: 40-60ml of umbilical cord blood from normal full-term cesarean section fetuses was collected under sterile conditions, anticoagulated with heparin at a concentration of 20U / ml, and all samples were isolated within 12 hours after collection. Dilute and mix with Hank'S balanced salt solution 1:1, superimpose on the Feoll-Hypaque, the relative density is 1.0779 / L, the column height ratio of the diluted blood to the Feoll-Hypaque solution is 2:1, centrifuge at 2000r / min for 20min, Take the mononuclear cells in the interface layer, add Hank'S balanced salt solution to centrifuge, wash twice, add to the medium, adjust...

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Abstract

The invention provides an isolation and cultivation method of prokaryotic cells of human umbilical cord blood mesenchyme stem cells, which is applied to a dimethyl sulfoxide induction process which enables the human umbilical cord blood mesenchyme stem cells to be differentiated into neuronal cells. The induction process uses serum-free dulbecco modified eagle medium (DMEM) containing dimethyl sulfoxide and butylated hydroxyanisole to induce the human umbilical cord blood mesenchyme stem cells to be differentiated into the neuronal cells. The isolation and cultivation method includes: collecting umbilical cord blood of a normal full-term cesarean section fetus under an aseptic condition, anti-freezing through heparin, separating single prokaryotic cell of the umbilical cord blood by using lymphocyte separation medium, cultivating and purifying by using partial acidic culture medium to obtain anchorage-dependent cells, obtaining application dimethyl sulfoxide which is amplified by a third generation to induce the mesenchymal stem cells (MSCs) to be differentiated towards the neuronal cells, and marking and displaying the induced MSCs in immunohistochemical mode to express a neurofilament protein (NF) and neuron specific endase (NSE). The human umbilical cord blood MSCs can be cultivated and amplified in vitro, can be differentiated towards neuron-like cells by using induction of the dimethyl sulfoxide, and provides a novel cell source for wound repair of central nervous systems.

Description

technical field [0001] The present invention relates to the field of biotechnology, in particular to a method for separating and culturing nuclear cells of human umbilical cord blood mesenchymal stem cells, which is applied to the dimethyl sulfoxide induction process of human umbilical cord blood mesenchymal stem cells differentiated into neuron cells . Background technique [0002] The injury and regeneration of the central nervous system (central nervous system, CNS) has long been a problem that plagues human health, and it is also the focus of scientists' research. Neural stern cells (NSCs) derived from embryonic tissue were once considered to be a hope for the treatment of central nervous system diseases, but their utilization was obviously limited due to the lack of sources and ethical constraints. In recent years, studies on bone marrow stromal stem cells that can produce skeletal muscle, cardiac muscle, liver cells, glial cells and neuron-like cells have shown that t...

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Application Information

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IPC IPC(8): C12N5/0775C12N5/0789C12N5/0793
Inventor 陆华
Owner 江苏迈健生物科技发展股份有限公司
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