Human placenta chorionic mesenchymal stem cell separation method

A technology of mesenchymal stem cells and separation methods, applied in the field of human placental chorionic mesenchymal stem cells, can solve the problems of decreased differentiation ability and cell number, strong immune rejection of allogeneic transplantation, and high chance of virus infection, etc., to reduce manual processing time and manpower consumption, the effect of increasing the area of ​​tissue adhering to the wall and reducing the chance of contamination

Active Publication Date: 2017-11-17
GUANGZHOU WOMEN AND CHILDRENS MEDICAL CENTER
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] At present, the mesenchymal stem cells used are mainly bone marrow mesenchymal stem cells, and their source is convenient and fast. As the age increases, the expansion, differentiation ability and cell number show an obvious downward trend; 2. There are ethical issues and the source is limited; 3. The donor feels uncomfortable and i

Method used

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  • Human placenta chorionic mesenchymal stem cell separation method
  • Human placenta chorionic mesenchymal stem cell separation method
  • Human placenta chorionic mesenchymal stem cell separation method

Examples

Experimental program
Comparison scheme
Effect test

Example Embodiment

[0046] Example 1:

[0047] 1) Take human placental chorionic tissue, cut it into 1~3cm block size, and wash it repeatedly with normal saline until there is no blood;

[0048] 2) Use a handheld electric homogenizer to treat the chorionic membrane to 0.1~0.3cm 3 For small particles, rinse again with saline to clear;

[0049] 3) Treat again to 0.1~0.5mm with a homogenizer 3 After the small particles are left and right, centrifuge 300r / min for 5min to remove the upper blood cells;

[0050] 4) Add the same volume of Trypsin and Collagenase II to the precipitate, digest at 37°C for 0.5h;

[0051] 5) After centrifugation and washing of the digested material with saline twice, add culture medium to the pellet and place it at 37℃, 5% CO 2 Culture in an incubator;

[0052] 6) On the 7th day, discard the tissue pieces and culture medium, rinse the bottom of the culture flask with normal saline, add fresh medium, then change the medium every 3 days until the cell confluence reaches 80-90%, and use t...

Example Embodiment

[0053] Example 2:

[0054] 1) Take human placental chorionic tissue, cut it into 1~3cm block size, and wash it repeatedly with normal saline until there is no blood;

[0055] 2) Use a handheld electric homogenizer to treat the chorionic membrane to 0.1~0.3cm 3 For small particles, rinse again with saline to clear;

[0056] 3) Treat again to 0.1~0.5mm with a homogenizer 3 Centrifuge the tiny particles at 500r / min for 5min to remove the upper blood cells;

[0057] 4) Add the same volume of Trypsin and Collagenase II to the precipitate, digest at 37°C for 1 hour;

[0058] 5) After centrifugation and washing of the digested material with saline twice, add culture medium to the pellet and place it at 37℃, 5% CO 2 Culture in an incubator;

[0059] 6) On the 7th day, discard the tissue pieces and culture medium, rinse the bottom of the culture flask with normal saline, add fresh medium, then change the medium every 3 days until the cell confluence reaches 80-90%, and use trypsin digestion and p...

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Abstract

The invention discloses a human placenta chorionic mesenchymal stem cell separation method, which comprises: 1) taking healthy human placenta chorionic tissue, cutting into strip blocks, and completely rinsing with physiological saline; 2) adding a proper amount of physiological saline, chopping the chorion into fine particles by using a homogenizer, and rinsing with physiological saline until the solution is clarified; 3) treating again with the homogenizer to achieve micro-particles, and carrying out centrifugation to remove the upper layer blood cells; 4) adding an enzyme to the precipitate, digesting, carrying out centrifugation, and separating the precipitate; 5) carrying out centrifugal washing with physiological saline twice, adding a culture medium to the precipitate, inoculating, and culturing; and 6) discarding the tissue blocks and the culture medium at the 7th day, rinsing the bottom of the culture flask by using physiological saline, adding a fresh culture medium, changing the culture medium every 3 days until the cell fusion degree achieves 80-90%, and carrying out digestion passage with trypsin to obtain the human placenta chorionic mesenchymal stem cells. According to the present invention, the operation is simple and rapid, and the high-quality human placenta chorionic mesenchymal stem cells can be obtained.

Description

technical field [0001] The invention belongs to the technical field of biomedicine, and in particular relates to a method for separating human placental chorionic mesenchymal stem cells. Background technique [0002] Mesenchymal stem cells are multipotent adult stem cells commonly found in different tissues. Mesenchymal stem cells have been isolated and cultured from bone marrow, fat, dental pulp, amniotic membrane, chorion, decidua, umbilical cord, umbilical cord blood and other tissues. Studies have shown that mesenchymal stem cells have strong differentiation ability, short doubling time, immunoregulatory effect, low immunogenicity, and easy transfection characteristics. They are an ideal seed cell and gene therapy carrier for regenerative medicine. Stem cells are a hotspot in clinical transformation application research, but how to easily and quickly isolate a large number of primary mesenchymal stem cells in vitro is the basis for mesenchymal stem cell research and ap...

Claims

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Application Information

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IPC IPC(8): C12N5/0775C12N5/0735
CPCC12N5/0605C12N5/0668C12N2509/00
Inventor 吴韶清欧志英金宇林李发涛唐婕吴洁莹陈劲松谢闺娥陆琰梁绮华刘东
Owner GUANGZHOU WOMEN AND CHILDRENS MEDICAL CENTER
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