Method for extracting, culturing and purifying amniotic stem cells

A technology of stem cells and amniotic membrane, applied in cell dissociation methods, biochemical equipment and methods, embryonic cells, etc., can solve the problems of inability to obtain amniotic membrane stem cells, complicated extraction and culture methods, etc., to improve extraction and purification efficiency and ensure activity Effect

Inactive Publication Date: 2018-06-01
宁波东钱湖旅游度假区靖芮医疗美容诊所有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the existing methods for extracting and culturing amniotic membrane stem cells are still rel

Method used

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Examples

Experimental program
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Effect test

Embodiment 1

[0030] A method for extracting, culturing and purifying amniotic membrane stem cells, the specific preparation steps are as follows:

[0031] S1: Material preparation: take 38±1 weeks of normal cesarean section placenta without hepatitis, syphilis, HIV and other infectious diseases, and the placenta is obtained within 4 hours after cesarean section, and then mechanically separate the amniotic membrane and Separated from the placenta, at the same time, remove a large piece of amniotic membrane around the umbilical cord, then put the removed amniotic membrane into a PBS container containing double antibodies and wash repeatedly to remove blood and debris;

[0032] S2: Extraction of amniotic membrane stem cells:

[0033] (1) Add the amniotic membrane piece in S1 directly into a container containing a homogeneous mixture of 0.25% trypsin and 0.02% EDTA digestion solution without cutting it into pieces, and place it in a 37°C incubator for 0.5h;

[0034] (2) Scrape off the amnion ...

Embodiment 2

[0043] A method for extracting, culturing and purifying amniotic membrane stem cells, the specific preparation steps are as follows:

[0044] S1: Material preparation: take 38±1 weeks of normal cesarean section placenta without hepatitis, syphilis, HIV and other infectious diseases, and the placenta is obtained within 4 hours after cesarean section, and then mechanically separate the amniotic membrane and Separated from the placenta, at the same time, remove a large piece of amniotic membrane around the umbilical cord, then put the removed amniotic membrane into a PBS container containing double antibodies and wash repeatedly to remove blood and debris;

[0045] S2: Extraction of amniotic membrane stem cells:

[0046] (1) Add the amnion slices in S1 directly into a container containing a homogeneous mixture of 0.25% trypsin and 0.02% EDTA digestion solution without cutting them into pieces, and place them in a 37°C incubator for 1 hour;

[0047] (2) Scrape off the amnion cell...

Embodiment 3

[0056] A method for extracting, culturing and purifying amniotic membrane stem cells, the specific preparation steps are as follows:

[0057] S1: Material preparation: take 38±1 weeks of normal cesarean section placenta without hepatitis, syphilis, HIV and other infectious diseases, and the placenta is obtained within 4 hours after cesarean section, and then mechanically separate the amniotic membrane and Separated from the placenta, at the same time, remove a large piece of amniotic membrane around the umbilical cord, then put the removed amniotic membrane into a PBS container containing double antibodies and wash repeatedly to remove blood and debris;

[0058] S2: Extraction of amniotic membrane stem cells:

[0059] (1) Add the amnion slices in S1 directly into a container containing a homogeneous mixture of 0.25% trypsin and 0.02% EDTA digestion solution without cutting them into pieces, and place them in a 37°C incubator for 1.5h;

[0060] (2) Scrape off the amnion cells ...

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Abstract

The invention discloses a method for extracting, culturing and purifying amniotic stem cells. The method comprises S1: material preparation: taking a normal placenta which is acquired in 4h after cesarean section in 38+/-1 weeks and has no hepatitis, syphilis, HIV and other infectious diseases, mechanically separating an amniotic membrane from the placenta, simultaneously, taking off a large areaof the amniotic membrane around the umbilical region, putting the amniotic membrane into a double-antibody-containing PBS container and repeatedly washing the amniotic membrane to remove blood and fragments, and S2: amniotic stem cell extraction: (1) directly adding the amniotic membrane pieces into a container containing an uniform mixture of 0.25% of trypsin and 0.02% of an EDTA digestion solution without cutting, putting the container into an incubator at 37 DEG C and carrying out treatment for 0.5h. The method separates, extracts, cultures and purifies the amniotic membrane pieces in the placenta tissue and carries out in-vitro expansion multiplication culture so that the amniotic stem cells can be simply and conveniently acquired in large scale.

Description

technical field [0001] The invention relates to the technical field of stem cell extraction and culture, in particular to a method for extracting, culturing and purifying amniotic membrane stem cells. Background technique [0002] In recent years, with the development of stem cell technology, amniotic membrane stem cell transplantation has become the development direction of new disease treatment. In clinical application, amniotic membrane stem cells have carried out a lot of clinical work in ophthalmology such as conjunctival injury repair, ocular surface reconstruction, symblepharon prevention and retinal transplantation. However, the existing methods for extracting and culturing amnion stem cells are still relatively complicated, and amnion stem cells with high purity and activity cannot be obtained. [0003] Therefore, it is necessary to invent a method for extracting, culturing and purifying amniotic membrane stem cells to solve the above problems. Contents of the in...

Claims

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Application Information

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IPC IPC(8): C12N5/073
CPCC12N5/0605C12N2509/00
Inventor 孟小锐
Owner 宁波东钱湖旅游度假区靖芮医疗美容诊所有限公司
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