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Method for preparing human amniotic membrane epithelial cells from human placental amniotic membrane and application thereof

A technology of epithelial cells and human amniotic membrane, applied in the field of stem cell therapy, can solve problems affecting cell separation and culture, high contamination rate of human amniotic membrane collection, poor cell activity, etc.

Active Publication Date: 2019-07-26
BOYALIFE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The invention solves the problems in the prior art that there is a high contamination rate of human amniotic membrane collection and poor cell activity after collection, which directly affects subsequent cell separation and culture

Method used

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  • Method for preparing human amniotic membrane epithelial cells from human placental amniotic membrane and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0076] Example 1: Preparation of human amniotic epithelial cells from placental amniotic membrane

[0077] (1) Take out the placenta from the placenta sample collection box, place it in a white porcelain dish, use a basic balanced salt solution (KH2PO4 of 0.4g, KH2PO4 of 0.06g, Na2HPO4.12H2O of 0.132g, NaCl of 8g, 0.35g of NaHCO3, the D-glucose of 1.0g, the streptomycin of 0.10g, the penicillin of 0.06g are dissolved in water and be settled into the solution of 1L, obtain) repeatedly wash the surface, and the placenta is disinfected;

[0078] (2) Slowly tear the outer layer of amniotic membrane with surgical forceps and place it in a 150mm glass dish, use basic balanced salt solution to repeatedly clean the surface of the blood, and cut it into small pieces of amniotic membrane tissue with a diameter of 5mm;

[0079] (3) Small pieces of amniotic membrane are evenly pasted on a 100mm petri dish with the amniotic membrane epithelium facing down. After drying for 30-60 minutes ...

Embodiment 2

[0083] Example 2: Preparation of human amniotic epithelial cells from placental amniotic membrane

[0084] (1) Take out the placenta from the placenta sample collection box, place it in a white porcelain dish, use a basic balanced salt solution (KH2PO4 of 0.4g, KH2PO4 of 0.06g, Na2HPO4.12H2O of 0.132g, NaCl of 8g, 0.35g of NaHCO3, the D-glucose of 1.0g, the streptomycin of 0.10g, the penicillin of 0.06g are dissolved in water and be settled into the solution of 1L, obtain) repeatedly wash the surface, and the placenta is disinfected;

[0085] (2) Slowly tear the outer layer of amniotic membrane with surgical forceps and place it in a 150mm glass dish, use basic balanced salt solution to repeatedly clean the surface of the blood, and cut it into small pieces of amniotic membrane tissue with a diameter of 5mm;

[0086] (3) Small pieces of amniotic membrane are evenly pasted on a 100mm culture dish with the amniotic membrane epithelial layer facing down. After drying for 30-60 ...

Embodiment 3

[0090] Example 3: Preparation of human amniotic epithelial cells from placental amniotic membrane

[0091] (1) Take out the placenta from the placenta sample collection box, place it in a white porcelain dish, use a basic balanced salt solution (KH2PO4 of 0.4g, KH2PO4 of 0.06g, Na2HPO4.12H2O of 0.132g, NaCl of 8g, 0.35g of NaHCO3, the D-glucose of 1.0g, the streptomycin of 0.10g, the penicillin of 0.06g are dissolved in water and be settled into the solution of 1L, obtain) repeatedly wash the surface, and the placenta is disinfected;

[0092] (2) Slowly tear the outer layer of amniotic membrane with surgical forceps and place it in a 150mm glass dish, use basic balanced salt solution to repeatedly clean the surface of the blood, and cut it into small pieces of amniotic membrane tissue with a diameter of 5mm;

[0093] (3) Small pieces of amniotic membrane are evenly pasted on a 100mm petri dish, with the amniotic membrane epithelial layer facing down, and after drying for 30-...

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Abstract

The invention relates to a method for preparing human amniotic membrane epithelial cells from human placenta amnion and application thereof, in particular to the method for preparing human amniotic membrane epithelial cells from the placenta amnion. The method comprises the following steps that the surface of a placenta is flushed with a basic balanced salt solution repeatedly and the placenta is sterilized; a foetal membrane outer layer amnion is torn and taken into a glass dish by surgical forceps, the grimy blood on the surface of the foetal membrane outer layer amnion is flushed with the basic balanced salt solution, and then the foetal membrane outer layer amnion is sheared into small blocks; the small amnion blocks are evenly attached to a culture dish, the epithelial layer of the amnion faces downwards, and after drying, a complete medium is added for culture; fluid infusion is conducted two days later, and then culture is continued; when epithelial cells are seen to crawl out, a tissue is removed, and medium change is conducted to continue culture; after multiple typical epithelial cell masses occur, the local fusion rate reaches 90%, and continuous cell culture is conducted; and harvesting is conducted, specifically, digestion is conducted by pancreatic enzymes, counting and freezing storage are conducted, and the human amniotic membrane epithelial cells are obtained. The invention further relates to the prepared human amniotic membrane epithelial cells and the application thereof. The method has the advantages showing in the specifications.

Description

technical field [0001] The invention belongs to the technical field of stem cell therapy, and relates to a method for preparing epithelial cells from human placenta amniotic membrane, human amniotic membrane epithelial cells prepared by the method, and their therapeutic application. Background technique [0002] Stem cells are the basis for the development of regenerative medicine, and their effective application is an effective way to realize the ultimate dreams of human health and longevity, anti-cancer and anti-aging, eternal youth, and continuous improvement of quality of life. In recent years, embryonic stem cells and adult stem cells have received much attention in the fields of cell biology and regenerative medicine research. However, despite the totipotency of embryonic stem cells that other cells cannot match, factors such as tumorigenicity, immune rejection after allogeneic transplantation, and unavoidable ethical disputes largely limit its clinical application. T...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/073A61K35/50
CPCA61K35/50C12N5/0605C12N2500/34C12N2500/46C12N2501/11
Inventor 许晓椿王正肖海蓉
Owner BOYALIFE
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