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Extraction and preparation method of placenta pluripotent stem cells

A technology of pluripotent stem cells and an extraction method, which is applied in the field of extraction and preparation of placental pluripotent stem cells, can solve the problems of immune response, high cost, and a small number of stem cells, and achieves the effect of high separation efficiency

Inactive Publication Date: 2016-09-28
ANHUI HUIEN BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the number of stem cells that can be extracted from the umbilical cord is rather small, making the process lengthy and expensive
In addition, stem cells isolated from the umbilical cord are not compatible with each recipient's immune system, which can cause an immune response leading to transplant rejection

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0018] Prepare D-Hanks solution: 800ml of freshly prepared triple distilled water, add 8.00g of sodium chloride, 0.40g of potassium chloride, 0.06g of disodium hydrogen phosphate, 0.06g of potassium dihydrogen phosphate, 0.35g of sodium bicarbonate, phenol red 0.02g, after fully dissolved, fill it with double-distilled water to 1000ml, measure pH 7.2-7.4, dispense into 250ml glass bottles, autoclave for 20min, cool at room temperature, store at 4°C, and heat to 37°C when used.

[0019] Prepare trypsin solution: Weigh 250mg trypsin and 20mg EDTA; take 80ml D-Hanks solution, dissolve trypsin completely, add D-Hanks solution to 100ml, and add EDTA solution to dissolve, the final concentration is 0.25% trypsin-0.02 %EDTA solution, sterilized by suction with a 0.1um filter and placed in a 4°C refrigerator for later use.

[0020] Preparation of type IV collagenase: Take 0.2ml of collagenase type IV stock solution, add D-Hanks solution to 100ml, use a 0.1um filter to filter and steri...

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Abstract

The invention discloses an extraction and preparation method of placenta pluripotent stem cells. The extraction and preparation method comprises the following steps: under a sterile condition, obtaining placenta tissues; repeatedly washing with PBS, disinfecting with betadine and sufficiently washing a placenta with a saline solution for at least three times; cutting the placenta tissues into fragments with the size of 1*1*1mm<3> and putting the fragments into a sterile centrifugal tube; digesting the placenta tissue fragments with 0.25% trypsase and 0.2% IV type collagenase; centrifuging and collecting to obtain cell sediment; re-suspending the cell sediment into a fetal calf blood serum alpha-MEM culture medium with the volume percent of 10% and freezing by a conventional method; and carrying out flow cytometry analysis on CD73, CD90 and CD105 cell expression. The placenta is obtained under the sterile condition and the placenta is subjected to a series of pre-treatment including PBS repeated washing, placenta surface disinfection and washing of the placenta with the saline solution, so as to guarantee that the obtained placenta is fresh and conveniently guarantee the activity of the pluripotent stem cells in the placenta tissues.

Description

technical field [0001] The invention belongs to the field of biotechnology, and relates to a method for extracting and preparing placental pluripotent stem cells. Background technique [0002] Stem cells are self-renewing and capable of differentiating into various cell lines. These cells can be divided into embryonic stem cells (ESCs) and non-embryonic or adult stem cells (ASCs). The latter cannot redifferentiate into various tissues (pluripotent) and proliferate at a slower rate than embryonic stem cells. Therefore, the applicability of embryonic stem cells is better than that of adult stem cells. Preliminary studies in animals and humans suggest that transplantation of stem cells grown outside the body may alleviate symptoms of diseases such as diabetes, Parkinson's disease, liver failure and congestive heart failure. However, progress in this field has been slow due to a number of reasons, such as the ethical concerns that accompany the use of human embryonic stem cel...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/073
CPCC12N5/0605C12N2509/00
Inventor 张正亮
Owner ANHUI HUIEN BIOTECH
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