Method for extracting hematopoietic stem cell from placental chorionic tissue and construction method of hematopoietic stem cell

A technology of hematopoietic stem cells and chorion is applied in the field of building hematopoietic stem cell banks and extracting CD133 positive stem cells. A large number of effects

Inactive Publication Date: 2012-10-31
BEIJING HEALTH & BIOTECH (H&B) CO LTD
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  • Abstract
  • Description
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  • Application Information

AI Technical Summary

Problems solved by technology

[0007] At present, the methods for isolating placental stem cells include mechanical separation, lavage, and collagenase digestion. The mechanical separation causes great damage to the cells and is time-consuming and laborious for separation, which is not conducive to industrialization. The lavage method is mainly used for hemangioblasts, and the cells are obtained. The rate is low; collagenase digestion method Neizhou victory, etc.
[0008] The new method of extracting hematopoietic stem cells from placental tissue for the establishment of hematopoietic stem cell bank (Patent No. 01131190.8) mentions that CD34-positive cells from tissue are mainly sorted, and a single collagenase is used to digest placental tissue for 15 minutes, but The above method can only obtain a very small number of cells, and the number of cells is far lower than that obtained by mechanical separation.

Method used

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  • Method for extracting hematopoietic stem cell from placental chorionic tissue and construction method of hematopoietic stem cell
  • Method for extracting hematopoietic stem cell from placental chorionic tissue and construction method of hematopoietic stem cell
  • Method for extracting hematopoietic stem cell from placental chorionic tissue and construction method of hematopoietic stem cell

Examples

Experimental program
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Effect test

Embodiment 1

[0025] Example 1: Isolation of placenta-derived cells

[0026] 1. Placenta collection, the placenta was collected with the informed consent of the parturient. Wash the placenta with sterile normal saline, then put the placenta into a special aseptic collection box, and add the collection solution. The placenta was then transported to the experimental site at 4°C, and the time for transporting the placenta did not exceed 48 hours.

[0027]2. Isolation of hematopoietic stem cells from placental chorionic tissue: (1) Separate the chorionic tissue by mechanical means; (2) Cut the tissue to 0.1-1 cubic centimeters, and then wash the tissue block repeatedly with phosphate solution for more than 5 times , so that the tissue block has no obvious blood clots and other sundries; (3) add collagenase, trypsin, hyaluronidase and DNase in a ratio of 1:1 to the volume of the tissue, and digest at a constant temperature of 37°C for 60 Minutes; (4) Add hydroxyethyl starch and digestive juice...

Embodiment 2

[0031] Sorting of CD133-positive cells derived from placental chorion:

[0032] The sorting method is the same as in Example 1, only the source is changed to placental chorionic tissue. The sorted cells were sorted out from placenta-derived cells by using a magnetic bead sorting method well known to professionals to separate CD133-positive cells.

[0033] figure 2 The results shown: After sorting by CD133 magnetic beads, the purity of CD133 positive cells obtained was 93.07% of the total cells.

Embodiment 3

[0035] Differentiation of placental chorion-derived CD133-positive cells into endothelial cells

[0036] image 3 The results shown: the CD31-positive ratio of cells cultured with mesenchymal stem cell culture medium (middle diagram) was 2.85%, and the CD31-positive ratio of cells cultured with endothelial cell culture medium (right diagram) was 74.36%.

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Abstract

The invention discloses a method for extracting a CD133 positive stem cell from a placental chorionic tissue. The method comprises the following steps: (1) collecting a placenta to a placenta collection box, and adding protection fluid into the collection box; (2) carrying out information registration and serological virus detection before the placenta separates the stem cell; (3) washing blood clot and hematocele in the placenta tissue; (4) separating the placenta stem cell by a mechanical method; (5) concentrating and purifying the placenta stem cell, and extracting the CD133 positive stem cell from the placenta; and (6)storing the CD133 positive stem cell. The stored CD133 positive stem cell can be used for the transplanting of a hematopoietic stem cell, angiogenesis, repairing of nerve injury and the like. The invention has the advantages that the CD133 positive stem cell extracting flow is convenient; theCD133 positive stem cell extracting cost is lower; and a whole placenta can be treated rapidly at a time, the placenta contains a large number of mononuclear cells, and the content of CD133 positive stem cells in the placenta reaches 2.0 to 5.0 percent. The technology is also applicable to constructing a CD133 positive stem cell bank, and the CD133 positive stem cells stored in the cell bank can be reused by adults for many times and stored for a long time, and are convenient to use at any time.

Description

technical field [0001] The invention relates to a method for extracting hematopoietic stem cells from placental chorionic tissue, in particular to a method for extracting CD133 positive stem cells from placental chorionic tissue, and a method for using the extracted CD133 positive stem cells to construct a hematopoietic stem cell bank. Background technique [0002] The stem of a hematopoietic stem cell (HSC), translated from English "stem", means "tree", "stem" and "origin". Similar to a tree trunk can grow branches, leaves, and blossom and bear fruit. In layman's terms, hematopoietic stem cells refer to immature cells, which are the origin of all hematopoietic cells and immune cells. They can not only differentiate into red blood cells, white blood cells and platelets, but also differentiate into cells of various tissues and organs across systems. Renewal, multilineage differentiation, and homing (ie, directed migration to hematopoietic tissues and organs) potential. Gene...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/0735C40B50/06
Inventor 韩忠朝王涛邵元康
Owner BEIJING HEALTH & BIOTECH (H&B) CO LTD
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