Method for culturing cord blood type embryonic stem cells as well as identification and application

A technology of embryonic stem cells and umbilical cord blood, which is applied in the field of culturing stem cells, can solve the problems of tumorigenicity, research and application limitations, and low instability, and achieve the effects of improving the degree of adhesion, firm cell adhesion, and shortening the time

Active Publication Date: 2015-05-27
天晴干细胞股份有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Although the current research has confirmed that human embryonic stem cells can be induced to differentiate into islet-like cells in vitro, there are still many problems: 1) The host has immune rejection to hES differentiated cells; 2) The induction differentiation efficiency is low and the instability is low; 3 ) Maturity of differentiated cells with potential tumorigenicity
4) The source of embryos involves ethical and moral issues, so research and application are limited, so it has little clinical significance in the near future
Problems in the clinical application of adult stem cells: 1) The access to adult stem cells is limited, the number is small and the proliferation ability is limit...

Method used

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  • Method for culturing cord blood type embryonic stem cells as well as identification and application
  • Method for culturing cord blood type embryonic stem cells as well as identification and application
  • Method for culturing cord blood type embryonic stem cells as well as identification and application

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specific Embodiment approach 1

[0050] Specific embodiment 1: The method for culturing cord blood-like embryonic stem cells of this embodiment is performed according to the following steps:

[0051] 1. Collection of cord blood

[0052] 2. Separation of cord blood

[0053] Mix the cord blood collected in step 1 with the separation solution in a volume ratio of 2:1, centrifuge to collect the mononuclear cell layer, then remove the red blood cells with red blood cell lysis solution, then wash the monocytes twice with PBS, count, and adjust the cell density To 2×10 6 / mL, spare;

[0054] 3. Coating of umbilical cord blood embryonic stem cell culture dishes

[0055] First, the laminin was coated on the bottom of the dish and kept for 24 hours, then washed twice with PBS, and then equilibrated with medium for 10-20 minutes, and then the cord blood cells separated in step 2 were pressed into 1×10 5 / mL is connected to the bottom of the dish, where the laminin is first prepared with PBS into a 10-fold concentration of lamini...

specific Embodiment approach 2

[0059] Specific embodiment two: This embodiment is different from specific embodiment one in that the separation liquid in step two is a lymphocyte separation liquid, and its density is 1.076 to 1.078 g / mL. Others are the same as the first embodiment.

specific Embodiment approach 3

[0060] Specific embodiment three: This embodiment is different from specific embodiment one in that the centrifugation condition in step two is 450g centrifugation for 20 minutes. Others are the same as the first embodiment.

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Abstract

The invention discloses a method for culturing cord blood type embryonic stem cells as well as identification and application, relating to a method for culturing stem cells as well as identification and application. The culture method comprises: 1, collection of cord blood; 2, separation of the cord blood; 3, coating of a cord blood type embryonic stem cell culture dish; and 4, culture amplification of the cord blood type embryonic stem cells. The cord blood type embryonic stem cells are used for assisting recovery of injured lymphocytes. The formula of the culture medium adopts multiple cell factors to stably maintain the biological characteristic of cells and accelerate cell proliferation. The culture dish is coated by adopting laminin, and the laminin is beneficial to wall attachment of cells, so that the wall attachment time of the cells is effectively shortened, the adhesion degree of the cells to the bottom of the dish is improved and the cells are attached to the wall more firmly.

Description

Technical field [0001] The invention relates to a method for culturing stem cells, identification and application. Background technique [0002] Type I diabetes (formerly known as juvenile diabetes or insulin-dependent diabetes) is a type of diabetes. It has a completely different pathogenesis from type II diabetes. It is an autoimmune disease and may be caused by the destruction of the pancreatic islets β that produce insulin by the autoimmune system. Caused by cells. According to data released by the International Diabetes Federation (IDF) in 2009, there are currently approximately 30 million type 1 diabetes patients in the world. [0003] Type I diabetes mainly has the following prevalence factors: 1) Genetic factors: It is reported that about 25%-50% of people have the disease due to genetic factors. The genetic factors are relatively certain regardless of type I or type II. According to the study of modern twins, type I CCP dominance is 50%, and the rest are environmental fa...

Claims

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Application Information

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IPC IPC(8): C12N5/078G01N33/68C12N5/0783
Inventor 张怡芦慧颖刘艳青
Owner 天晴干细胞股份有限公司
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