Extraction and separation method of embryonic stem cells

A technology for embryonic stem cells and separation methods, applied in cell dissociation methods, embryonic cells, biochemical equipment and methods, etc., can solve the problems of inner cell group damage, easily damaged cells, and high difficulty, and achieve the effect of vigorous proliferation

Pending Publication Date: 2021-03-19
广州北斗生物科技有限公司
View PDF1 Cites 1 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] Existing embryonic stem cell isolation mostly adopts immunosurgical methods and microsurgical methods. Since immunosurgical methods require special reagents to remove the zona pellucida and trophoblast, it is easy to cause damage to the inner cell population, while microsurgical methods require Specialized instruments and equipment, and high technical level requirements for personnel, are difficult to popularize and apply
[0004] To sum up, the existing embryonic stem cell isolation method is difficult, easy to damage the cells, and affect the isolation of embryonic stem cells. Therefore, we propose a method for the extraction and isolation of embryonic stem cells

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0022] S1. Take the 3-5 day old blastocysts of mice, remove the ovaries of mammals, and give exogenous hormones to make the embryos continue to develop, but delay implantation;

[0023] S2. After 4-6 days of development, the blastocysts were washed out from the uterus and cultured. As a result, the trophoblast cells grew and pushed away the feeder layer cells, and spread on the bottom wall of the culture dish;

[0024] S3. ICM cells proliferate and grow vertically upwards to form a cylindrical egg structure. The columnar structure is picked out with a fine glass needle under a microscope, and the original embryonic stem cell suspension is inoculated into adult cells that have been irradiated by radiation or treated with mitomycin C. Culture on the fibroblast feeder layer. After 1 week of culture, typical embryonic stem cell colonies were found. Cell colonies were picked and dispersed with low-concentration trypsin, then subcultured. Subcultured once every 3-5 days, after repeat...

Embodiment 2

[0027] S1. Take 9-10 day old pig blastocysts, remove the ovaries of mammals, and give exogenous hormones to make the embryos continue to develop, but delay implantation;

[0028] S2. After 4-6 days of development, the blastocysts were washed out from the uterus and cultured. As a result, the trophoblast cells grew and pushed away the feeder layer cells, and spread on the bottom wall of the culture dish;

[0029] S3. ICM cells proliferate and grow vertically upwards to form a cylindrical egg structure. The columnar structure is picked out with a fine glass needle under a microscope, and the original embryonic stem cell suspension is inoculated into adult cells that have been irradiated by radiation or treated with mitomycin C. Culture on the fibroblast feeder layer. After 1 week of culture, typical embryonic stem cell colonies were found. Cell colonies were picked and dispersed with low-concentration trypsin, then subcultured. Subcultured once every 3-5 days, after repeated subc...

Embodiment 3

[0032] S1. Take the 7-8 day old blastocysts of sheep, remove the ovaries of mammals, and give exogenous hormones to make the embryos continue to develop, but delay implantation;

[0033] S2. After 4-6 days of development, the blastocysts were washed out from the uterus and cultured. As a result, the trophoblast cells grew and pushed away the feeder layer cells, and spread on the bottom wall of the culture dish;

[0034] S3. ICM cells proliferate and grow vertically upwards to form a cylindrical egg structure. The columnar structure is picked out with a fine glass needle under a microscope, and the original embryonic stem cell suspension is inoculated into adult cells that have been irradiated by radiation or treated with mitomycin C. Culture on the fibroblast feeder layer. After 1 week of culture, typical embryonic stem cell colonies were found. Cell colonies were picked and dispersed with low-concentration trypsin, then subcultured. Subcultured once every 3-5 days, after repea...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

PUM

No PUM Login to view more

Abstract

The invention discloses an extraction and separation method of embryonic stem cells. The extraction and separation method comprises the following steps: S1, the ovary of a mammal is excised, and exogenous hormone is given to make an embryo continue to develop but implantation is delayed; S2, after the embryo develops for 4-6 days, blastocysts are taken by flushing from a uterus for culture, such that trophoblastic cells grow and feeder layer cells are pushed away, and are spread on the bottom wall of a culture dish; and S3, inner cell mass (ICM) cells proliferate, and vertically grow upward toform egg cylindrical structures, the cylindrical structures are picked out by using a fine glass needle under a microscope, and digestive passaging is performed to obtain embryonic stem cell-like colonies. According to the extraction and separation method, a tissue culture method is adopted to extract and separate the embryonic stem cell; by inoculating the blastocysts on a feeder layer, the implantation of blastocysts in vivo is simulated, which is closer to the natural development process; inner cell populations proliferate vigorously, and thus, embryonic stem cell-like colonies are relatively easy to obtain; and the problems that an existing separation method of the embryonic stem cells is difficult, the cells are easy to damage, and the separation of the embryonic stem cells is influenced are solved.

Description

technical field [0001] The invention relates to the technical field of embryonic stem cell separation, in particular to a method for extracting and isolating embryonic stem cells. Background technique [0002] Embryonic stem cells are a kind of totipotent cells isolated from the inner cell mass of pre-implantation embryos or primordial germ cells through in vitro differentiation inhibition culture. It has the characteristics of unlimited proliferation, self-renewal and multi-lineage differentiation in vitro. Embryonic stem cells have a similar morphological structure to early embryonic cells, with large nuclei, one or several nucleoli, mostly euchromatin in the nucleus, less cytoplasm and a simple structure. When cultured in vitro, the cells are closely arranged and grow in the form of colonies. Stained with alkaline phosphatase, ES cells were brown-red, while the surrounding fibroblasts were pale yellow. [0003] Existing embryonic stem cell isolation mostly adopts immun...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

Application Information

Patent Timeline
no application Login to view more
Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/0735C12N5/071
CPCC12N5/0606C12N2502/1323C12N2509/00
Inventor 朱瑜
Owner 广州北斗生物科技有限公司
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap