Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Human embryonic stem cells culture medium without dependent feeding cell

A culture medium and stem cell technology, applied in the field of human embryonic stem cell culture medium, can solve problems such as unseen

Inactive Publication Date: 2005-08-24
SHIYAN TAIHE HOSPITAL +1
View PDF0 Cites 1 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there are no relevant reports on the success of such work

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

[0075] Example 1 Cell Culture

[0076] 1. Purpose: To directly use LD-1 in hES cell culture, observe its growth and morphological changes, and set up a positive control group and a negative control group at the same time for simultaneous comparison.

[0077] 2. Grouping and methods: Inoculate 26-generation hES cells (H1) cultured on Matrigel with conditioned medium into 4-well plates coated with Matrigel, and divide them into three groups:

[0078] The first group is cultured with conditioned medium (positive control);

[0079] The second group was cultured with LD-1 medium (i.e.: normal medium + 40ng / ml Human FGF basic + 500ng / ml Recombinant Mouse Noggin / Fc Chimera);

[0080] The third group was cultured with normal medium (negative control).

[0081] According to 0.5ml culture medium / well, the culture medium was changed once a day, passaged once every 5-7 days, a total of 6 passages.

[0082] 3. Results: The cells in the first group and the second group grew well at the s...

Embodiment 2

[0084] Example 2 Immunohistochemical detection

[0085] 1. Objective: To detect the characteristics of some human embryonic stem cells cultured with LD-1 for 6 passages by immunohistochemical method.

[0086] 2. Method

[0087] 1. Cells: Plant LD-1 cultured hES cells that have undergone multiple passages into 4 wells of a 4-well plate, and plant hES cells cultured in conditioned medium and ordinary medium into the other two 4-well plates respectively. In 4 holes;

[0088] 2. Fix the cells: culture the above cells until the 4th day, suck off the culture medium, add 4% paraformaldehyde, and leave it at room temperature for 10 minutes to fix the cells;

[0089] 3. Cell grouping: the aforementioned fixed cells were divided into 4 groups, and each group contained 1 well of hES cells cultured in LD-1, conditioned medium and normal medium respectively.

[0090] 4. Primary antibody: a) Oct-4 (Chemicon MAB4305, mouse IgG1), b) SSEA-4 (Chemicon MAB4304, mouse IgG3), c) TRA-1-60 (Chem...

Embodiment 3

[0096] Example 3 Quantitative Analysis

[0097] 1. Objective: To quantitatively analyze hES cells after 6 passages of LD-1 culture by using flow cytometry.

[0098] 2. Materials hES cells from the first, second and third groups, anti-Oct4, etc.

[0099] 3. Methods Digest with trypsin / EDTA and resuspend in PBS (containing 10% goat serum) at 4°C for 15 minutes. After adding anti-Oct4 monoclonal antibody, incubate at 4°C for 30 min, and then add FITC fluorescein-labeled secondary antibody. The upper flow cytometer (Beckman Coulter Epics XL) was detected, and the data was analyzed by Coulter SYSTEMII software.

[0100] 4. Results The Oct4-positive rate of hES cells in the first and second groups was over 85%; the Oct4-positive rate of hES cells in the third group was less than 20%

[0101] V. Conclusion The hES cells in the first and second groups can basically maintain their undifferentiated state.

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

A human embryonic stem cell culture medium LD-1 independent to feeder cell (fibroblast or others) features that it contains some growth factor, such as fibre growth factor 2 (bFGF) and Noggin, which can support the self updating of human embryonic stem cells.

Description

Technical field: [0001] The present invention relates to a culture medium for human embryonic stem cells (LD1 cell culture medium) that does not depend on feeder cells, in particular to adding some factors that support the self-renewal of human embryonic stem cells to ordinary human embryonic stem cell culture medium, and does not depend on mouse embryogenesis Human embryonic stem cell culture medium for fibroblasts or any other cells. Background technique: [0002] In 1998, Professor Thomson of the University of Wisconsin in the United States successfully isolated the most primitive undifferentiated inner cell mass (Inner Cell Mass, ICM) in early human embryos (blastocysts 5-7 days after fertilization) and inoculated them into monolayer cultures. Mouse embryonic fibroblasts (MEFs) were continuously cultured and passaged in vitro, and kept in an undifferentiated state, and they were called human embryonic stem cells (hereinafter referred to as human embryonic stem cells or h...

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): C12N5/08
Inventor 李东升邓宏魁
Owner SHIYAN TAIHE HOSPITAL
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

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

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com