Physiochemical culture conditions for embryonic stem cells

• US 20040224401A1
• Filed: 03/26/2004
• Published: 11/11/2004
• Est. Priority Date: 03/28/2003
• Status: Abandoned Application

• Current Assignee: Wisconsin Alumni Research Foundation (University of Wisconsin System)
• Original Assignee: Wisconsin Alumni Research Foundation (University of Wisconsin System)
• Inventors: Thomson, James A., Ludwig, Tenneille E.
• Application Number:
  US10/811,423
• Publication Number:
  US 20040224401A1
• Time in Patent Office: Days
• US Class Current: 435/366
• CPC Class Codes: C12N 2500/02 Atmosphere, e.g. low oxygen conditions
• CPC Class Codes: C12N 2500/05 Inorganic components
• CPC Class Codes: C12N 2500/60 Buffer, e.g. pH regulation, osmotic pressure
• CPC Class Codes: C12N 2502/13 connective tissue cells; generic mesenchyme cells, e.g. so-called "embryonic fibroblasts"
• CPC Class Codes: C12N 5/0606 Pluripotent embryonic cells, e.g. embryonic stem cells [ES] embryonic germ cells C12N5/0611, induced pluripotent stem cells C12N5/0696

Abstract

Physiochemical parameters to improve the culturing and sub-culturing (here called cloning) of human embryonic stem cells have been investigated. Low levels of oxygen and higher than expected levels of osmolarity in the culture medium have both been found to contribute to the improved culture of human stem cells.

Claims

1. A method for culturing human embryonic stem cells comprising culturing the stem cells in a nutrient medium in which the stem cells will remain undifferentiated and in an atmosphere having no more than about 5% oxygen.

2. The method of claim 1 wherein the medium further comprises an antioxidant in the nutrient medium.

3. An improvement in methods for cloning cultures of human embryonic stem cells, the improvement comprising culturing the human embryonic stem cells prior to cloning in a nutrient medium and in an atmosphere having no more than about 5% oxygen.

4. The improvement of claim 3 further comprising adding an antioxidant to the nutrient medium in which the stem cells are cultured.

5. A method for culturing human embryonic stem cells comprising culturing the stem cells in a nutrient medium in which the stem cells can remain undifferentiated and in an atmosphere having no more than about 5% oxygen.

6. An improvement in a medium for the cultivation of human embryonic stem cells, the improvement comprising that the medium is adjusted to have an osmolarity in excess of 330 mOsMol.

7. The improvement as claimed in claim 6 wherein the osmolarity of the medium is about 350 mOsMol.

8. A stem cell culture comprising

a culture plate;

a nutrient medium in the culture plate;

growing human embryonic stem cells in the medium; and

the medium having an osmolarity in excess of 330 mOsMol.

9. A stem cell culture as claimed in claim 8 wherein the osmolarity is about 350 mOsMol.

10. A method for culturing human embryonic stem cells comprising culturing the stem cells in a nutrient medium in which the stem cells will remain undifferentiated, the medium having an osmolarity in excess of 330 mOsMol.

11. A method as claimed in claim 10 wherein the medium has an osmolarity of about 350 mOsMol.