METHOD FOR CELL EXPANSION

• US 20110070648A1
• Filed: 05/11/2009
• Published: 03/24/2011
• Est. Priority Date: 05/15/2008
• Status: Abandoned Application

• Current Assignee: GE Healthcare Bio-Sciences AB (GE Healthcare Technologies, Inc.)
• Original Assignee: GE Healthcare Bio-Sciences AB (GE Healthcare Technologies, Inc.)
• Inventors: Anneren, Cecilia, Pauler, Linnea
• Application Number:
  US12/991,930
• Publication Number:
  US 20110070648A1
• Time in Patent Office: Days
• US Class Current: 435/380
• CPC Class Codes: C12M 23/14 Bags
• CPC Class Codes: C12M 23/20 Material Coatings immunocoatings C12M25/00
• CPC Class Codes: C12M 25/16 Particles; Beads; Granular material; Encapsulation chemical or physical processes conducted in the presence of fluids and solid particles B01J8/00
• CPC Class Codes: C12N 2531/00 Microcarriers
• CPC Class Codes: C12N 5/0663 Bone marrow mesenchymal stem cells (BM-MSC)

Abstract

The present invention relates to a method for cell expansion. More closely, it relates to a method for expansion of cells, such as mesenchymal stem cells, on microcarriers in a plastic bag bioreactor. The invention enables expansion to therapeutic amounts of stem cells. The method comprises the following steps: a) addition of cells in cell culture medium and microcarriers to a plastic bag container; b) allowing the cells to adhere to the microcarriers while the container is kept substantially still; c) addition of further cell culture medium once the cells have adhered; d) culturing the cells under gentle and constant agitation; e) increase the surface area for continued culturing; and f) final harvesting of cells by an active detachment and separation step.

Claims

1. A method for cell expansion, comprising the following steps:

a) addition of cells in cell culture medium and microcarriers to a plastic bag container;

b) allowing the cells to adhere to the microcarriers while the plastic bag container is kept substantially still;

c) addition of further cell culture medium once the cells have adhered;

d) culturing the cells under gentle (<

7 rpm) and constant agitation;

e) increase the surface area for continued culturing; and

f) final harvesting of cells by an active detachment and separation step.

2. The method of claim 1, wherein step e) is repeated at least once.

3. The method of claim 1, comprising in step e) adding new carriers and allowing spontaneous migration of cells to the new carriers.

4. The method of claim 1, comprising in step e) an active detachment step of cells from the microcarriers and addition of new carriers.

5. The method of claim 1, wherein the microcarriers are provided with magnetic particles.

6. The method of claim 1, wherein the volume of the added cells and microcarriers in step a) is between 150-300 ml, the seeding density is 1-10 cells /microcarrier and the amount of microcarriers is at least 0.2 g (dry weight) per litre.

7. The method of claim 1, wherein the cells are nucleated cells from adipose tissue, bone marrow or cord blood.

8. The method of claim 7, wherein the cells are adult mesenchymal stem cells.

9. The method of claim 1, wherein steps e) and f) are repeated until 3-8 g microcarriers per litre medium is reached.

10. The method of claim 1, wherein the container is an inflated bioreactor bag.

11. The method of claim 1, wherein the cells are allowed to detach in step e) inside the container.

12. The method of claim 1, wherein the cells are allowed to detach in step e) outside the container, after which the cells and microcarriers are re-introduced into the container.

13. The method of claim 1, wherein the cells are allowed to detach by using enzymes, thermo responsive agents and/or pH responsive agents.

14. The method of claim 1, wherein the cells are expanded by allowing passive migration from confluent beads to newly added beads.