Suspension culture of human embryonic stem cells

Abstract

This disclosure provides an improved system for culturing human embryonic stem cells. The cells are cultured in suspension so as to maximize the production capacity of the culture environment. The new culture system of this invention allows for bulk proliferation of hES cells in a more cost-effective manner, which facilitates commercial production of important products for use in human therapy.

1 .- 18 . (canceled) 
     
     
         19 . A system or kit for culturing human pluripotent stem (hPS) cells in suspension, comprising a nutrient medium that contains at least 40 ng/mL fibroblast growth factor (FGF). 
     
     
         20 . The system or kit of  claim 19 , further comprising one or more extracellular matrix components or microparticles for adding to the medium. 
     
     
         21 . The system or kit of  claim 20 , wherein the one or more extracellular matrix components are selected from human laminin and human fibronectin. 
     
     
         22 . The system or kit of  claim 20 , wherein the microparticles are coated with one or more soluble or suspended extracellular matrix components. 
     
     
         23 . The system or kit of  claim 19 , wherein the medium further comprises one or more of transforming growth factor beta (TGFβ), stem cell factor (SCF), and Flt3 ligand (Flt3L). 
     
     
         24 . The system or kit of  claim 19 , wherein FGF is basic FGF (bFGF). 
     
     
         25 . The system or kit of  claim 24 , wherein the concentration of bFGF is about 80 ng/mL. 
     
     
         26 . The system or kit of  claim 19 , further comprising instructions for use. 
     
     
         27 . Use of the system or kit of  claim 19  for culturing cells wherein culturing cells comprises:
 a) suspending cells in the nutrient medium; 
 b) maintaining the cells in suspension while culturing; 
 c) changing the medium periodically; 
 d) optionally splitting the culture from time to time so as to reduce cell density; and finally 
 e) harvesting cells from the culture.