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US 10,472,612 B2Granted
Cell culture system
AssigneePresident and Fellows of Harvard College
InventorsDonald E. Ingber, Hyun Jung Kim
Life of this patent
Estimating the term…
Abstract
The embodiments of the invention described herein relate to systems and methods for culturing and/or maintaining intestinal cells, tissues and/or organoids in vitro. The cells, tissues and/or organoids cultured according to the methods and systems described herein can mimic or reproduce natural intestinal epithelial structures and behavior as well as support co-culture of intestinal microflora.
Claims34
A method for maintaining the viability of intestinal cells and bacterial cells in co-culture comprising:
- a) providing a fluidic device having:
- a fluid channel connected to a fluid source, the fluid source supplying culture media to the fluid channel;
- and
- a membrane positioned within the channel between membrane support elements;
- b) introducing, to at least one surface of the membrane viable intestinal epithelial cells;
- c) pre-culturing said intestinal epithelial cells such that a layer of cells is formed that is attached to said membrane, said layer exhibiting tight junctions;
- d) introducing viable bacterial cells to produce a co-culture comprising at least a portion of the bacterial cells adhered to the intestinal epithelial cells;
- e) flowing the culture media over the co-culture at a flow rate sufficient to wash non-adherent bacterial cells out of the fluidic device;
- and
- f) culturing the intestinal epithelial cells and bacterial cells for at least 48 hours wherein more than 95% of the intestinal epithelial cells are viable after at least 48 hours of culture.
7 more elements
A method for maintaining the viability of intestinal cells and bacterial cells in co-culture comprising:
- a) providing a fluidic device having:
- a fluid channel connected to a fluid source, the fluid source supplying culture media to the fluid channel;
- and
- a membrane positioned within the channel between membrane support elements;
- b) introducing, to at least one surface of the membrane, viable intestinal epithelial cells;
- c) pre-culturing said intestinal epithelial cells such that a layer of cells is formed that is attached to said membrane, said layer exhibiting barrier function;
- d) introducing viable bacterial cells to produce a co-culture comprising at least a portion of the bacterial cells adhered to the intestinal epithelial cells;
- e) flowing the culture media over the co-culture at a flow rate sufficient to wash non-adherent bacterial cells out of the fluidic device;
- and
- f) culturing the intestinal epithelial cells and bacterial cells for at least 48 hours wherein more than 95% of the intestinal epithelial cells are viable after at least 48 hours of culture.
7 more elements
A method for maintaining the viability of intestinal cells and bacterial cells in co-culture comprising:
- a) providing a fluidic device having:
- a fluid channel connected to a fluid source, the fluid source supplying culture media to the fluid channel;
- and
- a membrane positioned within the channel between membrane support elements;
- b) introducing, to at least one surface of the membrane:
- viable intestinal epithelial cells;
- c) pre-culturing said intestinal epithelial cells such that a layer of cells is formed that is attached to said membrane, said layer comprising polarized intestinal epithelial cells;
- d) introducing viable bacterial cells to produce a co-culture comprising at least a portion of the bacterial cells adhered to the intestinal epithelial cells;
- e) flowing the culture media over the co-culture at a flow rate sufficient to wash non-adherent bacterial cells out of the fluidic device;
- and
- f) culturing the intestinal epithelial cells and bacterial cells for at least 48 hours wherein more than 95% of the intestinal epithelial cells are viable after at least 48 hours of culture.
8 more elements
A method for maintaining the viability of intestinal cells and bacterial cells in co-culture comprising:
- a) providing a fluidic device having:
- a fluid channel connected to a fluid source, the fluid source supplying culture media to the fluid channel;
- and
- a membrane positioned within the channel between membrane support elements;
- b) introducing, to at least one surface of the membrane, viable intestinal epithelial cells;
- c) pre-culturing said intestinal epithelial cells such that a layer of cells is formed that is attached to said membrane, said layer comprising intestinal epithelial cells comprising villi structures;
- d) introducing viable bacterial cells to produce a co-culture comprising at least a portion of the bacterial cells adhered to the intestinal epithelial cells;
- e) flowing the culture media over the co-culture at a flow rate sufficient to wash non-adherent bacterial cells out of the fluidic device;
- and
- f) culturing the intestinal epithelial cells and bacterial cells for at least 48 hours wherein more than 95% of the intestinal epithelial cells are viable after at least 48 hours of culture.
7 more elements
Dependencies are inferred from the claim text.