Enhanced growth medium and method for culturing human mammary epithelial cells

An assay for determining the toxicity of a particular agent to human mammary epithelial cells, wherein viable colonies of human mammary epithelial cells are prepared by:

• digesting samples of human mammary tissue with an enzyme digestion mixture including at least one enzyme selected to break down the mammary tissue into clumps of epithelial cells substantially free from attached stromal cells;
• separating the clumps of epithelial cells from the stromal cells and other cellular material;
• and,
• culturing the clumps of epithelial cells in a medium including conditioned media obtained from cultures of cells selected from the group consisting of human fetal intestine epithelial cells and human bladder epithelial cells so that the cells in the clumps proliferate;
• separating individual cells from the clumps of epithelial cells;
• and
• plating a relatively small number of the individual cells onto a substrate in a medium including conditioned media from at least one media obtained from human fetal intestine epithelial cells and human bladder epithelial cells so that the cells proliferate in isolation from one another to form colonies;
• said assay comprising:
• exposing the viable colonies to the particular agent;
• and
• determining the survival rate of the colonies.

An assay for determining the toxicity of a particular agent to human mammary epithelial cells, said assay employing a culture of human mammary epithelial cells prepared by:

• digesting samples of human mammary tissue with an enzyme digestion mixture including at least one enzyme selected to break down the mammary tissue into clumps of epithelial cells substantially free from attached stromal cells;
• separating the clumps of epithelial cells from the stromal cells and other cellular material;
• and
• culturing the clumps of epithelial cells in a medium including conditioned media obtained from cultures of cells selected from the group consisting of human fetal intestine epithelial cells and human bladder epithelial cells so that the cells in the clumps proliferate;
• separating individual cells from the clumps of epithelial cells;
• and
• plating individual cells onto a substrate in a medium including conditioned media from at least one of the media obtained from human fetal intestine epithelial cells and human bladder epithelial cells so that the cells proliferate in isolation from one another to form colonies, wherein said substrate has been plated with feeder cells prior to culturing the epithelial cells;
• said assay comprising:
• exposing the plated cells to the agent;
• terminating the exposure of the colonies to the treatment;
• providing supplemental growth media further including feeder cells;
• allowing the colonies plated on the substrate to proliferate until it is possible to distinguish those colonies which continue to grow from those colonies which have ceased to grow prior to determining the survival rate;
• and
• determining the toxicity by comparing the number of viable colonies with the number of colonies which have ceased to grow.