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AssigneeThe Rockefeller University
InventorsRalph M. Steinman, Nina Bhardwaj, Gerold Schuler
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We describe an improved method for generating sizable numbers of mature dendritic cells from nonproliferating progenitors in human blood. The first step or “priming” phase is a culture of T cell depleted mononuclear cells in medium supplemented with GM-CSF and IL-4 to produce immature dendritic cells. The second step or “differentiation” phase requires the exposure to dendritic cell maturation factor such as monocyte conditioned medium. Using this two-step approach, substantial yields are obtained. The dendritic cells derive from this method have all the features of mature cells. They include a stellate cell shape, nonadherence to plastic, and very strong T cell stimulatory activity. The mature dendritic cells produced according to this invention are useful for activating T cells.
An in vitro method of producing mature CD83 positive dendritic cells from CD83 negative immature dendritic cells, said method comprising culturing the CD83 negative immature dendritic cells in the presence of a dendritic cell maturation factor for a time sufficient for said immature dendritic cells to mature and express characteristics of mature dendritic cells including an increase in CD83 expression which is stable for up to three days following removal of said dendritic cell maturation factor, and wherein the dendritic cell maturation factor comprises IFNα.
An in vitro method of producing mature CD83 positive dendritic cells, said method comprising contacting CD14 positive mononuclear pluripotential cells with at least one first factor to obtain immature CD83 negative dendritic cells and culturing the immature CD83 negative dendritic cells with a dendritic cell maturation factor for a time sufficient for said immature dendritic cells to mature and express characteristics of mature dendritic cells including an increase in CD83 expression which is stable for up to three days following removal of said dendritic cell maturation factor, and wherein the dendritic cell maturation factor comprises IFNα.
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