This invention provides methods to prepare and use immunostimulatory cells forenhancing an immune response. The invention provides a method for preparingmature dendritic cells (DCs), comprising the sequential steps of: (a)signaling isolated immature dendritic cells (iDCs) with a first signalcomprising an interferon gamma receptor (IFN-.gamma.R) agonist and/or a tumornecrosis factor alpha receptor (TNF-.alpha.R) agonist to produce signaleddendritic cells and (b) signaling said signaled dendritic cells with a secondtransient signal comprising an effective amount of a CD40 agonist to produceCCR7+ mature dendritic cells. Also provided by this invention are enrichedpopulations of dendritic cells prepared by the methods of the invention. Suchdendritic cells have enhanced immunostimulatory properties and increased IL-12secretion and/or decreased IL-10 secretion. CD40 signaling can be initiated byone or more of polypeptide translated from an exogenous polynucleotideencoding CD40L (e.g., mRNA or DNA), an agonistic antibody to CD40 receptor orby CD40 ligand polypeptide. The enriched populations can be further modifiedby the administration of an immunogen to the DC. The DC will take up andprocess the immunogen on its cell surface.
