The culture of mononuclear umbilical cord blood cells with low concentrations of interleukin-3 (IL-3) and interleukin-5 (IL-5) for 3 weeks or more results in the consistent production of large numbers of eosinophils. Density distribution analysis and electron microscopic analysis revealed the characteristics of cultured eosinophils to be partially different from peripheral eosinophils. In modified Boyden chamber studies, cultured eosinophils exhibited locomotive activity in response to platelet-activating factor (PAF), with a peak response at 10(-6) M, but not to N-formyl-L-methionyl-L-leucyl-L-phenylalanine. PAF-induced changes in intracellular calcium concentrations in peripheral eosinophils and cultured eosinophils were similar. PAF-induced actin polymerization was observed in cultured eosinophils for the first 30 s followed by depolymerization for the next 10 min. These results suggest that cultured eosinophils respond to PAF in a manner analogous to peripheral eosinophils, and will be useful for in vitro studies.