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A method is described allowing the isolation and culture of relatively pure preparations of murine Kupffer cells for long-term morphological and functional studies, In vivo labelling experiments showed no evidence of significant blood monocyte contamination. The use of a density gradient allowed removal of fibroblasts, cell debris and a relative enrichment of Kupffer cells. After 24 hr in culture the cells were phagocytic, contained non-specific esterase and exhibited an Fc receptor. During the first 3 weeks, cell clumps, epithelioid cells and multinucleate giant cells appeared. Mitoses were seen in the Kupffer cells, cell numbers increased and In vitro labelling with 3H-thymidine at 49 days showed that 33% of the cells underwent DNA synthesis in a 24-hr period. Experiments suggested that giant cell formation was produced both by cell fusion and nuclear division. The experiments show that Kupffer cells are capable of cell division and giant cell formation in culture. [ABSTRACT FROM AUTHOR]