The effects of dietary alpha-linolenic acid (18:3n-3) on fatty acid composition, eicosanoid production, and cell-mediated cytotoxic activity of immune cells before and after challenge with virus or poly I-C from BALB/c mice were studied. Weanling BALB/c mice were fed purified diets containing either 10%-by-weight corn oil or linseed oil providing a ratio of 18:3n-3 to 18:2n-6 of 1/32 or 2/1, respectively, for 6-10 weeks. Fatty acid analysis of splenocyte phospholipids showed an appreciable increase in the percentage of n-3, and a decrease in n-6, fatty acids in splenocytes from mice fed the linseed oil diet. Splenocyte prostaglandin E and peritoneal exudate cell leukotriene C production was significantly lower in the linseed oil-fed mice. In general, cell-mediated cytotoxic activity was similar for immune cells from linseed oil and corn oil-fed mice. However, 6 days after the viral challenge, splenocyte cell-mediated cytotoxic activity was significantly higher in linseed oil mice. This higher activity was associated with nonspecific cytotoxicity rather than that of viral-specific cytotoxic T-lymphocytes. Cell yields from the spleen and peritoneum were frequently significantly higher in linseed oil mice. Interactions between dietary 18:3n-3, eicosanoid production, and immune cell proliferation and/or migration are discussed. In summary, feeding mice a diet rich in 18:3n-3 elevates immune cell n-3 fatty acid content, reduces eicosanoid synthesis and, to a limited extent, enhances the cell-mediated cytotoxic response to a viral challenge.