The calcineurin inhibitors cyclosporin A (CsA), tacrolimus (FK506) and pimecrolimus (ASM981) are on the market for the oral treatment of psoriasis and atopic dermatitis and topical treatment of atopic dermatitis, respectively. The effect of these treatments on the immune response was investigated in this study after immunisation of rats with keyhole limpet hemocyanin (KLH). Male rats (10 per group) were orally administered pimecrolimus at 10 or 30 mg/kg/day), tacrolimus at 3 mg/kg/day or CsA at 20 mg/kg/day for 4 weeks. Control animals similarly received the vehicle only. The last five animals per group were immunised and challenged with KLH on days 16 and 24, respectively. Eight days after the last injection, the immune function was investigated by detecting KLH-specific antibodies in the serum and by examination of cell infiltration at the site of the KLH-challenge. In addition, a correlation between functional and structural changes was established by quantification of lymphocyte sub-populations in the blood or residing in lymphatic tissue. In KLH-immunised rats, CsA caused complete suppression of the KLH-specific IgM and IgG production, whereas only IgG production was affected by pimecrolimus at 30 mg/kg/day and more so by tacrolimus at 3 mg/kg/day. Immunophenotyping of lymphocyte sub-populations in spleen and lymph node indicated a decrease in T lymphocytes with pimecrolimus at 30 mg/kg/day, tacrolimus and CsA, whereas these changes were marginal for pimecrolimus at 10 mg/kg/day. Immunophenotyping of peripheral white blood cells (WBC) revealed a decrease in the absolute number of T lymphocytes with all three test items. In comparison with non-immunised animals, a slight increase in absolute numbers of T lymphocytes was observed in KLH-immunised animals treated with pimecrolimus at 10 or 30 mg/kg/day. In conclusion, the ability of the immune system to respond to KLH was not affected by pimecrolimus at 10 mg/kg/day whereas a decrease in immune function was noted in the other groups as follows: pimecrolimus (30 mg/kg/day) < tacrolimus (3 mg/kg/day) < CsA (20 mg/kg/day).