A method for monitoring circulating lymphocytes subsets in the rat on an automated flow cytometer with monoclonal antibodies was used to ascertain in vivo effects of various doses of immunosuppressive agents. The agents tested were anti-lymphocyte serum (ALS), azathioprine (AZA), cyclophosphamide (CTX), cyclosporin A (CsA) and methylprednisolone (MP). Each immunosuppressive agent varied in its capacity to induce changes in T cell subsets and B cell numbers. The rapidity of onset of action of the agents varied considerably; with ALS and MP maximal effects were seen within hours whilst the effects with CsA, cyclophosphamide (CTX) and azathioprine (AZA) took several days to develop. ALS had marked anti-T cell activity but did not selectively affect the T cell subsets. AZA and CTX both exerted their major effect upon the B cell (OX4+) subpopulation. CsA administration was associated with the appearance of many circulating lymphocytes which expressed the pan-T marker (W3/13) but neither of the T cell subset markers (W3/25, OX8). With CsA there was no significant alteration in the W3/25:OX8 ratio, although a persistent decrease in the number of all T lymphocytes was observed after administration of this drug at a dose of 45 mg/kg had ceased. MP was the only drug which had a marked selective effect on a T cell subset. The numbers of circulating Class II major histocompatibility complex (MHC) reactive lymphocytes (W3/25+) were significantly more depressed than the Class I MHC reactive subset (OX8+). This effect persisted for up to 31 days after the single injection of a depot preparation of this drug, and was found to be associated with prolonged survival of precultured endocrine xenografts.