Chicken anti-rat lymphocyte globulin (CARLG) is a new and uniquely specific immunosuppressant for which the mechanisms of immunosuppressive action are not yet known. This study defines its in vivo therapeutic dose range and in vitro allospecificity. A dose response study, using CARLG or normal chicken globulin (NCG) as the sole immunosuppressant, was conducted in Lewis (Ag-B1) recipients of Buffalo (Ag-B6) rat cardiac allografts. Groups of Lewis recipients were treated with 175 mg of NCG per kg or 30,100,175, or 300 mg of CARLG per kg for the first 8 post-transplant days. Doses of 175 and 300 mg of CARLG per kg resulted in significantly longer (P < 0.01) prolongation of graft survival than doses of 30 and 100 mg of CARLG per kg, which were not significantly different (P > 0.05) in their ability to prolong graft survival than a dose of 175 mg of NCG per kg. In vitro and in vivo absorption studies revealed that CARLG does not contain strain-specific alloanti-body. When Lewis lymphoid cells were used to absorb, in vitro, a batch of CARLG produced with Brown Norway (Ag-B3) lymphoid cells as immunogen, all antibody activity toward Brown Norway cells as quantitated by a two-stage lymphocy-totoxicity assay was completely removed. In another experiment, a preparation of CARLG, produced using Buffalo lymphoid cells as immunogen, underwent in vivo absorption as a result of i.v. injection into Lewis rats. Serum levels of CARLG-binding activity to Lewis or Buffalo lymphoid cells were quantitated in vitro at intervals after injection by an isotopic antiglobulin assay. Lewis rat tissues were able to remove completely serum CARLG-binding activity toward Buffalo antigens. A mechanism of action for CARLG derived from these observations is proposed and discussed.