We have investigated the effects of a single injection of cyclophosphamide (CY) in F1 host mice. One day after CY treatment, F1 host mice show increased susceptibility to graft-versus-host (GVH)-reactive parental strain spleen cells when measured by the popliteal lymph node (PLN) enlargement assay. F1 host mice challenged with parental strain cells 7 days after CY treatment show decreased PLN enlargement as compared with CY-untreated F1 hosts receiving parental strain cells. The PLN reactivity of normal F1 spleen cells injected into F1 hosts pretreated with CY was also measured. F1 spleen cells elicited marked PLN enlargement when injected into F1 hosts that were treated with CY 1 day previously. By day 7 after CY treatment of F1 hosts, F1 cells no longer produced significant PLN enlargement as compared to untreated F1 hosts receiving F1 cells. We discuss these results in terms of the possibility that CY initiates changes in host antigenicity that can lead both to an increase in responsiveness of GVH-reactive cells and to susceptibility to attack by syngeneic cells.