T cell receptor excision circles (TRECs) have been suggested to be useful markers of recent thymic output and graft-versus-host disease (GVHD) was thought to have an adverse effect on thymic output. On the other hand, TREC concentrations are diluted with T cell expansion. The C-type lectin superfamily inhibitory receptor CD94 expression on T cells might be a consequence of CD8 T cell expansion. Therefore, this study was designed to analyse TREC DNA level in CD4 and CD8 T cell subset expressing CD94 during GVHD. We investigated TREC DNA copy numbers (TREC levels) of CD4+, CD94+/CD8+ and CD94-/CD8+ T cells in 24 patients who had undergone allogeneic stem cell transplantation and also of in vitro activated and expanded CD94-expressing cells by immobilised anti-CD3 mAb and IL-15. TREC level of CD94+/CD8+ T cells in patients with chronic GVHD was lower than that in patients with no GVHD and with remission status of GVHD. In vitro activated and expanded CD94-expressing cells had a significantly lower TREC level than that in untreated CD8 cells. Therefore, the low TREC level of CD94+/CD8+ T cells is related to chronic GVHD and may reflect T cell expansion during chronic GVHD.