Single chicken thymic nurse cells (TNC) placed onto the chorionallantoic membrane (CAM), showed that intra-TNC lymphocytes (TNC-L) possess a strong graft-versus-host reactivity (GVHR) in allogeneic MHC combinations. This reaction shows the morphological, phenotypic, and functional characteristics of a classical GVH reaction (GVHR). The induction of a GVHR was significantly higher for TNC-L as compared with thymocytes or peripheral blood lymphocytes (PBL). The specificity of the GVHR was shown by serial transfer experiments onto appropriate allogeneic and syngeneic secondary embryonic hosts. In immunofluorescence analyses with monoclonal antibodies (mAb) to the chicken alpha/beta and gamma/delta T cell receptors (TCR) and the CD3, CD4, and CD8 equivalents, an enrichment of CD3+/CD4+/CD8- and CD3+/CD-4-/CD8+, TCR-alpha/beta + and TCR- gamma/delta + cells was observed inside TNC as compared with extra-TNC thymocytes. A large proportion of CD4+ and/or CD8+ TCR- gamma/delta + cells were demonstrated inside TNC. A minor population among TCR- gamma/delta extra-TNC thymocytes also expressed CD4 and/or CD8 molecules. Based on functional tests and double staining experiments, we propose that CD4+/CD8+ thymocytes enter the TNC where they may undergo positive selection for MHC restriction and further differentiation to CD4 or CD8 single-positive cells. Taken together these data support the concept that TNC contribute a specialized thymic microenvironment for T cell differentiation and maturation.