T-cell receptor (TCR) Valpha (TRAV) and Vbeta (TRBV) chains provide the T-cell specificity for recognition of major histocompatibility complex (MHC)-bound antigens. However, there is limited information on the diversity of TRAV use within an antigen response. Previous investigation of CD4(+) T-cell-mediated graft-versus-host disease (GVHD) in the minor histocompatibility antigen-mismatched C57BL/6 (B6)-->BALB.B irradiated murine model determined that Vbeta11(+) T cells were associated with disease severity. Polymerase chain reaction (PCR)-based complementarity-determining region 3 (CDR3)-sized spectratype analysis of B6 Vbeta11(+) T cells from the spleens of recipient BALB.B mice undergoing GVHD indicated biased use within the V(alpha)6, 9, 13, 14, 18, and 22 families. To probe deeper into this limited V(alpha) response, the current study was undertaken to further define TRAV-Jalpha (TRAJ) nucleotide sequences found in host-presensitized B6 Vbeta11(+) T cells proliferating in response to in vitro stimulation with BALB.B splenocytes. Using the nonpalindromic adaptor PCR method, we found dominant use of the TRAV13-TRAJ16 transcript combination. Then, using laser capture microdissection, we found use of the identical TRAV-TRAJ nucleotide sequence in areas dominated by infiltrating Vbeta11(+) CD4(+) T cells during the development of GVHD in both the rete-like prominences of the dorsal lingual epithelium and the ileal crypts of the small intestine.