The rat Tcra V gene locus is only poorly characterized, although rats are widely used in a variety of T-cell-mediated experimental animal models. Recently, we described the first monoclonal antibody, G99, directed against a rat Tcra V4 segment. We examined cDNA transcripts of G99-positively sorted T cells and show that the monoclonal antibody G99 most likely recognizes at least two members of the Tcra V4 family. Moreover, we analyzed the genomic repertoire of this VA family and report 15 novel Tcra V4 DNA sequences. Based on sequence and Southern blot analysis, the Tcra V4 family could be divided into four subgroups, which were also detected in mice. These findings corroborate previous findings of a similar genetic organization of the Tcra V loci in both species.