A gelatin sponge model of concomitant tumor immunity was employed in order to examine the clonality of T cells associated with progressing and rejected tumor sites. Here we show that freshly isolated T cells bearing TCR V_β1, CDR3 RPGTGN, J_β1.1 and TCR V_β8, CDR3 GD, J_β1.6 predominated progressing and rejected tumor sites. Despite the similarity in T cell populations, the T cells from rejected tumor sites were capable of killing the autologous tumor cells, whereas T cells from progressing tumor sites were not able to do so. The differing cytolytic ability could not be attributed to a difference in TCR ζ chain protein expression levels between both T cell populations. After a 5 day mixed lymphocyte tumor culture the T cells from the progressing tumor site were capable of killing autologous tumor cells, which suggested changes took place within the cell population during in vitro culture. Further TCR analysis revealed T cells bearing TCR V_β1, CDR3 RPGTGN, J_β1.1 and TCR V_β8, CDR3 GD, J_β1.6 were not expanded following the in vitro culture. These data suggest that the lack of cytotoxicity of freshly isolated tumor-infiltrating lymphocytes (TIL) was not due to abnormal TCR ζ chain expression or major differences in the TCR V_β usage. Additionally, the gain of TIL effector function did not correlate with an expansion of the TCR bearing T cells found to predominate the in vivo response. These data suggest that the predominant TCR V_β used by lymphocytes infiltrating regressing or rejected tumors may not represent the tumor reactive T cells that grow in culture or respond to the autologous tumor in vitro.