Figure 4.

Gene expression analysis of T cell-related genes in sorted CD4-1⁺ and CD8α⁺ lymphocytes. Trunk-kidney and spleen cells were stained with anti-CD4-1 and CD8α mAbs as described in Figure 3. Lymphocytes fraction from trunk-kidney and spleen were gated on FS and SS dot plot and doublets discrimination was performed. Anti-CD4-1 and CD8α mAb positive cells were sorted by FACS. Purity of FACS sorted cells was confirmed to be more than 95%. Total RNA was prepared from 5 × 10⁵ sorted cells and used for quantitative RT-PCR analysis. mRNA expression of cd3e, tcrb, zap-70, and lck in sorted T cells was normalized to internal control gene, ef1a. Statistical significance was calculated using t tests to each gene (ns, not significant p > 0.05).