Generation of CD8^low SP8 thymocytes correlates with reduced frequency of Melan-A–specific thymocytes. SP8 thymocytes from HLA-A2⁺ SCID-hu Thy/Liv mice (see Figure 5B) were enriched by CD4 depletion on magnetic beads and stained with MelanA pentamers or control CMV tetramers. (A) Representative examples in each group. (B-D) Frequency of MelanA⁺ SP8 thymocytes for all thymi (B), correlation with MHC-I up-regulation on DP thymocytes (C), and correlation with CD8α expression in MelanA⁺ SP8 thymocytes (D). P values were calculated using the Mann-Whitney test for group analysis and by Spearman rank correlation for correlation test. R² from the Pearson coefficient of correlation is also indicated.