Quantitative assessment of mRNA expression in single cells. Single cells of each CD8⁺ T-cell subset were sorted, and cells expressing each gene identified, as shown in Figures 2–3. In all cells that scored positive for the expression of any particular gene, the mRNA expression levels of that gene were further quantified. (A) Each graph shows 3 cells from each population. We show GZMA and RANTES amplification because these genes can be found in all cell sets, and thus their expression levels can be directly compared. (B) Correlation between the number of mRNA molecules expressed by every single cell of T_EM and T_EMRA compounding subsets. Individual cells expressing GZMA, IL-10Rα, and RANTES from either T_EM or T_EMRA were ordered according to the expression level of that gene and attributed an ordinal point. The number of mRNA molecules associated with each ordinal point from T_EM subsets was plotted against the correspondent ordinal point values of the equivalent T_EMRA subset. The dots in the graphs represent the intersection of the values derived from the number of mRNA molecules expressed by 1 T_EM cell (horizontal axis) and 1 T_EMRA cell (vertical axis). Data are from 1 of 3 donors.