Decreased in vitro apoptosis of AKR/J TN thymocytes. Total thymocytes were isolated from 2-month-old AKR/J or control B10.BR mice and cultured for 12 hours in medium supplemented with 5% serum. Cells were stained with a cocktail of Cy–(anti-CD4, anti-CD8, anti-TCRαβ) antibodies together with anti-CD25–APC and anti-CD44–FITC. Subsequently, apoptotic cells were detected by FACS-TUNEL staining revealed by addition of PE-conjugated streptavidin. FACS profiles (A) represent frequency of apoptotic cells, as determined by TUNEL positive staining, among gated TN (CD4^–CD8^–TCRαβ^–) thymocytes and among each cell subset within TN thymocytes: CD44^–CD25^–, CD44^–-CD25⁺, CD44⁺CD25^–, and CD44⁺CD25⁺. FACS profiles (B) represent cell size analysis, as determined by FSC channel, among CD44^–CD25⁺ TN thymocytes. This cell subset is divided into small- and large-size cells, and frequency of apoptotic cells among each population is determined by TUNEL staining versus FSC. Results (A-B) are representative of 3 independent experiments with n = 5 mice for each experiments. FACS profiles (C) represent analysis of in vitro apoptosis after addition of 5 ng/mL recombinant mouse IL-7. Analysis of FSC scatter and TUNEL staining are shown among CD44^–CD25⁺ TN thymocytes. Results (C) are representative of 3 independent experiments with n = 3 mice for each experiments. Numbers shown in FACS profiles denote percentages of cells in each quadrant.