Lfng overexpression sustains proliferation of DN3 and DN4 thymocytes on OP9-DL1. DN3 (A) or DN4 (B) thymocytes were sorted from Lfng Tg⁺ or WT mice and cultured on OP9-DL1 cells. Beginning at day 7, cells were passaged every 4-5 days onto fresh OP9-DL1 monolayers. At each time point, an aliquot of cells was harvested and stained with anti-CD4–FITC versus anti-CD8–phycoerythrin (PE; left panels). The 2-parameter contour plots (5% probability) depict CD4 versus CD8 expression (left panels) or forward scatter (FSC) versus side scatter (SSC, right panels) of progeny at each time point. Numbers depict the percentage of cells in each quadrant or gate. NS, not sufficient (the number of cells present was not sufficient to be analyzed). The fold expansion for each time point is summarized in supplemental Table 2. Similar results were obtained in 2 independent experiments.