Induction of T-cell activation markers following stimulation of human cord blood–derived HSCs induced to differentiate on OP9-DL1 cells. CB-derived HSCs cocultured with OP9-DL1 cells for 52 days were stimulated with anti-CD3 plus anti-CD28 mAbs in the presence or absence of recombinant human IL-2 added to the media. (A) The expression of CD27 and CD69 was analyzed before stimulation and 2 and 5 days after stimulation on CD3⁺-gated cells. Anti-CD3 plus anti-CD28 mAb in IL-2–enriched cocultures are compared to IL-2–enriched or control (no IL-2 added) cocultures alone. (B) The expression of CD28 (histograms presented in top row) and CTLA-4 (histograms presented in bottom row) was analyzed before stimulation and 2 and 5 days after stimulation on CD3⁺-gated cells from cocultures treated without IL-2 (dotted thin black line) or with IL-2 (solid thin black line), or with IL-2 and anti-CD3 plus anti-CD28 (solid thick black line) added and then compared to the appropriate isotype control (thin dotted gray line).