CD34⁺ progenitors transduced with nef, mutated in regions important for CD4/CD8β down-regulation, still show an impaired T-cell development in FTOC. Bivariate dot plots of flow cytometric measurement of control, Nef WT (LAI), and mutant Nef (WLAA, DDGA, LLAA) transduced cells. LAI WT is representative for the other Nef WT constructs. (A) CD8β-PE, CD4-APC versus EGFP expression of transduced thymocytes at day 22 of FTOC. (B) CD8β-PE versus CD4-APC expression from thymocytes shown in panel A, gated on EGFP⁺ cells. Quadrants were set to include 99% of cells stained with isotypic controls and EGFP^– cells in lower left quadrant. Values indicate percentage (rounded off to a whole number) of cells present in corresponding quadrants (< 100 events are shown in some plots). (C) The figure shows the mean thymocyte generation ratio and standard deviations calculated from the data generated in at least 3 independent experiments. An asterisk (*) over a bar indicates a statistically significant difference between the experimental construct and control data sets (P < .05).