Expression of gp91^phox in cultured X-CGD CD34⁺PBSCs under optimum conditions for ex vivo transduction with lentivector or oncoretrovirus vector.

Shown is the percentage of 17-day cultured CD34⁺PBSCs expressing gp91^phox comparing nontransduced normal CD34⁺PBSCs (open bar) with X-CGD CD34⁺PBSCs that had been subjected to 3 daily 7-hour transductions under growth-stimulating conditions (5 cytokines) using either 3rd-generation SIN lentivector–gp91^phox (dotted bar) or MFGS–gp91^phox (hatched bar). Cultured CD34⁺PBSCs were labeled with FITC-conjugated anti-gp91^phox and analyzed by flow cytometry, and the data were expressed as the percentage of cells that exhibit an expression of gp91^phox that is higher than the 95^thpercentile of isotype antibody control. By this criterion nontransduced X-CGD CD34⁺PBSCs do not give rise to any cells that are positive for the expression of gp91^phox (not shown). Normal control CD34⁺PBSCs differentiating in culture first gave rise to differentiated myeloid cells that expressed native gp91^phox by approximately day 8 of culture, and this increased to a steady state maximum by approximately day 15, which, in the example shown at day 17 of culture, demonstrated approximately 24% of cells positive for the expression of gp91^phox (open bar). Results are representative of 2 experiments.