Reconstitution of protein expression in mouse neutrophils by untagged, wt p40^phox. (A) Retroviral vectors used for mouse bone marrow reconstitution. Maps of empty vector pMIG R1-IRES-GFP (top) and a construct containing untagged, wt p40^phox (bottom). (B) Position of mutated residues in the primary protein structure of p40^phox. (C) Flow cytometric analysis of GFP expression in circulating neutrophils, as a measure of the efficiency of reconstitution of peripheral blood and bone marrow by transduced donor cells. Mouse FL cells on a p40^phox−/− background were transduced with retroviral supernatants and injected into irradiated recipients. Four weeks after injection, allowing sufficient time to repopulate the bone marrow compartment, peripheral blood samples were subjected to flow cytometric analysis of GFP-positive neutrophils, using Gr1 as an indicator for neutrophils. Each data point represents one individual mouse. Data are mean ± SEM. (D) Expression of p40^phox (top), p67^phox (middle), and p47^phox (bottom) in BMNs isolated from p40^phox−/− mice heterologously expressing GFP or untagged p40^phox (p40-IRES-GFP) or wt mice. Neutrophils were sonicated into SDS sample buffer, subjected to SDS-PAGE (4 × 10⁵ cells/lane), and immunoblotted for phox components as described in “Methods.” Shown is a representative blot of 2, performed in duplicate.