Cytofluorimetric analysis of transduced splenocytes in SCID mice.

Expression of cell differentiation antigens by cytofluorimetric analysis on gated GFP⁺ splenocyte population from SCID mice shown in Figure 7C. The black area in the overlaid FACS profiles corresponds to the expression of specific membrane markers on the basis of immunostaining by means of PE-conjugated antibodies specific for each of these different membrane markers (CD54, CD11c, CD80, MHC class II, F4/80, CD11b, CD86, CD62E, CD31) or via indirect staining (CD105 and CD106). Controls included gated GFP⁺ cells that were stained with an isotype-matched antibody specific for an irrelevant antigen (TNP) either directly conjugated with PE or used in conjunction with a secondary PE-conjugated antibody (as control for CD105 and CD106) (gray contours). The percentage of positive cells was determined with the use of appropriate FACS software by subtracting the background profiles from the experimental profiles. Consequently, only the fluorescent cells that fall outside the background area (gray contour) were counted and considered positive. The mean percentage of fluorescent cells ± SD was mentioned, and one representative profile for each of the markers was depicted. Analysis was performed 1 week after transduction.