Figure 5. Quantification of rAd5 association with LSEC and KC.

We assessed the whereabouts of rAd5 in 4-color fluorescence images like the representative ones shown in Fig 3, Fig 4 and Supplementary Figs S1 and S2, employing 4 combinations of complementary markers to identify cells and virus. By counting total virus color (pixel area x mean intensity) associated with both LSEC and KC, we found that 83–96% of virus was associated with LSEC and 4–17% with KC and none with hepatocytes, as shown in the bar graphs as mean ± SD for each of 4 different experiments. Data are graphed for both strategies for labeling virus, both Cy3-labeling (Cy3) and labeling with anti-Ad5 antibody (Ab). Two different markers identified both LSEC and KC. The table below the Fig illustrates that the 4 experiments were comparable by several parameters including relative fluorescence intensity scored RFI, numbers of mice used (mice), numbers of KC and sinusoid lumens scored (lumens), number of images examined, and the area of tissue examined microscopically (µm²).