PtDd release from infected cells and effect on epithelial junctions.

T84 cells were cultured in transwell chambers until the TEER was constant, i.e. epithelial junctions had formed. A) Confocal immunofluorescence microscopy of T84 cells. The left panel shows an XY image of the cell surface. The right panel shows stacked XZ images. Antibodies against E-cadherin (red) and DSG2 (green) visualize epithelial junction proteins. B) Release of PtDd from mu-Ad3GFP or wt-Ad3GFP-infected T84 cells. Polarized T84 cells were infected with mu-Ad3GFP or wt-Ad3GFP at an MOI of 10 vp/cell (added into the inner chamber) for 2 hours. 24 hours post infection, cells were fixed in 4% PFA for 15 minutes and then permeabilized with cold methanol for 10 minutes. Immunofluorescence staining was performed with anti-Ad3 PtDd polyclonal antibody (red). Nuclei were counterstained with DAPI (blue). C) Cells were infected as described for B). Cells were stained for the junction protein E-cadherin (red) 72 hours pos-tinfection. Virally infected cells express GFP (green). The scale bar of all images is 20 µm.