FIG. 4.
Digital confocal image of HEp-2 cells infected with various viruses and fixed in methanol at 16 h.p.i. Using an MOI of 5, HEp-2 cells were infected with wild-type HSV-1 (WT), UL34 null mutant virus vRR1072(tk+) (34−), repaired UL34 mutant virus vRR1072Rep (34R), UL31 null mutant virus R5132 (31−), or a repaired UL31 mutant virus (R5132Rep) (UL31R). Cells were stained with UL31 protein-specific antiserum (green), UL34 protein-specific antiserum (red), and ICP4-specific ascites fluid (blue). Bound immunoglobulins specific for UL31 protein, UL34, and ICP4 by reaction with FITC-conjugated donkey anti-rabbit Ig, Texas Red-conjugated donkey anti-chicken Ig, and Cy5-conjugated goat anti-mouse Ig antibody, respectively. Cells were analyzed by confocal laser scanning microscopy by acquiring images through the middle of the cell. For illustrative purposes, Cy5 staining was pseudocolored blue in the figure. Areas of UL31 and UL34 staining colocalization are indicated by a yellow color upon merging of the separate images (Merge). Original magnification, X600.