Cell lines expressing epitope-tagged HSV-2 antigens: relative abundance of gD-, ICP8-, RR-1-, and VP5-specific antibodies in HSV-2 0ΔNLS antiserum.

<p>Western blots of (UI) uninfected Vero cells, cells inoculated with 2.5 pfu/cell of HSV-2 MS, or Vero cell lines that stably express the following, recombinant HSV-2 proteins: gD-FLAG, ICP8-FLAG, RR-1-FLAG, or VP5-FLAG incubated with 1:20,000 dilutions of serum from (<b>A</b>) naïve mice or (<b>C, E</b>) mice immunized with <b>(C)</b> gD-2 + alum/MPL adjuvant or <b>(E)</b> HSV-2 0ΔNLS. Following incubation with mouse serum, blots were rinsed and re-probed with mouse α-FLAG antibody to validate the relative amount of FLAG-tagged HSV-2 protein on each blot. (<b>B, D, F</b>) Normalized amount of IgG antibody bound to recombinant gD, FLAG, RR-1, or VP5 on blots incubated with (<b>B</b>) naïve mouse serum (n = 3), (<b>D</b>) gD-2 antiserum (n = 3), or (<b>F</b>) 0ΔNLS antiserum (n = 6). Levels of bound IgG antibody were normalized to account for blot-to-blot variance in the relative amount of each target based on the relative amount of α-FLAG antibody that bound each recombinant protein. In panel F, ‘**’ denotes that IgG antibody in 0ΔNLS antiserum bound RR-1 to significantly greater levels than gD, ICP8, or VP5 (p<0.001, one-way ANOVA and Tukey’s post-hoc-t-test).