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Additional file 2: Figure S1. Flow cytometry-gating strategy for splenocytes. Aged (18–22 months) BALB/c mice were immunized twice with H1-VLP, split-virion vaccine or naïve. Three weeks post-boost (6–24 mice/group), splenocytes were collected and stimulated ex vivo for 18 h with H1-VLP.

Additional file 3: Figure S2. Higher dose IM + IN vaccination: Effects on lung immune response 42 days post-vaccination, viral load 3 dpi and survival after A/California/07/2009 challenge. Aged (18–22 months) BALB/c mice were immunized twice with H1-VLP, split-virion vaccine or PBS. Lungs were collected six weeks after vaccination. Percent of lung A) CD8+ T cells and expressing IFNγ presented as background subtracted (stimulated - unstimulated). Fold-change of tissue-resident B) CD4+ T cells from the PBS group. For statistical analysis, two-way ANOVA was performed followed by Tukey’s multiple comparison test (** p 20% of their initial weight. A log-rank (Mantel-Cox) test was used to compare survival curves with the PBS control group. (** p

Additional file 1: Table S1. Frailty Index measurements for day 0 and day 25 post-infection. Table S2. Statistics comparing groups for weight loss, using Tukeyâ s multiple comparison test. Table S3. Percent of mouse splenocyte T cells (CD4+ or CD8+) above the PBS average compared to the vaccinated animals

Additional file 3: Figure S2. Higher dose IM + IN vaccination: Effects on lung immune response 42 days post-vaccination, viral load 3 dpi and survival after A/California/07/2009 challenge. Aged (18–22 months) BALB/c mice were immunized twice with H1-VLP, split-virion vaccine or PBS. Lungs were collected six weeks after vaccination. Percent of lung A) CD8+ T cells and expressing IFNγ presented as background subtracted (stimulated - unstimulated). Fold-change of tissue-resident B) CD4+ T cells from the PBS group. For statistical analysis, two-way ANOVA was performed followed by Tukey’s multiple comparison test (** p 20% of their initial weight. A log-rank (Mantel-Cox) test was used to compare survival curves with the PBS control group. (** p

Additional file 2: Figure S1. Flow cytometry-gating strategy for splenocytes. Aged (18–22 months) BALB/c mice were immunized twice with H1-VLP, split-virion vaccine or naïve. Three weeks post-boost (6–24 mice/group), splenocytes were collected and stimulated ex vivo for 18 h with H1-VLP.