Figure 3.

Influenza sa-mRNA bicistronic A/H5N1 vaccines elicit robust cellular immune responses in BALB/c mice

Female BALB/c mice, 8–10 weeks old, were immunized (10 mice/group) on days 1 and 22 with bilateral 50 μL intramuscular injections in the rear quadriceps. Spleens were collected from euthanized animals on day 43. The frequency of intracellular cytokine expression among CD4+ (A) and CD8+ (B) in response to stimulation is displayed. Spleens from five mice/group were pooled and processed, and splenocytes were stimulated with anti-mouse CD28 in the absence or presence of synthetic peptides representing immunodominant epitopes in the H5 or N1 proteins encoded by the vaccine. Cells were stained with fluorescently tagged antibodies to cell surface markers CD3, CD4, CD8, and intracellular cytokines interleukin-2 (IL-2), tumor necrosis factor α (TNFα), interferon γ (IFNγ), IL-5, and IL-13 were analyzed by flow cytometry. Each column represents a mean from duplicate cultures of the five pooled spleens.