Cytokine secretion from immunized mouse splenocytes.

<p>ELIspot assays were used to detect the proliferation of antigen-specific CD4+ secreting INF-γ (<b>A</b>) or IL-4 (<b>B</b>) cells as described in “<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0013972#s4" target="_blank">Materials and Methods</a>”. Splenocytes were isolated from immunized mice 4 weeks after the boosting immunization and seeded into pre-coated Multiscreen plates (Millipore). Defined peptides (MHC I or MHC II) or inactivated PR8 viruses were added to stimulate antigen-specific T cell proliferation. Cytokine secreting clones were visualized by spot staining following the manufacturer's instruction. Representative data were mean ± S.D. of spot number in 1×10⁶ splenocytes of six mice.