Precursor frequencies of subdominant influenza-specific CD8+ T cells by ELISpot. CD8+ T cells were obtained from control donor PBMCs and analyzed by ELISpot directly ex vivo. A. Shows a representative example of ELISpot wells. T2 cells were pulsed with either PB1413-421 peptide, M158-66 peptide, or no peptide. CD8+ T cells were plated at a 1:1 ratio with peptide pulsed T2 cells. B. Summary of IFNg secreting CD8+ T cells determined using an ELISpot assay. The results are an average mean of triplicates with background subtracted. All displayed data points are statistically significant (p < 0.05), with a cut off value of 5 SFC/100000 CD8+ T cells.