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. 2022 Jun 23;7:61. doi: 10.1038/s41541-022-00483-z

Fig. 4. Ad5.F35+Lm enhances the avidity and polyfunctionality of the GUCY2C-specific CD8+ T-cell pool.

Fig. 4

BALB/cJ mice (n = 5–8/group) were immunized using Ad-GUCY2C (prime) or Ad-GUCY2C + Lm-GUCY2C (prime-boost) vaccination regimens. At the peak effector response, (14 days following prime and 6 days following prime-boost vaccination), splenocytes were collected and stimulated with GUCY2C254-262 peptide to assess GUCY2C-specific CD8+ T-cell avidity quantified by IFNγ ELISpot (a) and polyfunctionality quantified by flow cytometry (b–d). a Non-linear regression (solid line) of GUCY2C-specific CD8+ T cell avidity is depicted with 95% confidence intervals (dashed lines). b The proportion of live CD8+ T cells staining positive for IFNγ, TNFα, or MIP1α cytokines after stimulation with GUCY2C254-262 peptide is shown. c, d Of cytokine+ CD8+ T cells, the percentage of cells staining positive for two and three cytokine-markers (c) as well as overall polyfunctionality (d) are shown. Polyfunctionality comparisons were made using two-sided T-tests comparing prime vs prime-boost with Holm-Sidak correction for multiple comparisons. Error bars indicate mean +/− SEM. Representative FACS plots are shown in Supplementary Fig. S8.