Figure 4.

Vaccination-induced memory cells have increased functional potential after in vivo recall

PBMCs were analyzed pretransfer for the frequency of tetramer⁺ cells. Cells were divided into two equal fractions and transferred to NSG-HLA:A^∗02 transgenic mice. On the same day, animals were infected i.p. with 2 × 10⁵ PFU mCMV-COVID or mCMV-FLU. The next day, mice were injected i.p. with human IL-2 (100 ng/mouse). 7 days post-infection, donor cells in peritoneal exudate cells (PECs) were analyzed (n = 25).

(A) Experimental setup.

(B) Line graph showing expansion of cells of the same donor as fold increase over pretransfer, comparing C19- with FLU-specific CD8⁺ T cells.

(C) Representative FACS plots of CD8⁺ T cell stained with HLA-A^∗02 tetramers pretransfer or 7 days post-infection. Non-HLA-A^∗02 cells are included as negative control. Numbers indicate percentages. Gated is for live hCD45⁺hCD8⁺ cells.

(D) Fold increase of antigen-specific cells segregated by C19 groups.

(E) Correlation between time after last antigen exposure and the expansion of virus-specific cells after mCMV-C19 infection.

(F–G) On day 7 post-infection, PECs were re-stimulated in vitro with PMA/IONO for 4 h in the presence of Brefeldin A and analyzed by flow cytometry. Shown are (F) cytokine production of live hCD45⁺hCD8⁺tetramer⁺ cells and (G) representative FACS plots gated for hCD45⁺hCD8⁺tetramer⁺ cells. Indicated are means ± SEM.

p values were calculated using paired Student t test (B) and Kruskal-Wallis rank-sum test with Dunn’s post hoc test for multiple comparisons (F). ^∗p < 0.05, ^∗∗p < 0.01, and ^∗∗∗p < 0.001.

See also Figure S4.