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. 2024 Nov 28;14:29609. doi: 10.1038/s41598-024-80636-3

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© This is a U.S. Government work and not under copyright protection in the US; foreign copyright protection may apply 2024

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 4 — Adjuvanted S1 protein vaccine efficacy in aged mice. Aged (> 18 months old) B6 (WT) mice were immunized with S1 protein alone or S1 protein co-delivered IL-15 with TLR-Ls. Humoral immune responses were assessed following immunization by analysis of total serum IgG responses to (A) S1, (B) ectodomain, or (C) RBD of SARS-CoV-2 spike protein over time evaluated by ELISA. (D) Serum antibody neutralization potencies at days 14, 28, and 50 are presented as SNI% in 1:30 dilutions of sera against RBD-hACE2 binding. The dashed horizontal line corresponds to the lower limit of detection (LLOD = 20%) of the assay used. The data are represented as the mean ± SEM and represent at least two independent experiments. p values are shown for entirety of study using one (D) or two-way ANOVA (A-C) test with post hoc Tukey’s multiple comparisons; *, p < 0.05; **, p < 0.01; ***, p < 0.001; ****, p < 0.0001. Cellular immune responses were assessed at day 75 post-immunization by stimulating spleen cells ex vivo with either vehicle control or overlapping peptide pools comprising the RBD protein at 0.1 µg/ml of all peptides for 6 h and then assessed by flow cytometry for RBD-tetramer⁺ CD8⁺ T cells and intracellular production of IFNγ in CD8⁺ T cells. (E) Representative flow cytometry contour plots and (F) graphed values of individual mice. *, p < 0.05; **, p < 0.01; ***, p < 0.001; ****, p < 0.0001 determined using a paired two-tailed Student’s t-test.