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. 2020 Aug 5;10:13167. doi: 10.1038/s41598-020-69897-w

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© The Author(s) 2020

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 license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license 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 license, visit http://creativecommons.org/licenses/by/4.0/.

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Intranasal delivery of rOVA-3 subunit vaccine plus αGalCer elicits CD8⁺ TRM responses. B7.2^tg mice (n = 5) were primed and boosted i.n. as in Fig. 6. On day 30 after boost, mice were injected i.v. with anti-CD45.2-APC mAb. After 3–5 min., airway cells were collected by perfusing the lungs with PBS. Airway, lungs and spleen single cell suspensions were stained for surface markers and antigen specificity, and gated to identify B8R_70–78-specific, CD8⁺ TRM cells in the airways and lung interstitium (IST TRM), lung CD8⁺ MV TEM cells, and splenic TCM cells. Expression of surface activation markers was assessed on the indicated populations. Representative gating strategy (A), overlay histograms (B), and geometric mean fluorescent intensity (gMFI) of surface indicated markers on specified populations (C) are shown. Data are cumulative mean ± sem. Airway TRM (n = 2); and IST TRM; MV TEM, TCM: n = 5. ns, not significant (P > 0.05), *P ≤ 0.05, **P ≤ 0.001, ***P ≤ 0.0001.