Characterization of lung T cells from young and middle-aged mice.

<p>BALB/cAnNCr mice were immunized at 2 months (A, C, and E) or 11 months old (B, D, and F) as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0153195#pone.0153195.g001" target="_blank">Fig 1</a> or left unvaccinated (naïve). Lung T cells were prepared and analyzed by flow cytometry. (A, B) Total numbers of K^d-NP_147–155-tetramer⁺ CD8⁺ T cells recovered from the lungs 12–13 weeks after vaccination were determined by multicolor flow cytometry. Bars show mean ± SEM of 3–4 animals per group for the two-month-old mice (A). For 11- month-old mice, cell yield after Ficoll was low and lung cells had to be pooled to permit analysis (B). (C, D) Lung CD8⁺ T cell phenotypic analysis: Proportions of activated effector memory (CD62L^lo, CD127^lo), effector memory (CD62L^lo, CD127^hi) and central memory (CD62L^hi) among tetramer⁺ CD8⁺ T cells from A/NP+M2 vaccinated animals are displayed as pie charts. (E, F) Activation status of pooled lung CD8⁺ T cells characterized by positive staining for both tetramer and CD69 marker. Fluorescence minus one (FMO) controls were used to identify gating boundaries and adjusted based on internal negative controls for each treatment group. Percent of total CD8⁺ T cells are displayed per quadrant.