Extended Data Fig. 4: Monoclonal donor T_RM cells maintain resident phenotype after adoptive transfer.

Congenically marked HR OT-I T_RM (green) and HR OT-I T_CM cells (blue), that were FACS-purified from SI IEL and spleen of L.m.-OVA -infected mice, respectively, were co-transferred into naïve recipients, and distribution and phenotype of donor cells were analyzed 14 days later. a, Dot plot shows distribution of HR OT-I T_RM and T_CM cells within the total donor cell population (left panel) and histograms display expression of phenotypic markers by OT-I T_RM and T_CM cells (right panel) prior to transfer. b, Graphic scheme depicts setting of adoptive transfer experiments. c, Representative flow cytometry plots show expression of the congenic markers CD45.1 and CD45.2 to identify donor T_RM-derived (green), donor T_CM-derived (blue) and host-derived (grey) cells within the memory (CD44^high) CD8⁺ T cell population in the indicated tissues at two weeks after transfer. d, The frequency of T_RM- and T_CM-derived CD44^high cells was determined in indicated tissues. Dotted line indicates detection limit, as determined by analysis of non-transferred mice. e, Representative flow cytometry plots show expression of CD62L and CD69 on CD44^high T_RM and T_CM donor cells in liver. f, The frequency of CD69^– CD62L⁺ cells within the donor populations was quantified. Two-tailed paired t-test, *P = 0.0173. g, Representative flow cytometry plots show expression of tdTomato and CD69 on CD44^high T_RM and T_CM donor cells in liver. h, The frequency of CD69⁺ tdTomato⁺ cells within the donor populations was quantified. Two-tailed paired t-test, ***P = 0.0005. Combined data from two independent experiments (n = 4 mice). Symbols represent individual mice; bars represent the mean. Error bars represent mean ± SEM.