FIGURE 3.

GMM is presented by CD1c to T cells in macaques. (A) PBMC from an Indian-origin rhesus macaque that was vaccinated intradermally with BCG were stained with Mamu CD1c-GMM (PE) and Mamu CD1c mock-loaded (BV421) tetramers to identify CD1c-GMM–specific T cells. (B) T cells staining with Mamu CD1c-GMM but not Mamu CD1c mock-loaded tetramer were sorted and expanded in vitro for 2 wk to generate a T cell line (cG52R). (C) The species origin of cG52R was confirmed by staining with an anti-CD3ε Ab (clone UCHT1) that does not bind Mamu CD3ε as well as an anti-CD3ε Ab (clone SP34-2) that binds CD3ε from both species. (D) T cells staining with Mamu CD1c-GMM tetramer present within cG52R were further examined for coreceptor expression using Abs targeting CD4 and CD8α. (E) cG52R was simultaneously stained with Mamu CD1c-GMM tetramer (G780) and H. sapiens CD1c-GMM tetramer (PE). Fewer than 5% of T cells staining with Mamu CD1c-GMM tetramer costain with H. sapiens CD1c-GMM tetramer. (F) cG52R was coincubated with K562-CD1c or K562-EV APCs in the presence or absence of GMM. IFN-γ detection by ELISPOT demonstrates the Mamu-derived T cell line is partially activated by H. sapiens CD1 proteins. Data are representative of at least three (A–E) or two (F) independent experiments.