Mucosa associated invariant T (MAIT) cells belong to the unconventional T cell compartment, are enriched in mucosal sites and their effector phenotype allows rapid activation upon antigen encounter. They have a semi-invariant αβTCR that recognizes riboflavin pathway metabolites presented on the monomorphic Major histocompatibility complex (MHC)-Ib-related molecule MR1. MAIT cells have important antimicrobial functions against pulmonary infections and a role in maintaining barrier integrity. However, many aspects of the immunobiology of human lung MAIT cells are still unexplored. In this project, we aimed to characterize the phenotype and function of MAIT cells in the lung and lung-associated lymph nodes (LN) from human organ donors. Flow cytometry data show that MAIT cells are present in the lung and lung LN and express tissue residency markers CD69 and CD103 in contrast to paired blood samples. MAIT cells in the lung and lung LN have distinct phenotypic profiles. PMA/ionomycin stimulation induces a stronger and more polyfunctional response in LN MAIT cells as compared to lung MAIT cells, resembling their blood counterparts in that respect. TCR stimulation is sufficient to induce cytokine response and granzyme B production in LN MAIT cells, while in the lung MAIT cell responses rely more on the cytokine milieu. The cytokine response in the lung is further enhanced by inflammatory cytokines in addition to the TCR stimulus. In summary, these data indicate that lung LN and lung tissue MAIT cells display a differential profile and sensitivity to stimuli that reflects location and adaptation to non-lymphoid versus lymphoid tissue sites.