Assessing lineage and cytolytic functional potential of murine tissue-resident innate lymphocytes

Group 1 innate lymphocytes are heterogeneous, and their ontogeny and function remain ambiguous. Here, we describe a protocol to measure cell ontogeny and effector functions of natural killer (NK) and ILC1 subsets based on current understanding of their differentiation pathways. We use cre drivers to genetically fate-map cells, tracking plasticity between mature NK and ILC1. We describe innate lymphoid cell precursor transfer studies that determine ontogeny of granzyme-C-expressing ILC1. Additionally, we detail in vitro killing assays that test cytolytic potential of ILC1s. For complete details on the use and execution of this protocol, please refer to Nixon et al. (2022).1 [Display omitted] •Three experiments that together address group 1 innate lymphocyte ontogeny and function•Genetic fate mapping of mature effector cells explores potential further differentiation•Innate lymphoid cell (ILC) precursor transfer differentiates NK cells from ILC1•Killing assay demonstrates cell subset potential for perforin-dependent cytotoxicity Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. Group 1 innate lymphocytes are heterogeneous, and their ontogeny and function remain ambiguous. Here, we describe a protocol to measure cell ontogeny and effector functions of natural killer (NK) and ILC1 subsets based on current understanding of their differentiation pathways. We use cre drivers to genetically fate-map cells, tracking plasticity between mature NK and ILC1. We describe innate lymphoid cell precursor transfer studies that determine ontogeny of granzyme-C-expressing ILC1. Additionally, we detail in vitro killing assays that test cytolytic potential of ILC1s.