Immunometabolic analysis of primary murine Group 2 Innate Lymphoid Cells: a robust step-by-step approach

Group 2 Innate Lymphoid Cells (ILC2s) have recently been shown to exert key regulatory functions in innate and adaptive immune response networks that drive the establishment and progression of type 2 immunity and its associated pathologies. Although mainly tissue resident, ILC2s and their crosstalk within tissue microenvironments influences both local and systemic metabolism. In turn, the metabolic status shapes the diverse ILC2 phenotypes and effector functions. Hence, deciphering the metabolic networks of ILC2s is essential in understanding ILC2’s roles in health as well as pathophysiologies. Here we detail a framework of experimental approaches to study key immunometabolic states of primary murine ILC2s and link them to phenotypes and functionality. Utilizing flow cytometry, Single Cell ENergetIc metabolism by ProfilIng Translation inhibition (SCENITH) as well as the Seahorse platform we provide a framework that allows in-depth analysis of cellular bioenergetic states to determine the immunometabolic wiring of ILC2. Linking immunometabolic states and networks to ILC2 phenotypes and effector functions will allow in-depth studies that assess the potential of novel pharmaceutics to alter ILC2 functionality in experimental and clinical settings.