Severe allergic asthma is characterized by the presence of eosinophils in the airways and systemic eosinophilia. As a therapeutic strategy, eosinophils are depleted with biologics, although the long-term safety of depleting eosinophils is currently unknown. Using mouse models of allergic asthma, two subsets of eosinophils have been identified; "inflammatory" eosinophils expressing CD101 and "homeostatic" lung-resident CD101-negative eosinophils. We have discovered the dynamic transition of three waves of eosinophil differentiation during the allergic response, where "homeostatic" become "inflammatory" in consecutive eosinophil waves of different time-dependent origins; 1. lung-resident 2. blood-derived and 3. bone marrow-derived. This is supported by spatio-temporal single cell RNA sequencing of eosinophils. At the moment, it is completely unknown in what way these waves differ and contribute to the inflammatory response across time. In the proposal, I will investigate the transcriptome, proteome and metabolic program of eosinophils in the lung during allergic inflammation. I will make use of the newest and complementary single cell technologies, including CITEseq, spatial transcriptomics, high dimensional metabolic single cell profiling and genetic knockout using eosinophil-specific transgenic animals. These findings will lead to a more detailed understanding of eosinophil functioning in allergic lung inflammation and provide novel targets for immune therapy.