Autofluorescence enables detection and quantification of EETosis in human leukocyte cultures

Eosinophils are leukocytes involved in defense against multicellular parasites and other pathogens. Like neutrophils, eosinophils can undergo a cytolytic form of cell death known as eosinophil extracellular trap cell death (EETosis), during which they release DNA and cytoplasmic granules to form eosinophil extracellular traps (EETs). Here, we demonstrate that eosinophil granules released during EETosis exhibit distinct autofluorescence in both purified human eosinophil cultures and mixed leukocyte cultures. This autofluorescence allowed detection of EETosis in mixed leukocyte cultures without additional staining or eosinophil isolation. We further show that 2 vanilloid compounds, previously identified as inhibitors of NETosis, suppress phorbol 12-myristate 13-acetate-induced EETosis. By measuring the autofluorescence intensity of released eosinophil granules, we successfully quantified the inhibitory effects of these compounds on EETosis. Our autofluorescence-based assay enables rapid and convenient detection of EETosis and provides a useful tool for in vitro studies of eosinophil biology.