Quantification of heterotypic granule fusion in human neutrophils by imaging flow cytometry.

摘要: Human neutrophils are filled with intracellular storage organelles, called granules and secretory vesicles, which differ in their content of soluble matrix proteins membrane-bound molecules. To date, at least four distinct granule/vesicle subsets have been identified. These organelles may secrete extracellularly following mobilization to fusion the plasma membrane, but some them also fuse internal membrane-enclosed typically a membrane-derived phagosome. There instances where different appear one another, process that would enable mixing membrane components. Such granule enables e.g., myeloperoxidase-processing intragranular oxygen radicals, key event formation neutrophil extracellular traps (Bjornsdottir et al., 2015) [1]. Described herein data show quantification such heterotypic granule-granule by use imaging flow cytometry, technique combines cytometry microscopy. The analysis described is based on immunofluorescent staining established markers (lactoferrin and/or NGAL for subset; specific granules, CD63 another subset, azurophil granules) calculation colocalization score resting PMA-stimulated neutrophils.