Human leukocyte phagocytosis of zymosan particles measured by flow cytometry

Human leukocyte phagocytosis of fluorescein-isothiocyanate (FITC)-labelled zymosan particles was studied by a flow cytometric (FCM) assay allowing discrimination of adhered and ingested zymosan particles. Free zymosan particles, non-phagocytes and phagocytes could be discriminated and quantified by simultaneous registration of fluorescence and light scatter. All leukocytes capable of phagocytosis were phagocytosing, and within 15 min 80% of the zymosan particles were adhered or ingested. Compared to the FITC-fluorescence of free zymosan particles, the mean fluorescence of phagocyte-associated zymosan particles was reduced by about 35%, indicating ingestion and processing of zymosan particles. Abolishing the FITC-fluorescence of extracellular zymosan particles by crystal violet, the number of zymosan particles adhered and ingested could be calculated from FCM measurements of phagocyte fluorescence. This showed that in 15 min 83% of the phagocyte-associated zymosan particles were actually ingested.