Simultaneous evaluation of phagocytosis and Fc gamma R-mediated oxidative burst in human monocytes by a simple flow cytometry method.
In this study we describe a fast, simple, and objective flow cytometry method to quantify simultaneously phagocytosis and Fc gamma R-mediated oxidative burst in human monocytes. Peripheral blood monocytes isolated by elutriation were sequentially incubated with 2',7'-dichlorofluorescein-diacetate (DCFH-DA) and sheep erythrocytes opsonized with rabbit IgG (SRBC-IgG), and then analysed by flow cytometry. Two parameters were studied simultaneously in the same cell population: forward scatter modifications (grade of phagocytosis) and fluorescence intensity of DCFH oxidation (index of oxidative burst). We found a statistically significant correlation (r = 0.96274) between the forward scatter increase and the amount of phagocytosis as determined by light microscopy. We also found a significant correlation between the oxidative burst and phagocytosis studied both microscopically (r = 0.7714) and by flow cytometry (r = 0.78056). As the presence of DCFH did not impair monocyte phagocytosis, we conclude that this simple method could be a useful tool in functional studies of monocytes.
Casado
J A
JA
Department of Immunology, Clinica Universitaria, University of Navarra, Pamplona, Spain.
Merino
J
J
Cid
J
J
Subira
M L
ML
Sanchez-Ibarrola
A
A
D000818:Animals; D005434:Flow Cytometry / Q000379:methods*; D005452:Fluoresceins; D006801:Humans; D009000:Monocytes / Q000276:immunology / Q000502:physiology*; D010587:Phagocytosis; D017452:Receptors, IgG / Q000502:physiology*; D016897:Respiratory Burst; D012756:Sheep
D005452:Fluoresceins; D017452:Receptors, IgG; C037631:2',7'-dichlorofluorescein