Flow Cytometry Assessment of Bacterial and Yeast Induced Oxidative Burst in Peripheral Blood Phagocytes

Abstract

Abstract Objective: The aim of this study was to verify in our laboratory conditions the performance criteria of a commercial kit (PhagoburstTM, Glycotope Biotechnology) as described by the producers. We have also partially altered the use of the available kit by introducing a non-opsonized Candida albicans stimulus, in addition to the opsonized Escherichia coli stimulus provided by the manufacturer. Material and methods: The peripheral blood samples of 6 clinically healthy adults were tested in triplicate according to the manufacturer recommendations. The intraassay imprecision as well as the ranges of neutrophil and monocyte burst activation triggered by various stimuli were assessed. Results: The activation range of granulocytes and monocytes was similar to the one described by the producer in the presence of E. coli (granulocytes: 78.45-99.43% versus 99.6-99.95%, average %CV of 1.53% versus 0.1%, monocytes: 54.63-92.33% versus 81.80-96.67, average %CV 6.92% versus 1.1%). The leukocyte range of activation in the presence of non-opsonized C. albicans was comparable to the one triggered by the fMLP (N-formyl-methionyl-leucyl-phenylalanine) stimulus. Conclusion: The intra-assay precision obtained in our laboratory conditions, as well as the ranges of activated leukocytes, are comparable to the ones described by the producer when using E. coli as a stimulus. The present study shows that introducing an extra fungal stimulus for burst oxidation assessment could provide additional information regarding the non-specific cellular immune response, particularly in patients at risk for candidemia.