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

Objective: The aim of this study was to verify in our laboratory conditions the performance criteria of a commercial kit (Phagoburst^TM, 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 intra-assay 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.