Respiratory Burst Activity
Generation of ROS was evaluated by flow cytometry using the technique of
Smith and Wiedemann22. Cells were incubated with DHR
123 (50 μL) and PBS (450 μL) at 37°C for 10 minutes. Cells were
stimulated with 1 μL (16 μM) phorbol 12-myrisate 13-acetate (PMA) for 20
minutes at 37°C. The reaction was then halted by placing samples on ice.
Fluorescence intensity was assessed by flow cytometry and expressed as
mean fluorescent intensity (MFI). DHR 123 has been shown to detect
intracellular H2O2 and OH radical
production22.