In vitro determination of phagocyte activity by luminol- and lucigenin-amplified chemiluminescence

Abstract

Amplified chemiluminescence (CL) detects most sensitively biologically important reactive oxygen species (ROS) which are generated by phagocytes by the respiratory burst permitting the determination of cell activity in vitro. Different murine phagocyte populations were used in combination with various ROS-catabolizing enzymes and some of their inhibitors to determine the possible advantages of one of the two main presently used amplifiers, i.e. luminol and lucigenin. Lucigenin appeared to react mainly with the first of the generated ROS the superoxide anion radical (O-.2) and thus records cell activity via the respiratory burst much more reliable than luminol. The more commonly employed luminol reacts mainly with hydrogen peroxide (H2O2) and probably the singlet oxygen (1O2) which result in photon emission. However, it seems not to react with the hydroxyl radical (OH.). The dependence of luminol-amplified CL upon the generation of the chain reaction intermediate H2O2 and its three main catalysts catalase, myeloperoxidase and glutathione makes this reaction prone to different artifacts if cell activity is to be determined. Lucigenin-amplified CL offers great advantages to study cell activating or inhibiting properties of drugs and kinetics in vitro because of the biological relevance of O-.2 determination, its sensitivity, reproducibility and ease in handling.