title: Chemiluminescence microscopy reveals functional heterogeneity in single neutrophils undergoing oxygen burst.
abstract: Activated polymorphonuclear leukocytes (PMN) respond to various triggers with an oxygen burst, during which the release of reactive oxygen species (ROS) plays a key role in microbial killing. The biological function of the ROS-associated light emissions is not known. However, this particularly weak cell-derived chemiluminescence (CL) may serve as a parameter for the identification of PMN activation. In this study we describe a novel technique which we termed CL microscopy. A microscope-based low-light image-processing system was applied which was sensitive enough to detect single photons, capable of two-dimensional signal accumulation, and digital image analysis. This technique permitted, for the first time, the visualization of the oxygen burst in single cells. Furthermore, quantitative evaluation of cell-derived luminol-enhanced CL revealed functional heterogeneity. Single-cell investigations of activated living PMN of normal human donors showed clear differences in kinetics and intensity of the oxygen burst related to different stimuli. The chemical agent (phorbol 12-myristate 13-acetate) induced CL in 83% of PMN. In contrast, the complement-mediated phagocytic stimulation by opsonized zymosan gave much higher light intensities of individual cells, but only in part of the PMN population (30%). CL microscopy presents a new and highly sensitive technique with considerable potential for single-cell analysis in immunological research.
authors: Institute of Clinical Immunology, Inselspital, Bern, Switzerland.
authors: R
authors: R
authors: Fritzsche
authors: A L
authors: AL
authors: De Weck
mesh_terms: D005609:Free Radicals; D006801:Humans; D007091:Image Processing, Computer-Assisted; D008163:Luminescent Measurements; D008853:Microscopy / Q000295:instrumentation / Q000379:methods*; D009504:Neutrophils / Q000145:classification* / Q000378:metabolism / Q000502:physiology; D010100:Oxygen / Q000378:metabolism
chemical_list: D005609:Free Radicals; D010100:Oxygen