Single-Cell Optical Imaging of the Phagocyte NADPH Oxidase
- 1 September 2006
- journal article
- research article
- Published by Mary Ann Liebert Inc in Antioxidants and Redox Signaling
- Vol. 8 (9-10), 1509-1522
- https://doi.org/10.1089/ars.2006.8.1509
Abstract
The phagocyte NADPH oxidase is a key component of the innate immune response against invading microorganisms, because the generation of superoxide (O2−) inside the phagocytic vacuole by this enzyme is responsible for microbial killing by mechanisms that are directly or indirectly dependent on reactive oxygen species (ROS) formation. Most of what is known about the membrane-embedded and cytosolic NADPH oxidase subunits and their intricate network of interactions on assembly and activation has been derived from biochemical and biophysical studies involving subcellular fractionation or reconstituted cell-free systems. Such investigations can be complemented by single-cell microscopy on phagocytes, which may reveal spatial and/or temporal details about NADPH oxidase assembly that cannot be obtained from fractionated-cell assays. In recent years, we have investigated the NADPH oxidase in neutrophils using two complementary optical imaging techniques: Raman microscopy, a vibrational spectroscopic technique that does not require protein labeling, and live-cell fluorescence microscopy, which sheds light on the dynamics of NADPH oxidase assembly in individual cells. Here, we briefly introduce these techniques, compare their characteristics, and show their potential for studying NADPH oxidase at the single-cell level. New microscopy data are presented to illustrate the versatility of Raman and fluorescence microscopy on intact neutrophils.Keywords
This publication has 98 references indexed in Scilit:
- FOURIER TRANSFORM INFRARED VIBRATIONAL SPECTROSCOPIC IMAGING: Integrating Microscopy and Molecular RecognitionAnnual Review of Physical Chemistry, 2005
- Infrared spectroscopic imaging for histopathologic recognitionNature Biotechnology, 2005
- Expression of gp91phox/Nox2 in COS-7 cells: cellular localization of the protein and the detection of outward proton currentsBiochemical Journal, 2005
- Neutrophil NADPH Oxidase Is Reduced at theAnaplasma phagocytophilumPhagosomeInfection and Immunity, 2004
- Anaplasma phagocytophilumUtilizes Multiple Host Evasion Mechanisms To Thwart NADPH Oxidase-Mediated Killing during Neutrophil InfectionInfection and Immunity, 2004
- Unique targeting of cytosolic phospholipase A2 to plasma membranes mediated by the NADPH oxidase in phagocytesThe Journal of cell biology, 2003
- Creation of a genetic system for analysis of the phagocyte respiratory burst: high-level reconstitution of the NADPH oxidase in a nonhematopoietic systemBlood, 2002
- The p67 Activation Domain Regulates Electron Flow from NADPH to Flavin in Flavocytochromeb 558Journal of Biological Chemistry, 1999
- Spectroscopic identification of the heme axial ligation of cytochrome b558 in the NADPH oxidase of porcine neutrophilsFEBS Letters, 1995
- Kinetics of fusion of the cytoplasmic granules with phagocytic vacuoles in human polymorphonuclear leukocytes. Biochemical and morphological studies.The Journal of cell biology, 1980