p67phox

P67PHOX is a 6-element fingerprint that provides a signature for neutrophil cytosol factor 2 (p67phox). Regulation of the respiratory burst takes place at the phagocytic vacuole. The process is mediated by NADPH oxidase, which transports electrons across the plasma membrane to form superoxide (an oxygen molecule with an extra electron that is toxic to normal cells) in the vacuole interior. The electrons are carried across the membrane by a short electron transport chain in the form of an unusual flavocytochrome b(1).\nNADPH Oxidase is comprised of flavocytochrome b558 (made up of p22 phox and gp91 phox), p40 phox, p47 phox and p67 phox. NADPH Oxidase mediates the final steps of electron transfer to oxygen, resulting in the generation of superoxide ion. A growing body of evidence suggests a potential role for oxygen-derived radicals such as superoxide anions and hydrogen peroxide as intracellular signaling molecules that may be involved in cell growth, apoptosis and cellular senescence. To conserve NADPH and avoid the toxic effects of superoxide, the oxidase remains inactive until it receives an appropriate stimulus. Activation involves the participation of a number of cytosolic proteins, some of which attach to the flavocytochrome. P47phox, p67phox and p40phox are specialized components of phagocytic cells, all of which contain SH3 domains, by means of which they attach to proline-rich regions of other proteins. Upon activation, p47phox and p67phox are phosphorylated and, with p40phox, translocate to the region of the plasma membrane forming the phagocytic vacuole, where they associates with hydrophilic regions of the flavocytochrome (1).\n\nApplications: \nSuitable for use in Western Blot, Immunoprecipitation and Immunocytochemistry. Other applications not tested.\n\nRecommended Dilution:\nWestern Blot: 1:500-1:2000 detects p67-phox in RIPA lysates from HL-60 cells. Note: p67-phox is highly susceptible to proteases. \nImmunoprecipitation: 4ul immunoprecipitated p67-phox from 500ug of HL-60 RIPA lysate. \nImmunocytochemistry: 1:500-1:4000 shows positive cytosolic immunostaining for p67-phox in HL-60 cells fixed with 4% paraformaldehyde and permeabilized with 0.1% Triton X-100.\nOptimal dilutions to be determined by the researcher. \n\nRecommended Secondary Antibodies:\nI1904-39 IgG, X-Adsorbed (HRP) Pab Gt x Rb\nI1904-40A IgG, H&L, X-Adsorbed (HRP) Pab Gt x Rb\nI1904-40P IgG, Fc, X-Adsorbed (HRP) Pab Gt x Rb\n\nStorage and Stability:\nMay be stored at 4 degrees C for short-term only. For long-term storage and to avoid repeated freezing and thawing, add sterile 40-50% glycerol, aliquot and store at -20 degrees C. Aliquots are stable for at least 12 months at -20 degrees C. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap. Further dilutions can be made in assay buffer.

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