NADPH Oxidases pp 461-472 | Cite as

Isolation of Redox-Active Endosomes (Redoxosomes) and Assessment of NOX Activity

  • Weam S. Shahin
  • John F. EngelhardtEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 1982)


Reactive oxygen species (ROS) convey signals essential for proliferation, maintenance, and senescence of a growing list of cell types. Compartmentalization of these signals is integral to cell viability as well as the signaling pathways ROS direct. Redox-active endosomes (redoxosomes) are formed downstream of several ligand-activated receptors. NADPH oxidase (NOX) is a main component of redoxosomes, which recruits multiple proteins (Rac1, NOX2, p67phox, SOD1). Isolation of redoxosomes and evaluation of how superoxide (O2˙) production directs receptor signaling at the level of the endosome have enabled a better understanding of biologic processes controlled by ROS. In this chapter, we will first review the major signaling pathways that utilize redoxosomes and components that control its redox-dependent functions. We will then outline biochemical and biophysical methods for the isolation and characterization of redoxosome properties.

Key words

ROS Redoxosomes Iodixanol Immuno-affinity isolation Lucigenin EPR NOX Rac1 TRAF 



This work was supported by NIH grant R24 DK096518 (to J.F.E.).


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Authors and Affiliations

  1. 1.Department of Anatomy and Cell BiologyCarver College of Medicine, University of IowaIowa CityUSA
  2. 2.Department of Internal MedicineCarver College of Medicine, University of IowaIowa CityUSA
  3. 3.Center for Gene TherapyCarver College of Medicine, University of IowaIowa CityUSA

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