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J Vis Exp. 2014 Jul 7;(89). doi: 10.3791/51328.

Analysis of oxidative stress in zebrafish embryos.

Author information

1
Department of Molecular Biotechnology and Health Science, University of Torino.
2
Department of Molecular Biotechnology and Health Science, University of Torino; Laboratory of Endothelial Molecular Biology, Vesalius Research Center, VIB; massimo.santoro@unito.it.

Abstract

High levels of reactive oxygen species (ROS) may cause a change of cellular redox state towards oxidative stress condition. This situation causes oxidation of molecules (lipid, DNA, protein) and leads to cell death. Oxidative stress also impacts the progression of several pathological conditions such as diabetes, retinopathies, neurodegeneration, and cancer. Thus, it is important to define tools to investigate oxidative stress conditions not only at the level of single cells but also in the context of whole organisms. Here, we consider the zebrafish embryo as a useful in vivo system to perform such studies and present a protocol to measure in vivo oxidative stress. Taking advantage of fluorescent ROS probes and zebrafish transgenic fluorescent lines, we develop two different methods to measure oxidative stress in vivo: i) a "whole embryo ROS-detection method" for qualitative measurement of oxidative stress and ii) a "single-cell ROS detection method" for quantitative measurements of oxidative stress. Herein, we demonstrate the efficacy of these procedures by increasing oxidative stress in tissues by oxidant agents and physiological or genetic methods. This protocol is amenable for forward genetic screens and it will help address cause-effect relationships of ROS in animal models of oxidative stress-related pathologies such as neurological disorders and cancer.

PMID:
25046434
PMCID:
PMC4212721
DOI:
10.3791/51328
[Indexed for MEDLINE]
Free PMC Article

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