Abstract：
:Reactive oxygen species (ROS) are continuously produced in the cell as a consequence of aerobic metabolism, and are controlled by several antioxidant mechanisms. An accurate measurement of ROS is essential to evaluate the redox status of the cell, or the effects of molecules with the pro-oxidant or antioxidant activity. Here we report a cytofluorimetric technique for measuring simultaneously, at the single-cell level, hydrogen peroxide and superoxide anion, reduced glutathione (a main intracellular antioxidant) and cell viability. The staining is performed with the fluorescent dyes 2',7'-dichlorodihydrofluorescein diacetate (H2DCFH-DA), hydroethidine (HE), monobromobimane (MBB) and TO-PRO-3. This analysis is possible with new-generation flow cytometers equipped with several light sources (in our case, four lasers and an UV lamp), which excite different fluorochromes. This approach is extremely useful to study the balance between ROS content and antioxidants in cells receiving different stimuli, and to analyze the relationship between oxidative stress and cell death.
journal_name
Nat Protocjournal_title
Nature protocolsauthors
Cossarizza A,Ferraresi R,Troiano L,Roat E,Gibellini L,Bertoncelli L,Nasi M,Pinti Mdoi
10.1038/nprot.2009.189subject
Has Abstractpub_date
2009-01-01 00:00:00pages
1790-7issue
12eissn
1754-2189issn
1750-2799pii
nprot.2009.189journal_volume
4pub_type
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