Several studies with purified enzymes have suggested that nitration of critical tyrosine residues adversely affects enzyme activity, but there is as yet no conclusive evidence in vivo for biological or cellular changes as a direct result of tyrosine nitration [16, 20] . For instance, tyrosine nitration was suggested to have an effect on cellular pathways by affecting cytoskeletal proteins or tyrosine phosphorylation, thereby affecting processes involved in, for example, cell proliferation or differentiation [16, 26] . Recent studies have provided support for selective tyrosine nitration within certain proteins [27, 28] and of selective cellular targets for nitration by RNS (see, for example, [29, 30] ), and such specificity might indicate a potential physiological role for this protein modification. However, in none of these cases could tyrosine nitration be linked directly to changes in enzyme function. Chemical studies have indicated that tyrosine nitration by RNS accounts for only a minor fraction of oxidant involved, and reactions with other biological targets (thiols, selenoproteins, or transition metal ions) are much more prominent [5, 6] . Indeed, the extent of tyrosine nitration in vivo is very low (1-1000 per 10 6 tyrosine residues according to best estimates [16]), although different analytical methods used to detect 3-nitrotyrosine in biological systems have often given inconsistent results. It is important to note that recent rigorous studies have unveiled substantial sources of artifact during sample preparation, which might frequently have led to an overestimation of tyrosine nitration in vivo in previous studies [31] .