Fig. 4.
Protein tyrosine nitration as analyzed by Western blot analysis. (A and C) Lanes 1–5, control aortas (CON, n = 5); lanes 6–11, atherosclerotic aortas (HC, n = 6); all samples contained 20 μg of protein. (A) Polyclonal antiserum against NT recognized one specific band at 38 kDa (p38nt), which was greatly diminished in the atherosclerotic samples. Preincubation of the antibody for 60 min with 50 μM free NT completely blocked formation of the band (data not shown). Rabbit IgG heavy chains (50 kDa) were recognized by the secondary antibody. (B) Densitometric analysis of bands from A; data are represented as mean ± SEM (***, P < 0.001; AU, arbitrary units). (C) Monoclonal anti-NO2-BSA antibody recognized a single but different protein band at 63 kDa (p63nt), which was significantly increased in the atherosclerotic samples but not readily blocked by preincubation with free NT. (D) Densitometric analysis of bands from C (**, P < 0.01).