FIGURE 3.

ApoA-I nitrated on tyrosine 166 present in human atherosclerotic lesions is not located on HDL-like particles. Proteins in atherosclerotic lesion homogenate or after sequential D₂O/sucrose buoyant density ultracentrifugation fractionation into HDL-like particle and lipoprotein-depleted fractions from the indicated density ranges were separated by gradient (5–15%) reducing SDS-PAGE. A, SYPRO Ruby-stained gel of the indicated protein samples (5 μg) from homogenate and the indicated density ranges obtained from different atherosclerotic lesion tissue samples (n = 5). B, top panel, Western blot membrane of a duplicate-run gel as in A probed with anti-NO₂-Tyr¹⁶⁶-apoA-I mAb 4G11.2. B, bottom panel, overexposure of Western blot to show NO₂-Tyr¹⁶⁶-apoA-I in the HDL-like fraction. Monomers, dimers, and trimers (multimers) of apoA-I-immunoreactive NO₂-Tyr¹⁶⁶-apoA-I bands are indicated by arrowheads. Molecular mass markers are indicated.