Abstract
In this study we established a murine monoclonal antibody that recognizes oxidized LDL using homogenates of human atheromatous plaque as immunogen. This antibody, FOH1a/DLH3, reacted with oxidized LDL but did not with either native, acetylated or malondialdehyde-treated LDL on ELISA. The antibody cross-reacted with oxidized HDL, suggesting that particular sequences of the apoB are not essential for specificity of the antigen recognition by the antibody. Immunohistochemical analysis of thin paraffin-sections from human colonary arteries showed that the foam cells derived from macrophages in atherosclerotic lesion were stained conspicuously with this antibody. Other compositions in the lesion including cellular debris in necrotic cores, swollen collagen fibers and endothelial cells were moderately stained. The epitope of this antibody was characterized by a model antigen generating system using ferrous ion-induced peroxidation of lipids. When the total lipid fraction extracted from LDL was treated with the ferrous ion-induced peroxidation system, the reaction mixture was recognized by the antibody. Antigenic product(s) was produced from phosphaidylcholine (PC) but not from the other lipids in the ferrous ion-induced peroxidation system. To detect a complex of an antigenic product with a polypeptide, reaction mixture of PC oxidized in the presence of a synthetic peptide was added to a microtiter well precoated with the monoclonal antibody FOH1a/DLH3. After washing, the peptide remained in the well was detected with a rabbit antiserum against the peptide, while no reactivity was obtained when the peptide alone was added to the well. Binding of the complex of oxidized PC and the peptide to FOH1a/DLH3 was competed by oxidized PC that was genarated in the absence of any polypeptide. We conclude that oxidized phospholipid product(s) is the epitope of the monoclonal antibody which forms complexes with polypeptides, and that the antigenic materials are detected in foam cells in atherosclerotic lesion.
