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(Arteriosclerosis, Thrombosis, and Vascular Biology. 1995;15:704-713.)
© 1995 American Heart Association, Inc.

Articles

Generation, Characterization, and Histochemical Application of Monoclonal Antibodies Selectively Recognizing Oxidatively Modified ApoB-Containing Serum Lipoproteins

Astrid Hammer; Gerd Kager; Gottfried Dohr; Hans Rabl; Irmgard Ghassempur; Günther Jürgens

From the Institute for Medical Biochemistry (A.H., G.K., G.J.), the Institute for Histology and Embryology (G.D., I.G.), and the Department for Surgery (H.R.), Medical Faculty, Karl-Franzens Universität Graz, Graz, Austria.

Correspondence to Dr Günther Jürgens, Institute for Medical Biochemistry, Harrachgasse 21, A-8010 Graz, Austria.

Abstract To investigate either the role oxidized LDL plays in atherosclerosis or structural changes on the surface of oxidized LDL, monoclonal antibodies (mAbs) are an important tool. After immunizing mice with Cu²⁺-oxidized LDL (oxLDL) and fusion of splenocytes, hybridoma supernatants were screened and cloned. Two mAbs, OB/04 and OB/09 (IgG and IgM), were further characterized. In solid-phase fluorescence immunoassays and Western blot analysis both mAbs reacted with oxLDL, LDL oxidized by a free radical–generating azo compound, or oxVLDL but not with native LDL, acetylated LDL, oxHDL₃, azo-oxidized HDL₃, or HDL₃ modified with malondialdehyde (MDA). In competitive immunoassays with LDL modified by oxidized fatty acid–derived aldehydes, mAb OB/09 strongly reacted with MDA-LDL or MDA-VLDL and LDL modified with 4-hydroxyhexenal followed by 4-hydroxynonenal but not with 4-hydroxyoctenal or hepta-2,4-dienal. mAb OB/04 had a weak affinity for LDL after modification with these aldehydes except for MDA-LDL. LDL modified with arachidonic acid oxidation products (AAOPs) was also recognized by this mAb. However, albumin modified either by the aldehydes applied or by AAOPs did not react with either mAb. Thus, the data indicate that each of the mAbs recognizes a different epitope that is expressed only on apoB-containing lipoproteins upon oxidative modification. An immunostaining with mAb OB/04 was obtained in areas rich in macrophages and in connective tissue of a human atherosclerotic lesion.

Key Words: modified lipoproteins • apoB • lipid peroxidation • monoclonal antibodies • atherosclerosis

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