Immunogenicity of malondialdehyde-modified low density lipoproteins. Studies with monoclonal antibodies.
Malondialdehyde (MDA)-modified low density lipoprotein (LDL) can stimulate the accumulation of cholesteryl esters in cultured macrophages through its interaction with specific scavenger receptors. It has been speculated that such interaction occurs in vivo thus contributing to the formation of foam cells within atherosclerotic lesions. This report describes the development of new tools in the form of a specific assay for MDA-LDL to investigate this hypothesis. We have immunized BALB/c mice with malondialdehyde mouse low density lipoproteins and antibodies against malondialdehyde human low density lipoproteins were generated. Monoclonal antibodies were produced using hybridoma techniques and one particular clone (EB 7-3) was expanded for further studies. The immunoreactivity of several antigens was tested using antibody EB 7-3 in an enzyme-linked immunosorbent assay (ELISA). In a typical assay malondialdehyde human LDL (with at least 40% of lysines modified) was coated (2 micrograms/ml, 100 microliter) in 96-well microtiter plates. Antibody plus one of several antigens were then added and the interaction between the antibody and coated antigen was measured using alkaline phosphatase-conjugated affinity purified goat anti-mouse immunoglobulin. The binding of antibody EB 7-3 to wells coated with malondialdehyde-LDL was competitively inhibited by malondialdehyde-LDL added in solution, with half maximal inhibition occurring at 150 +/- 80 ng/ml. In addition, the ability of malondialdehyde-LDL to inhibit this interaction was proportional to the degree of modification: the more lysines were modified the more did malondialdehyde-LDL inhibit the binding of antibody EB 7-3 to coated malondialdehyde-LDL.(ABSTRACT TRUNCATED AT 250 WORDS)
D000911:Antibodies, Monoclonal / Q000276:immunology*; D000918:Antibody Specificity; D001011:Aorta / Q000032:analysis; D001059:Apoproteins / Q000276:immunology; D001666:Binding Sites, Antibody; D001667:Binding, Competitive; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000276:immunology; D006801:Humans; D008077:Lipoproteins, LDL / Q000276:immunology*; D008314:Malonates / Q000276:immunology*; D008315:Malondialdehyde / Q000276:immunology*; D011487:Protein Conformation
D000911:Antibodies, Monoclonal; D001059:Apoproteins; D000939:Epitopes; D008077:Lipoproteins, LDL; D008314:Malonates; D008315:Malondialdehyde