POLYCLONAL ANTIBODY DIRECTED AGAINST HOMOCYSTEINE THIOLACTONE MODIFIED LDL

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Homocysteinemia is presumed to be responsible for the development of atherosclerosis, however, the precise etiology is unclear. We examined the possibility that the homocystamide-LDL adduct, a product of the reaction between homocysteine thiolactone and apo-B lysyl residues, was immunogenic. New Zealand white rabbits were immunized with this adduct at 6-week intervals. Antibody titers (ca. 100,000) were determined in antisera collected following the third immunization using solid-phase ELISA techniques. In competition-based ELISAs, homocysteine thiolactone-treated LDL competed for binding with the antiserum, as the 50% inhibitory concentration was approximately 10 micrograms/ml. Neither homocysteine, homocystine, nor Cu(2+)-oxidized LDL competed for binding. LDL in which lysyl residues were derivatized by acetylation or methylation were not recognized by the antiserum. All lipoprotein fractions from homocysteine thiolactone-treated plasma competed for binding to the antiserum. We conclude that homocysteine thiolactone-modified LDL is highly immunogenic and specific for homocystamide-lysyl adducts. This antibody has the potential to be used as a diagnostic tool for measuring plasma homocysteine and for delineating the role of homocysteinemia in vascular disease.